Home » Intelligent Design » The Ubiquitin System: Functional Complexity and Semiosis joined together.

The Ubiquitin System: Functional Complexity and Semiosis joined together.

Spread the love

This is a very complex subject, so as usual I will try to stick to the essentials to make things as clear as possible, while details can be dealt with in the discussion.

It is difficult to define exactly the role of the Ubiquitin System. It is usually considered mainly a pathway which regulates protein degradation, but in reality its functions are much wider than that.

In essence, the US is a complex biological system which targets many different types of proteins for different final fates.

The most common “fate” is degradation of the protein. In that sense, the Ubiquitin System works together with another extremely complex cellular system, the proteasome. In brief, the Ubiquitin System “marks” proteins for degradation, and the proteasome degrades them.

It seems simple. It is not.

Ubiquitination is essentially one of many Post-Translational modifications (PTMs): modifications of proteins after their synthesis by the ribosome (translation). But, while most PTMs use simpler biochemical groups that are usually added to the target protein (for example, acetylation), in ubiquitination a whole protein (ubiquitin) is used as a modifier of the target protein.

 

The tool: Ubiquitin

Ubiquitin is a small protein (76 AAs). Its name derives from the simple fact that it  is found in most tissues of eukaryotic organisms.

Here is its aminoacid sequence:

MQIFVKTLTGKTITLEVEPSDTIENVKAKIQDKEGIPPD

QQRLIFAGKQLEDGRTLSDYNIQKESTLHLVLRLRGG

Essentially, it has two important properties:

  1. As said, it is ubiquitous in eukaryotes
  2. It is also extremely conserved in eukaryotes

In mammals, ubiquitin is not present as a single gene. It is encoded by 4 different genes: UBB, a poliubiquitin (3 Ub sequences); UBC, a poliubiquitin (9 Ub sequences); UBA52, a mixed gene (1   Ub sequence + the ribosomal protein L40); and RPS27A, again a mixed gene (1 Ub sequence + the ribosomal protein S27A). However, the basic ubiquitin sequence is always the same in all those genes.

Its conservation is one of the highest in eukaryotes. The human sequence shows, in single celled eukaryotes:

Naegleria: 96% conservation;  Alveolata: 100% conservation;  Cellular slime molds: 99% conservation; Green algae: 100% conservation; Fungi: best hit 100% conservation (96% in yeast).

Ubiquitin and Ubiquitin like proteins (see later) are characterized by a special fold, called  β-grasp fold.

 

The semiosis: the ubiquitin code

The title of this OP makes explicit reference to semiosis. Let’s try to see why.

The simplest way to say it is: ubiquitin is a tag. The addition of ubiquitin to a substrate protein marks that protein for specific fates, the most common being degradation by the proteasome.

But not only that. See, for example, the following review:

Nonproteolytic Functions of Ubiquitin in Cell Signaling

Abstract:

The small protein ubiquitin is a central regulator of a cell’s life and death. Ubiquitin is best known for targeting protein destruction by the 26S proteasome. In the past few years, however, nonproteolytic functions of ubiquitin have been uncovered at a rapid pace. These functions include membrane trafficking, protein kinase activation, DNA repair, and chromatin dynamics. A common mechanism underlying these functions is that ubiquitin, or polyubiquitin chains, serves as a signal to recruit proteins harboring ubiquitin-binding domains, thereby bringing together ubiquitinated proteins and ubiquitin receptors to execute specific biological functions. Recent advances in understanding ubiquitination in protein kinase activation and DNA repair are discussed to illustrate the nonproteolytic functions of ubiquitin in cell signaling.

Another important aspect is that ubiquitin is not one tag, but rather a collection of different tags. IOWs, a tag based code.

See, for example, here:

The Ubiquitin Code in the Ubiquitin-Proteasome System and Autophagy

(Paywall).

Abstract:

The conjugation of the 76 amino acid protein ubiquitin to other proteins can alter the metabolic stability or non-proteolytic functions of the substrate. Once attached to a substrate (monoubiquitination), ubiquitin can itself be ubiquitinated on any of its seven lysine (Lys) residues or its N-terminal methionine (Met1). A single ubiquitin polymer may contain mixed linkages and/or two or more branches. In addition, ubiquitin can be conjugated with ubiquitin-like modifiers such as SUMO or small molecules such as phosphate. The diverse ways to assemble ubiquitin chains provide countless means to modulate biological processes. We overview here the complexity of the ubiquitin code, with an emphasis on the emerging role of linkage-specific degradation signals (degrons) in the ubiquitin-proteasome system (UPS) and the autophagy-lysosome system (hereafter autophagy).

A good review of the basics of the ubiquitin code can be found here:

The Ubiquitin Code 

(Paywall)

It is particularly relevant, from an ID point of view, to quote the starting paragraph of that paper:

When in 1532 Spanish conquistadores set foot on the Inca Empire, they found a highly organized society that did not utilize a system of writing. Instead, the Incas recorded tax payments or mythology with quipus, devices in which pieces of thread were connected through specific knots. Although the quipus have not been fully deciphered, it is thought that the knots between threads encode most of the quipus’ content. Intriguingly, cells use a regulatory mechanism—ubiquitylation—that is reminiscent of quipus: During this reaction, proteins are modified with polymeric chains in which the linkage between ubiquitin molecules encodes information about the substrate’s fate in the cell.

Now, ubiquitin is usually linked to the target protein in chains. The first ubiquitin molecule is covalently bound through its C-terminal carboxylate group to a particular lysine, cysteine, serine, threonine or N-terminus of the target protein.

Then, additional ubiquitins are added to form a chain, and the C-terminus of the new ubiquitin is linked to one of seven lysine residues or the first methionine residue on the previously added ubiquitin.

IOWs, each ubiquitin molecule has seven lysine residues:

K6, K11, K27, K29, K33, K48, K63

And one N terminal methionine residue:

M1

And a new ubiquitin molecule can be added at each of those 8 sites in the previous ubiquitin molecule. IOWs, those 8 sites in the molecule are configurable switches that can be used to build ubiquitin chains.

Her are the 8 sites, in red, in the ubiquitin molecule:

MQIFVKTLTGKTITLEVEPSDTIENVKAKIQDKEGIPPD

QQRLIFAGKQLEDGRTLSDYNIQKESTLHLVLRLRGG

Fig 1 shows two ubiquitin molecules joined at K48.

Fig 1 A cartoon representation of a lysine 48-linked diubiquitin molecule. The two ubiquitin chains are shown as green cartoons with each chain labelled. The components of the linkage are indicated and shown as orange sticks. By Rogerdodd (Own work) [CC BY-SA 3.0 (https://creativecommons.org/licenses/by-sa/3.0)], via Wikimedia Commons

The simplest type of chain is homogeneous (IOWs, ubiquitins are linked always at the same site). But many types of mixed and branched chains can also be found.

Let’s start with the most common situation: a poli-ubiquitination of (at least) 4 ubiqutins, linearly linked at K48. This is the common signal for proteasome degradation.

By the way, the 26S proteasome is another molecular machine of incredible complexity, made of more than 30 different proteins. However, its structure and function are not the object of this OP, and therefore I will not deal with them here.

The ubiquitin code is not completely understood, at present, but a few aspects have been well elucidated. Table 1 sums up the most important and well known modes:

Code

Meaning

Polyubiquitination (4 or more) with links at K48 or at K11 Proteasomal degradation
Monoubiqutination (single or multiple) Protein interactions, membrane trafficking, endocytosis
Polyubiquitination with links at K63 Endocytic trafficking, inflammation, translation, DNA repair.
Polyubiquitination with links at K63 (other links) Autophagic degradation of protein substrates
Polyubiquitination with links at K27, K29, K33 Non proteolytic processes
Rarer chain types (K6, K11) Under investigation

 

However, this is only a very partial approach. A recent bioinformatics paper:

An Interaction Landscape of Ubiquitin Signaling

(Paywall)

Has attempted for the first time a systematic approach to deciphering the whole code, using synthetic diubiquitins (all 8 possible variants) to identify the different interactors with those signals, and they identified, with two different methodologies,  111 and 53 selective interactors for linear polyUb chains, respectively. 46 of those interactors were identified by both methodologies.

The translation

But what “translates” the complex ubiquitin code, allowing ubiquinated proteins to met the right specific destiny? Again, we can refer to the diubiquitin paper quoted above.

How do cells decode this ubiquitin code into proper cellular responses? Recent studies have indicated that members of a protein family, ubiquitin-binding proteins (UBPs), mediate the recognition of ubiquitinated substrates. UBPs contain at least one of 20 ubiquitin-binding domains (UBDs) functioning as a signal adaptor to transmit the signal from ubiquitinated substrates to downstream effectors

But what are those “interactors” identified by the paper (at least 46 of them)? They are, indeed, complex proteins which recognize specific configurations of the “tag” (the ubiquitin chain), and link the tagged (ubiquinated) protein to other effector proteins which implement its final fate, or anyway contribute in deffrent forms to that final outcome.

 

The basic control of the procedure: the complexity of the ubiquitination process.

So, we have seen that ubiquitin chains work as tags, and that their coded signals are translated by specific interactors, so that the target protein may be linked to its final destiny, or contribute to the desired outcome. But we must still address one question: how is the ubiquitination of the different target proteins implemented? IOWs, what is the procedure that “writes” the specific codes associated to specific target proteins?

This is indeed the first step in the whole process. But it is also the most complex, and that’s why I have left it for the final part of the discussion.

Indeed, the ubiquitination process needs to realize the following aims:

  1. Identify the specific protein to be ubiquitinated
  2. Recognize the specific context in which that protein needs to be ubiquitinated
  3. Mark the target protein with the correct tag for the required fate or outcome

We have already seen that the ubiquitin system is involved in practically all different cellular paths and activities, and therefore we can expect that the implementation of the above functions must be a very complex thing.

And it is.

Now, we can certainly imagine that there are many different layers of regulation that may contribute to the general control of the procedure, specifically epigenetic levels, which are at present poorly understood. But there is one level that we can more easily explore and understand, and it is , as usual, the functional complexity of the proteins involved.

And, even at a first gross analysis, it is really easy to see that the functional complexity implied by this process is mind blowing.

Why? It is more than enough to consider the huge number of different proteins involved. Let’s see.

The ubiquitination process is well studied. It can be divided into three phases, each of which is implemented by a different kind of protein. The three steps, and the three kinds of proteins that implement them, take the name of E1, E2 and E3.

 

Fig. 2 Schematic diagram of the ubiquitylation system. Created by Roger B. Dodd: Rogerdodd at the English language Wikipedia [GFDL (http://www.gnu.org/copyleft/fdl.html) or CC-BY-SA-3.0 (http://creativecommons.org/licenses/by-sa/3.0/)], via Wikimedia Commons

 The E1 step of ubiquitination.

This is the first thing that happens, and it is also the simplest.

E1 is the process of activation of ubiquitin, and the E1 proteins is called E1 ubiquitin-activating enzyme. To put it simply, this enzyme “activates” the ubiquitin molecule in an ATP dependent process, preparing it for the following phases and attaching it to its active site cysteine residue. It is not really so simple, but for our purposes that can be enough.

This is a rather straightforward enzymatic reaction. In humans there are essentially two forms of E1 enzymes, UBA1 and UBA6, each of them about 1000 AAs long, and partially related at sequence level (42%).

 

The E2 step of ubiquitination.

The second step is ubiquitin conjugation. The activated ubiquitin is transferred from the E1 enzyme to the ubiquitin-conjugating enzyme, or E2 enzyme, where it is attached to a cysteine residue.

This apparently simple “transfer” is indeed a complex intermediate phase. Humans have about 40 different E2 molecules. The following paper:

E2 enzymes: more than just middle men

details some of the functional complexity existing at this level.

Abstract:

Ubiquitin-conjugating enzymes (E2s) are the central players in the trio of enzymes responsible for the attachment of ubiquitin (Ub) to cellular proteins. Humans have ∼40 E2s that are involved in the transfer of Ub or Ub-like (Ubl) proteins (e.g., SUMO and NEDD8). Although the majority of E2s are only twice the size of Ub, this remarkable family of enzymes performs a variety of functional roles. In this review, we summarize common functional and structural features that define unifying themes among E2s and highlight emerging concepts in the mechanism and regulation of E2s.

However, I will not go into details about these aspects, because we have better things to do: we still have to discuss the E3 phase!

 

The E3 step of ubiquitination.

This is the last phase of ubiquitination, where the ubiquitin tag is finally transferred to the target protein, as initial mono-ubiquitination, or to build an ubiquitin chain by following ubiqutination events. The proteins which implement this final passage are call E3 ubiquitin ligases. Here is the definition from Wikipedia:

A ubiquitin ligase (also called an E3 ubiquitin ligase) is a protein that recruits an E2 ubiquitin-conjugating enzyme that has been loaded with ubiquitin, recognizes a protein substrate, and assists or directly catalyzes the transfer of ubiquitin from the E2 to the protein substrate.

It is rather obvious that the role of the E3 protein is very important and delicate. Indeed it:

  1. Recognizes and links the E2-ubiquitin complex
  2. Recognizes and links some specific target protein
  3. Builds the appropriate tag for that protein (Monoubiquitination, mulptiple monoubiquitination, or poliubiquitination with the appropriate type of ubiquitin chain).
  4. And it does all those things at the right moment, in the right context, and for the right protein.

IOWs, the E3 protein writes the coded tag. It is, by all means, the central actor in our complex story.

So, here comes the really important point: how many different E3 ubiquitin ligases do we find in eukaryotic organisms? And the simple answer is: quite a lot!

Humans are supposed to have more than 600 different E3 ubiquitin ligases!

So, the human machinery for ubiquitination is about:

2 E1 proteins  –  40 E2 proteins – >600 E3 proteins

A real cascade of complexity!

OK, but even if we look at single celled eukaryotes we can already find an amazing level of complexity. In yeast, for example, we have:

1 or 2 E1 proteins  –  11 E2 proteins – 60-100 E3 proteins

See here:

The Ubiquitin–Proteasome System of Saccharomyces cerevisiae

Now, a very important point. Those 600+ E3 proteins that we find in humans are really different proteins. Of course, they have something in common: a specific domain.

From that point of view, they can be roughly classified in three groups according to the specific E3 domain:

  1. RING group: the RING finger domain ((Really Interesting New Gene) is a short domain of zinc finger type, usually 40 to 60 amino acids. This is the biggest group of E3s (about 600)
  2. HECT domain (homologous to the E6AP carboxyl terminus): this is a bigger domain (about 350 AAs). Located at the C terminus of the protein. It has a specific ligase activity, different from the RING   In humans we have approximately 30 proteins of this type.
  3. RBR domain (ring between ring fingers): this is a common domain (about 150 AAs) where two RING fingers are separated by a region called IBR, a cysteine-rich zinc finger. Only a subset of these proteins are E3 ligases, in humans we have about 12 of them.

See also here.

OK, so these proteins have one of these three domains in common, usually the RING domain. The function of the domain is specifically to interact with the E2-ubiquitin complex to implement the ligase activity. But the domain is only a part of the molecule, indeed a small part of it. E3 ligases are usually big proteins (hundreds, and up to thousands of AAs). Each of these proteins has a very specific non domain sequence, which is probably responsible for the most important part of the function: the recognition of the specific proteins that each E3 ligase processes.

This is a huge complexity, in terms of functional information at sequence level.

Our map of the ubiquinating system in humans could now be summarized as follows:

2 E1 proteins  –  40 E2 proteins – 600+ E3 proteins + thousands of specific substrates

IOWs, each of hundreds of different complex proteins recognizes its specific substrates, and marks them with a shared symbolic code based on uniquitin and its many possible chains. And the result of that process is that proteins are destined to degradation by the proteasome or other mechanisms, and that protein interactions and protein signaling are regulated and made possible, and that practically all cellular functions are allowed to flow correctly and smoothly.

Finally, here are two further compoments of the ubuquitination system, which I will barely mention, to avoid making this OP too long.

Ubiquitin like proteins (Ubl):

A number of ubiquitin like proteins add to the complexity of the system. Here is the abstract from a review:

The eukaryotic ubiquitin family encompasses nearly 20 proteins that are involved in the posttranslational modification of various macromolecules. The ubiquitin-like proteins (UBLs) that are part of this family adopt the β-grasp fold that is characteristic of its founding member ubiquitin (Ub). Although structurally related, UBLs regulate a strikingly diverse set of cellular processes, including nuclear transport, proteolysis, translation, autophagy, and antiviral pathways. New UBL substrates continue to be identified and further expand the functional diversity of UBL pathways in cellular homeostasis and physiology. Here, we review recent findings on such novel substrates, mechanisms, and functions of UBLs.

These proteins include SUMO, Nedd8, ISB15, and many others.

Deubiquitinating enzymes (DUBs):

The process of ubiquitination, complex as it already is, is additionally regulated by these enzymes which can cleave ubiquitin from proteins and other molecules. Doing so, they can reverse the effects of ubiquitination, creating a delicately balanced regulatory network. In humans there are nearly 100 DUB genes, which can be classified into two main classes: cysteine proteases and metalloproteases.

 

By the way, here is a beautiful animation of the basic working of the ubiquitin-proteasome system in degrading damaged proteins:

 

 

A summary:

So, let’s try a final graphic summary of the whole ubiquitin system in humans:

Fig 3 A graphic summary of the Ubiquitin System

 

Evolution of the Ubiquitin system?

The Ubiqutin system is essentially an eukaryotic tool. Of course, distant precursors for some of the main components have been “found” in prokaryotes. Here is the abstract from a paper that sums up what is known about the prokaryotic “origins” of the system:

Structure and evolution of ubiquitin and ubiquitin-related domains.

(Paywall)

Abstract:

Since its discovery over three decades ago, it has become abundantly clear that the ubiquitin (Ub) system is a quintessential feature of all aspects of eukaryotic biology. At the heart of the system lies the conjugation and deconjugation of Ub and Ub-like (Ubls) proteins to proteins or lipids drastically altering the biochemistry of the targeted molecules. In particular, it represents the primary mechanism by which protein stability is regulated in eukaryotes. Ub/Ubls are typified by the β-grasp fold (β-GF) that has additionally been recruited for a strikingly diverse range of biochemical functions. These include catalytic roles (e.g., NUDIX phosphohydrolases), scaffolding of iron-sulfur clusters, binding of RNA and other biomolecules such as co-factors, sulfur transfer in biosynthesis of diverse metabolites, and as mediators of key protein-protein interactions in practically every conceivable cellular context. In this chapter, we present a synthetic overview of the structure, evolution, and natural classification of Ub, Ubls, and other members of the β-GF. The β-GF appears to have differentiated into at least seven clades by the time of the last universal common ancestor of all extant organisms, encompassing much of the structural diversity observed in extant versions. The β-GF appears to have first emerged in the context of translation-related RNA-interactions and subsequently exploded to occupy various functional niches. Most biochemical diversification of the fold occurred in prokaryotes, with the eukaryotic phase of its evolution mainly marked by the expansion of the Ubl clade of the β-GF. Consequently, at least 70 distinct Ubl families are distributed across eukaryotes, of which nearly 20 families were already present in the eukaryotic common ancestor. These included multiple protein and one lipid conjugated forms and versions that functions as adapter domains in multimodule polypeptides. The early diversification of the Ubl families in eukaryotes played a major role in the emergence of characteristic eukaryotic cellular substructures and systems pertaining to nucleo-cytoplasmic compartmentalization, vesicular trafficking, lysosomal targeting, protein processing in the endoplasmic reticulum, and chromatin dynamics. Recent results from comparative genomics indicate that precursors of the eukaryotic Ub-system were already present in prokaryotes. The most basic versions are those combining an Ubl and an E1-like enzyme involved in metabolic pathways related to metallopterin, thiamine, cysteine, siderophore and perhaps modified base biosynthesis. Some of these versions also appear to have given rise to simple protein-tagging systems such as Sampylation in archaea and Urmylation in eukaryotes. However, other prokaryotic systems with Ubls of the YukD and other families, including one very close to Ub itself, developed additional elements that more closely resemble the eukaryotic state in possessing an E2, a RING-type E3, or both of these components. Additionally, prokaryotes have evolved conjugation systems that are independent of Ub ligases, such as the Pup system.

 

As usual, we are dealing here with distant similarities, but there is no doubt that the ubiquitin system as we know it appears in eukaryotes.

But what about its evolutionary history in eukaryotes?

We have already mentioned the extremely high conservation of ubiquitin itself.

UBA1, the main E1 enzyme, is rather well conserved from fungi to humans: 60% identity, 1282 bits, 1.21 bits per aminoacid (baa).

E2s are small enzymes, extremely conserved from fungi to humans: 86% identity, for example, for UB2D2, a 147 AAs molecule.

E3s, of course, are the most interesting issue. This big family of proteins behaves in different ways, consistently with its highly specific functions.

It is difficult to build a complete list of E3 proteins. I have downloaded from Uniprot a list of reviewed human proteins including “E3 ubiquitun ligase” in their name: a total of 223 proteins.

The mean evolutionary behavior of this group in metazoa is rather different from protein to protein. However, as a group these proteins exhibit an information jump in vertebrates which is significantly higher than the jump in all other proteins:

 

Fig. 4 Boxplots of the distribution of human conserved information jump from pre-vertebrates to vertebrates in 223 E3 ligase proteins and in all other human proteins. The difference is highly significant.

 

As we already know, this is evidence that this class of proteins is highly engineered in the transition to vertebrates. That is consistent with the need to finely regulate many cellular processes, most of which are certainly highly specific for different groups of organisms.

The highest vertebrate jump, in terms of bits per aminoacid, is shown in my group by the E3 ligase TRIM62. also known as DEAR1 (Q9BVG3), a 475 AAs long protein almost absent in pre-vertebrates (best hit 129 bits, 0.27 baa in Branchiostoma belcheri) and which flaunts an amazing jump of 1.433684 baa in cartilaginous fish (810 bits, 1.705263 baa).

But what is this protein? It is a master regulator tumor suppressor gene, implied in immunity, inflammation, tumor genesis.

See here:

TRIM Protein-Mediated Regulation of Inflammatory and Innate Immune Signaling and Its Association with Antiretroviral Activity

and here:

DEAR1 is a Chromosome 1p35 Tumor Suppressor and Master Regulator of TGFβ-Driven Epithelial-Mesenchymal Transition

This is just to show what a single E3 ligase can be involved in!

An opposite example, from the point of view of evolutionary history, is SIAH1, an E3 ligase implied in proteosomal degradation of proteins. It is a 282 AAs long protein, which already exhibits 1.787234 baa (504 bits) of homology in deuterostomes, indeed already 1.719858 baa in cnidaria. However, in fungi the best hit is only 50.8 bits (0.18 baa). So, this is a protein whose engineering takes place at the start of metazoa, and which exhibits only a minor further jump in vertebrates (0.29 baa), which brings the protein practically to its human form already in cartilaginous fish (280 identities out of 282, 99%). Practically a record.

So, we can see that E3 ligases are a good example of a class of proteins which perform different specific functions, and therefore exhibit different evolutionary histories: some, like TRIM62, are vertebrate quasi-novelties, others, like SIAH1, are metazoan quasi-novelties. And, of course, there are other behaviours, like for example BRCA1, Breast cancer type 1 susceptibility protein, a protein 1863 AAs long which only in mammals acquires part of its final sequence configuration in humans.

The following figure shows the evolutionary history of the three proteins mentioned above.

 

Fig. 5 Evolutionary history in metazoa of three E3 ligases (human conserved functional information)

 

An interesting example: NF-kB signaling

I will discuss briefly an example of how the Ubiquitin system interacts with some specific and complex final effector system. One of the best models for that is the NF-kB signaling.

NK-kB is a transcription factor family that is the final effector of a complex signaling pathway. I will rely mainly on the following recent free paper:

The Ubiquitination of NF-κB Subunits in the Control of Transcription

Here is the abstract:

Nuclear factor (NF)-κB has evolved as a latent, inducible family of transcription factors fundamental in the control of the inflammatory response. The transcription of hundreds of genes involved in inflammation and immune homeostasis require NF-κB, necessitating the need for its strict control. The inducible ubiquitination and proteasomal degradation of the cytoplasmic inhibitor of κB (IκB) proteins promotes the nuclear translocation and transcriptional activity of NF-κB. More recently, an additional role for ubiquitination in the regulation of NF-κB activity has been identified. In this case, the ubiquitination and degradation of the NF-κB subunits themselves plays a critical role in the termination of NF-κB activity and the associated transcriptional response. While there is still much to discover, a number of NF-κB ubiquitin ligases and deubiquitinases have now been identified which coordinate to regulate the NF-κB transcriptional response. This review will focus the regulation of NF-κB subunits by ubiquitination, the key regulatory components and their impact on NF-κB directed transcription.

 

The following figure sums up the main features of the canonical activation pathway:

 

Fig. 6 A simple summary of the main steps in the canonical activayion pathway of NF-kB

 

Here the NF-κB TF is essentially the heterodimer RelA – p50. Before activation, the NF-κB (RelA – p50) dimer is kept in an inactive state and remains in the cytoplasm because it is linked to the IkB alpha protein, an inhibitor of its function.

Activation is mediated by a signal-receptor interaction, which starts the whole pathway. A lot of different signals can do that, adding to the complexity, but we will not discuss this part here.

As a consequence of receptor activation, another protein complex, IκB kinase (IKK), accomplishes the Phosphorylation of IκBα at serines 32 and 36. This is the signal for the ubiquitination of the IkB alpha inhibitor.

This ubiqutination targets IkB alpha for proteosomal degradation. But how is it achieved?

Well, things are not so simple. A whole protein complex is necessary, a complex which implements many different ubiquitinations in different contexts, including this one.

The complex is made by 3 basic proteins:

  • Cul1 (a scaffold protein, 776 AAs)
  • SKP1 (an adaptor protein, 163 AAs)
  • Rbx1 (a RING finger protein with E3 ligase activity, 108 AAs)

Plus:

  • An F-box protein (FBP) which changes in the different context, and confers specificity.

In our context, the F box protein is called beta TRC (605 AAs).

 

Fig. 7 A simple diagram of the SKP1 – beta TRC complex

 

Once the IkB alpha inhibitor is ubiquinated and degraded in the proteasome, the NF-κB dimer is free to translocate to the nucleus, and implement its function as a transcription factor (which is another complex issue, that we will not discuss).

OK, this is only the canonical activation of the pathway.

In the non canonical pathway (not shown in the figure) a different set of signals, receptors and activators acts on a different NF-κB dimer (RelB – p100). This dimer is not linked to any inhibitor, but is itself inactive in the cytoplasm. As a result of the signal, p100 is phosphorylated at serines 866 and 870. Again, this is the signal for ubiquitination.

This ubiquitination is performed by the same complex described above, but the result is different. P100 is only partially degraded in the proteasome, and is transformed into a smaller protein, p52, which remains linked to RelB. The RelB – p52 dimer is now an active NF-κB Transcription Factor, and it can relocate to the nucleus and act there.

But that’s not all.

  • You may remember that RelA (also called p 65) is one of the two components of NF-kB TF in the canonical pathway (the other being p 50). Well, RelA is heavily controlled by ubiquitination after it binds DNA in the nucleus to implement its TF activity. Ubiquitination (a very complex form of it) helps detachment of the TF from DNA, and its controlled degradation, avoiding sustained expression of NF-κB-dependent genes. For more details, see section 4 in the above quoted paper: “Ubiquitination of NF-κB”.
  • The activation of IKK in both the canonical and non canonical pathway after signal – receptor interaction is not so simple as depicted in Fig. 6. For more details, look at Fig. 1 in this paper: Ubiquitin Signaling in the NF-κB Pathway. You can see that, in the canonical pathway, the activation of IKK is mediated by many proteins, including TRAF2, TRAF6, TAK1, NEMO.
  • TRAF2 is a key regulator on many signaling pathways, including NF-kB. It is an E3 ubiquitin ligase. From Uniprot:  “Has E3 ubiquitin-protein ligase activity and promotes ‘Lys-63’-linked ubiquitination of target proteins, such as BIRC3, RIPK1 and TICAM1. Is an essential constituent of several E3 ubiquitin-protein ligase complexes, where it promotes the ubiquitination of target proteins by bringing them into contact with other E3 ubiquitin ligases.”
  • The same is true of TRAF6.
  • NEMO (NF-kappa-B essential modulator ) is also a key regulator. It is not an ubiquinating enzyme, but it is rather heavily regulated by ubiquitination. From Uniprot: “Regulatory subunit of the IKK core complex which phosphorylates inhibitors of NF-kappa-B thus leading to the dissociation of the inhibitor/NF-kappa-B complex and ultimately the degradation of the inhibitor. Its binding to scaffolding polyubiquitin seems to play a role in IKK activation by multiple signaling receptor pathways. However, the specific type of polyubiquitin recognized upon cell stimulation (either ‘Lys-63’-linked or linear polyubiquitin) and its functional importance is reported conflictingly.”
  • In the non canonical pathway, the activation of IKK alpha after signal – receptor interaction is mediated by other proteins, in particular one protein called NIK (see again Fig. 1 quoted above). Well, NIK is regulated by two different types of E3 ligases, with two different types of polyubiquitination:
    • cIAP E3 ligase inactivates it by constant degradation using a K48 chain
    • ZFP91 E3 ligase stabilizes it using a K63 chain

See here:

Non-canonical NF-κB signaling pathway.

In particular, Fig. 3

These are only some of the ways the ubiquitin system interacts with the very complex NF-kB signaling system. I hope that’s enough to show how two completely different and complex biological systems manage to cooperate by intricate multiple connections, and how the ubiquitin system can intervene at all levels of another process. What is true for the NF-kB signaling pathway is equally true for a lot of other biological systems, indeed for almost all basic cellular processes.

But this OP is already too long, and I have to stop here.

As usual, I want to close with a brief summary of the main points:

  1. The Ubiquitin system is a very important regulation network that shows two different signatures of design: amazing complexity and an articulated semiotic structure.
  2. The complexity is obvious at all levels of the network, but is especially amazing at the level of the hundreds of E3 ligases, that can recognize thousands of different substrates in different contexts.
  3. The semiosis is obvious in the Ubiquitin Code, a symbolic code of different ubiquitin configurations which serve as specific “tags” that point to different outcomes.
  4. The code is universally implemented and shared in eukaryotes, and allows control on almost all most important cellular processes.
  5. The code is written by the hundreds of E3 ligases. It is read by the many interactors with ubiquitin-binding domains (UBDs).
  6. The final outcome is of different types, including degradation, endocytosis, protein signaling, and so on.
  7. The interaction of the Ubiquitin System with other complex cellular pathways, like signaling pathways, is extremely complex and various, and happens at many different levels and by many different interacting proteins for each single pathway.

PS:

Thanks to DATCG for pointing to this video in three parts by Dr. Raymond Deshaies, was Professor of Biology at the California Institute of Technology and an Investigator of the Howard Hughes Medical Institute. On iBiology Youtube page:

A primer on the ubiquitin-proteasome system

 

Cullin-RING ubiquitin ligases: structure, structure, mechanism, and regulation

 

Targeting the ubiquitin-proteasome system in cancer

950 Responses to The Ubiquitin System: Functional Complexity and Semiosis joined together.

  1. Oooooooooooooohhhh more fun 🙂

    I need time to read later and look forward to another good discussion.

    I made a quick comment on Spliceosome post by you and scientist Arthur Hunt’s disappearance from the discussion a moment ago.

    Scientist Arthur Hunt: https://www.researchgate.net/profile/Arthur_Hunt

    Thought he would contribute to a good debate and discussion points in opposition to you Gpuccio on the Spliceosome, it’s evolution, Design or not.

    But he seems to have disappeared without any rebuttal to your detailed responses…

    Spliceosome – Defy non-Design Explanations

    .

  2. DATCG:

    Thank you for opening the discussion! 🙂

    I understand that the OP is rather long and touches a lot of stuff. I hope that you and others who are interested in this kind of approach can give a very good contribution to a discussion about this fascinating issue.

    Of course, UB is specially invited to comment: he is the master of the semiosis arguments, and I am sure that he will be interested in the things discussed here. 🙂

    Like you, I have been a little disappointed of the apparent disappearance of Arthur Hunt from the spliceosome discussion. He could certainly have given a great contribution. However, not knowing his reasons, I cannot certainly judge.

  3. Looking forward to it. Thanks again for your interesting post. And yes, very much like this approach. Hope others join in.

    Nothing like peering through signals, interpretations and post-translations and meta layers of information code on top of code.

    Conditional processing is a feature here I think as well as specificity.

    Agree with you. Not judging Arthur. If my speculation is correct, then I’m defending him. And hope he may show up in the future.

    But onward 🙂

    To the Ubiquitin Code.

    And hope it’s OK to add a video overview in 3 parts for those who might like to see some animation along with explanations.

    By Dr. Raymond Deshaies, was Professor of Biology at the California Institute of Technology and an Investigator of the Howard Hughes Medical Institute. On iBiology Youtube page.

    A primer on the ubiquitin-proteasome system…
    https://www.youtube.com/watch?v=ILdEOXCfgUc

    There’s more of course, but thought he added some good insights.

  4. DATCG:

    Thank you for the link to the video. I have added it (alll three parts) at the end of the OP. I have not yet had the time to see them all, but I will, and I will comment on the points I find most interesting! 🙂

    Yes, codes are of supreme importance in cellular life, and they do scream design.

    You say:

    “Conditional processing is a feature here I think as well as specificity.”

    That’s exactly the point! This ubiquitin code is a perfect implementation of conditional processing, an equivalent of complex “if – else” structures.

    Using an universal tag with internal configurable switches is really a brilliant solution for that.

    And it also reminds me of Lego construction toys! 🙂

  5. DATCG:

    At timepoint 2:39 of the first video there is already a very important point which deserves discussion: why proteins, and especially regulator proteins, need to be unstable so that adjustments in their steady state can be achieved quickly.

    It means that biological systems invest a lot of resources to achieve flexibility and quick (and very intelligent) adaptation to changing conditions.

    I suppose that can be achieved because one of life’s features is to depend so critically on far from equilibrium processes.

    Food for thought.

  6. DATCG:

    At timepoint 23:25 of the first video there is a brief explanation of the role of phosphorylation in regulating ubiquitination.

    I have given an example of that in the OP, when I speak of the regulation of NF-kB, where phosphorylation provides the ubiquitination activating signal, both in the canonical and non canonical pathway, acting on the substrate. As shown in the video, phosphorylarion can act both on the substrate and on the E3 ligase, and finally it can also have an inhibitory role.

    This is fascinating, because we have a simpler semiotic system (phosphorylation) which controls a much more complex semiotic system (ubiquitination) to regulate the working of an effector system (NF-kB signaling) which is, itself, highly semiotic.

  7. DATCG:

    The second video is very interesting too. Starting at 1:33, the structure of Cal1 E3 ligases (of which the one shown at Fig. 7 in the OP is a very good example) seems to be a wonderful example of modular design.

    70 F-boxes.

    40 SOCS boxes.

    250 different assemblies.

    Really amazing!

  8. gpuccio,

    Excellent OP, as usual. Thanks.

    Also very interesting discussion with DATCG, who writes insightful comments and pointed to an interesting biology video series on the same fascinating topic of the OP.

    Here’s a link to a full text PDF copy of one of the papers you referenced in the OP:

    https://www.researchgate.net/profile/Lakshminarayan_Iyer/publication/221848141_Structure_and_Evolution_of_Ubiquitin_and_Ubiquitin-Related_Domains/links/57436c4a08aea45ee84d1061/Structure-and-Evolution-of-Ubiquitin-and-Ubiquitin-Related-Domains.pdf

    Note the 21 citations to the given paper.

    PS, my current activities are keeping me from commenting lately. Perhaps that’s pleasant news to some folks out there who were annoyed by my posts and even kept statistical track of their frequency and volume, but never dared to address them seriously. What else is new?

    The project I’ve been working on has proceeded to another phase that requires more attention to difficult for me issues.

  9. Dionisio:

    That will perhaps be pleasant news to some folks, but believe me, you will be missed by many others, including me.

    I am happy that your project is proceeding. However, I hope that you can still find some time to be with us, maybe with less frequency and volume, but with your usual ingenuity! 🙂

  10. I think that the title of the following 2016 paper about the ubiquitin system could be of some interest:

    Design Principles Involving Protein Disorder Facilitate Specific Substrate Selection and Degradation by the Ubiquitin-Proteasome System”

    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4807260/

    (Emphasis mine. Public access.)

    The Abstract is very interesting, too:

    The ubiquitin-proteasome system (UPS) regulates diverse cellular pathways by the timely removal (or processing) of proteins. Here we review the role of structural disorder and conformational flexibility in the different aspects of degradation. First, we discuss post-translational modifications within disordered regions that regulate E3 ligase localization, conformation, and enzymatic activity, and also the role of flexible linkers in mediating ubiquitin transfer and reaction processivity. Next we review well studied substrates and discuss that substrate elements (degrons) recognized by E3 ligases are highly disordered: short linear motifs recognized by many E3s constitute an important class of degrons, and these are almost always present in disordered regions. Substrate lysines targeted for ubiquitination are also often located in neighboring regions of the E3 docking motifs and are therefore part of the disordered segment. Finally, biochemical experiments and predictions show that initiation of degradation at the 26S proteasome requires a partially unfolded region to facilitate substrate entry into the proteasomal core.

    (Emphasis mine)

    The theme of intrinsically disordered regions in proteins is becoming ever more relevant.

    See also here:

    “Classification of Intrinsically Disordered Regions and Proteins”

    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4095912/

    Public access.

    I quote just the title of the first section:

    1.1. Uncharacterized Protein Segments Are a Source of Functional Novelty

  11. This is a funny video. She does not use Lego, but something like that…

    https://www.youtube.com/watch?v=miZYmuDKO2s

    About histone ubiquitination, which I have not touched in the OP, here is an interesting paper:

    “Histone Ubiquitination and Deubiquitination in Transcription, DNA Damage Response, and Cancer”

    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3355875/

    (Public access)

    Both monoubiquitination and polyubiquitination seem to happen at histone level.

  12. About branched ubiquitin chains and, again, NF-kB regulation:

    The K48-K63 Branched Ubiquitin Chain Regulates NF-kB Signaling.

    http://www.cell.com/molecular-.....16)30563-9

    (Public access)

    Abstract:

    Polyubiquitin chains of different topologies regulate diverse cellular processes. K48- and K63-linked chains, the two most abundant chain types, regulate proteolytic and signaling pathways, respectively. Although recent studies reported important roles for heterogeneous chains, the functions of branched ubiquitin chains remain unclear. Here, we show that the ubiquitin chain branched at K48 and K63 regulates nuclear factor ?B (NF-?B) signaling. A mass-spectrometry-based quantification strategy revealed that K48-K63 branched ubiquitin linkages are abundant in cells. In response to interleukin-1?, the E3 ubiquitin ligase HUWE1 generates K48 branches on K63 chains formed by TRAF6, yielding K48-K63 branched chains. The K48-K63 branched linkage permits recognition by TAB2 but protects K63 linkages from CYLD-mediated deubiquitylation, thereby amplifying NF-?B signals. These results reveal a previously unappreciated cooperation between K48 and K63 linkages that generates a unique coding signal: ubiquitin chain branching differentially controls readout of the ubiquitin code by specific reader and eraser proteins to activate NF-?B signaling.

  13. I have added to the OP a beautiful animation of the ubiquitin-proteasome system in degrading damaged proteins.

  14. In the OP I have barely mentioned the role of ubiquitination in DNA repair.

    This is interestng, because ubiquitination acts mainly at the level of histones.

    We know that Post Translational Modifications (PTMs) of histones are in themselves a code, the histone code. This code is, too, highly symbolic, and very complex indeed. See, for example, Wikipedia:

    https://en.wikipedia.org/wiki/Histone_code

    However, the histone code is mainly written by simpler PTMs, especially methylation (mono, di and tri methylation) and acetylation. The role of ubiquitin here is minor.

    But in the case of DNA damage, especially Double-Strand Breaks (DSB), ubiquitin becomes the absolute protagonist, and directs the repair process in an extremely complex and detailed way. See, for example, this paper:

    Writers, Readers, and Erasers of Histone Ubiquitylation in DNA Double-Strand Break Repair

    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4923129/

    The main role seems to be played by two E3 ligases, RNF8 and RNF 168, but a lot of other proteins are involved.

    So, this is another example of two complex symbolic codes (histone code and ubiquitin code) strongly interacting, in a highly dynamic and articulated pattern.

  15. This is the most amazing thing I have ever heard.

  16. butifnot:

    There are a lot of highly amazing things in biology.

    But we have to dig deep into them, to really understand and appreciate their complexity and value.

    As in many other contexts, the devil is in the details! 🙂

  17. And, of course, deubiquitinating enzymes have a key role in regulating DNA repair. See here:

    Fine-tuning the ubiquitin code at DNA double-strand breaks: deubiquitinating enzymes at work

    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4561801/

    If you love simplicity, just look at Figure 1! And carefully read the Figure legend… 🙂

  18. gpuccio @16:

    There are a lot of highly amazing things in biology

    That’s an understatement. 🙂

  19. Mitosis ensures equal segregation of the genome and is controlled by a variety of ubiquitylation signals on substrate proteins.

    However, it remains unexplored how the versatile ubiquitin code is read out during mitotic progression.

    Here, we identify the ubiquitin receptor protein UBASH3B as an important regulator of mitosis.

    UBASH3B interacts with ubiquitylated Aurora B, one of the main kinases regulating chromosome segregation, and controls its subcellular localization but not protein levels.

    UBASH3B is a limiting factor in this pathway and is sufficient to localize Aurora B to microtubules prior to anaphase.

    Importantly, targeting Aurora B to microtubules by UBASH3B is necessary for the timing and fidelity of chromosome segregation in human cells.

    Our findings uncover an important mechanism defining how ubiquitin attachment to a substrate protein is decoded during mitosis.

    Ubiquitin Receptor Protein UBASH3B Drives Aurora B Recruitment to Mitotic Microtubules.

    Krupina, Ksenia & Kleiss, Charlotte & Metzger, Thibaud & Fournane, Sadek & Schmucker, Stephane & Hofmann, Kay & Fischer, Benoit & Paul, Nicodeme & Porter, Iain & Raffelsberger, Wolfgang & Poch, Olivier & Reese Swedlow, Jason & Brino, Laurent & Sumara, Izabela. (2016).

    Developmental Cell. 36. 63-78. 10.1016/j.devcel.2015.12.017.

    http://www.cell.com/developmen.....0826-6.pdf

  20. gpuccio @17:

    That figure 1 legend has material to fill post-doc course textbooks.

  21. This thread has too much bad news for the ‘modern synthesis’ and the ‘third way’ clubs. 🙂

  22. The cell division cycle is driven by a collection of enzymes that coordinate DNA duplication and separation, ensuring that genomic information is faithfully and perpetually maintained. The activity of the effector proteins that perform and coordinate these biological processes oscillates by regulated expression and/or posttranslational modifications. Ubiquitylation is a cardinal cellular modification and is long known for driving cell cycle transitions. In this review, we emphasize emerging concepts of how ubiquitylation brings the necessary dynamicity and plasticity that underlie the processes of DNA replication and mitosis. New studies, often focusing on the regulation of chromosomal proteins like DNA polymerases or kinetochore kinases, are demonstrating that ubiquitylation is a versatile modification that can be used to fine-tune these cell cycle events, frequently through processes that do not involve proteasomal degradation. Understanding how the increasing variety of identified ubiquitin signals are transduced will allow us to develop a deeper mechanistic perception of how the multiple factors come together to faithfully propagate genomic information. Here, we discuss these and additional conceptual challenges that are currently under study toward understanding how ubiquitin governs cell cycle regulation.

    Gilberto, Samuel & Peter, Matthias. (2017). Dynamic ubiquitin signaling in cell cycle regulation. The Journal of Cell Biology. 216. jcb.201703170. 10.1083/jcb.201703170.

    http://jcb.rupress.org/content.....l-text.pdf

  23. a deubiquitinating enzyme complex regulates the mitotic spindle assembly factor NuMA.

    BRISC binds and deubiquitinates the spindle assembly factor NuMA

    Spindle assembly at a BRISC pace

    Compared with a control cell (left), microtubules (green) form a multipolar spindle in a cell (right) lacking the BRISC component ABRO1 (red). DNA is labeled blue.

    Yan et al. describe how a deubiquitinating enzyme complex regulates the mitotic spindle assembly factor NuMA.
    BRCC36 is a deubiquitinating enzyme that preferentially cleaves lysine 63–linked polyubiquitin chains. As part of the Rap80 complex, BRCC36 regulates the repair of DNA double-strand breaks, but the enzyme can also assemble into a distinct complex, known as BRISC.
    Yan et al. found that knocking down BRCC36, or a subunit unique to the BRISC complex called ABRO1, caused mitotick cells to assemble multipolar spindles that frequently aligned and segregated chromosomes incorrectly. These defects could not be rescued by a catalytically inactive version of BRCC36. In wild-type mitotic cells, BRISC accumulated at the spindle poles, bound to the minus ends of stable, kinetochore-attached microtubules. Early in mitosis, BRISC also localized near the kinetochores themselves, where it promoted the chromosome-dependent nucleation of spindle microtubules.
    Yan et al. discovered that BRISC binds and deubiquitinates the spindle assembly factor NuMA, which captures and focuses microtubules at spindle poles. In the absence of BRISC, ubiquitinated NuMA showed an increased association with both importin-? and dynein, which regulate the protein’s function.
    BRISC therefore promotes bipolar spindle assembly by deubiquitinating NuMA. Senior author Genze Shao says that BRISC may have other mitotic substrates as well. Because some BRISC-deficient cells progress through mitosis, despite their disorganized spindles, he is particularly interested in whether the deubiquitinase can regulate the spindle assembly checkpoint.

    • Yan, K., et al0
    . 2015. J. Cell Biol.
    doi:10.1083/jcb.201503039
    • Ben Short

  24. Ubiquitination is a post-translational modification that defines the cellular fate of intracellular proteins. It can modify their stability, their activity, their subcellular location, and even their interacting pattern. This modification is a reversible event whose implementation is easy and fast. It contributes to the rapid adaptation of the cells to physiological intracellular variations and to intracellular or environmental stresses. E2F1 (E2 promoter binding factor 1) transcription factor is a potent cell cycle regulator. It displays contradictory functions able to regulate both cell proliferation and cell death. Its expression and activity are tightly regulated over the course of the cell cycle progression and in response to genotoxic stress. I discuss here the most recent evidence demonstrating the role of ubiquitination in E2F1’s regulation.

    Dubrez, Laurence. (2017). Regulation of E2F1 Transcription Factor by Ubiquitin Conjugation. International Journal of Molecular Sciences. 18. 2188. 10.3390/ijms18102188.

  25. Human gut Bacteroides species produce different types of toxins that antagonize closely related members of the gut microbiota. Some are toxic effectors delivered by type VI secretion systems, and others are non-contact-dependent secreted antimicrobial proteins. Many strains of Bacteroides fragilis secrete antimicrobial molecules, but only one of these toxins has been described to date (Bacteroidales secreted antimicrobial protein 1 [BSAP-1]). In this study, we describe a novel secreted protein produced by B. fragilis strain 638R that mediated intraspecies antagonism. Using transposon mutagenesis and deletion mutation, we identified a gene encoding a eukaryotic-like ubiquitin protein (BfUbb) necessary for toxin activity against a subset of B. fragilis strains. The addition of ubb into a heterologous background strain conferred toxic activity on that strain. We found this gene to be one of the most highly expressed in the B. fragilis genome. The mature protein is 84% similar to human ubiquitin but has an N-terminal signal peptidase I (SpI) signal sequence and is secreted extracellularly. We found that the mature 76-amino-acid synthetic protein has very potent activity, confirming that BfUbb mediates the activity. Analyses of human gut metagenomic data sets revealed that ubb is present in 12% of the metagenomes that have evidence of B. fragilis. As 638R produces both BSAP-1 and BfUbb, we performed a comprehensive analysis of the toxin activity of BSAP-1 and BfUbb against a set of 40 B. fragilis strains, revealing that 75% of B. fragilis strains are targeted by one or the other of these two secreted proteins of strain 638R.

    Chatzidaki-Livanis, Maria & Coyne, Michael & G. Roelofs, Kevin & R. Gentyala, Rahul & M. Caldwell, Jarreth & E. Comstock, Laurie. (2017). Gut Symbiont Bacteroides fragilis Secretes a Eukaryotic-Like Ubiquitin Protein That Mediates Intraspecies Antagonism. mBio. 8. e01902-17. 10.1128/mBio.01902-17.

  26. Gpuccio,

    Nice, glad videos are of help 🙂

    I’ve yet to get through the papers you and Dionisio have shared.

    Like at your #14 post…
    “So, this is another example of two complex symbolic codes (histone code and ubiquitin code) strongly interacting, in a highly dynamic and articulated pattern.”

    So which code arrived 1st? Simultaneous Code building by random mutations and natural selection?

    Doubtful.

    Obviously Histone must be in place or can you imagine the journey along the length of DNA not packaged by Histones?

    And did ubiquitin just pop up one day and say, voila, I’m code and recognize histone binding targets and building pathways for DNA repair?

    Obviiously, I’m being a bit loose and humorous, but there’s a point. And simple questions.

    If the two codes do not arise at same time or not built simultaneously working together, then what?

    So neo-Darwinist have addressed this, but how adequate is their explanation?

    What happens if ubiguitin code and enzyme regulation are not available?

    What happens to DNA Repair?

    And how often is repair required, double-stranded that is?

    Curious what neo-Darwinist propose as evolutionary history for multiple codes, pathways, signal recognition and repair functions rising together. As well as coordination of these systems.

    Will have more time to review tonight.

    Very good work Gpuccio! Thanks again 🙂

  27. Dionisio:

    I have been missing your support! 🙂

    Yes, mitosis and cell division seem to be under the realm of ubiquitin control, too.

    About the paper you referenced at #22, it is rather surprising how some concept constantly recur in ubiquitin literature. For example, just from the abstract of this paper:

    dynamicity
    plasticity
    versatile
    fine-tune
    variety
    signals
    conceptual challenges

    Maybe my focus on semiosis in the OP is not so bad! 🙂

  28. Dionisio:

    Fig. 1 of the same paper is a very good summary of:

    “Writing, reading, and editing ubiquitin.”

    Semiosis everywhere!

    Fig. 2 from the same paper is a very good summary of cullin based modular proteins, and it adds the even more complex APC/C structure (at least 14 different subunits!). From the Legend of the Figure:

    Although the APC/C is closely related to CRLs and contains the cullin-homology subunit APC2 (Yu et al., 1998), it is structurally divergent. The APC/C is composed of at least 14 different subunits, including the RING subunit APC11, plus one of two coactivators (CDC20 and CDH1) that also participate in substrate binding (Sivakumar and Gorbsky, 2015). The APC/C operates in mitosis and G1 and is mostly known for its ability to degrade mitotic cyclins and other mitotic factors so that chromosomes are separated and mitotic exit ensues (Zhou et al., 2016).

  29. DATCG:

    Thank you again for you very good thoughts.

    I would like to add some comments about the points you make, but I have not the time now. I will come back later! 🙂

  30. DATCG:

    Histone code, Ubiquitin code, cell signaling, and many other things, are essentially new layers of complexity that appear in eukaryotes, and become more stratified in metazoa.

    My personal idea is that eukaryotes and metazoa require, just from the beginning, a whole new design approach that is based on many interacting levels of regulation networks.

    And the key word is: semiosis and regulation.

    Of course, high levels of semiosis and regulation are already present in prokaryotes. But the eukaryotic world seems to be structured according to new concepts, and here is where we start to see these many parallel levels of control, each of them extremely important for practically all cell functions, each of them relatively independent, and yet each of them intertwined with all the others.

    So we have all the epigenetic layers of transcription regulation, which multiply the possibilities of a rather static genome: DNA methylation, histone code, chromatin remodeling, and so on.

    And we have the infinitely intricate netwotk of TFs, with their combinatorial working.

    And then the complex post-transcriptional regulation, the intron system, the spliceosome, miRNAs, lncRNAs.

    And a lot of different PTMs, including the Ubiquitination System which is the object of this discussion.

    And, last but not least, cell to cell signaling, and the multitude of signals and receptors and of transmission pathways that convey the symbolic meaning of the signal from the membrane receptor, through definite intemediate steps, and ultimately to the TF network and DNA.

    The amazing thing is that there is not one of these levels which does not control the same things as all the other levels, but in different ways, and that is not strictly interconnected with all the other levels. It’s like a multiple control strategy whose complexity grows exponentially, and whose ultimate purpose can only be to attain functional outcomes which would be absolutely impossible without control, or with single level control.

    You ask: “So which code arrived 1st?” I think that the obsvious answer is: they are all part of the same project, they were conceived and implemented in a parallel process, just from the beginning, to realize a design of increasing complexity and efficiency of which we no equivalent is known.

  31. Dionisio:

    The bacteroides paper you link is really intriguing. It seems an obvious case or HGT with adaptation, but it is certainly atypical and stimulating.

    The most amazing fact is that this ubiquitin-like protein is used as a toxin against similar bacterial species. Infortunately, the mechanism of action of the protein in that role has not been clarified. that would really be interesting.

    It’s interesting that the authors have no doubts that the bacterial protein was acquired, either from eukaryotes or from giant viruses. So, they opt for HGT just from the beginning.

    That seems obvious to me too, with the protein having 67% identities and 84% positives with human ubiquitin (bitscore 108 bits, E value 2e-38). It is clearly a derivation from eukaryotic ubiquitin. But I think a pure neo-darwinist could also propose some form of convergent evolution, or what?

    The simple truth is: 108 bits of information require an explanation. That’s why the authors state (rightly): “The source from which ubb was acquired is not clear from existing genomic sequences.”

    OK, but what about the thousands of bits that appear in thousand of individual protein, for example, at the origin of vertebrates? Or in eukaryotes? Don’t they deserve some explanation too?

    Am I asking too much? 🙂

  32. Hello GP, I am just now seeing your fantastic OP.

    Man’o man, GP, what a wonderful job you’ve done in explaining the system and highlighting the scope of the issue. If you don’t write a book, then there is something truly wrong with the world.

    I’ve just read the OP, haven’t read the comments.

    It all functions via semiosis — from the very start.

  33. GP, you and I once spoken briefly about Marcello Barbieri holding his annual “Code Biology” conference in Italy. I suspect this is precisely the type of information covered in those labs and presentations.

  34. GP, your presentation is just excellent. Where are your opponents, my friend? I don’t see them. Are there no faithful left to carry the RV+NS banner into battle?

  35. Reading this:

    But what “translates” the complex ubiquitin code, allowing ubiquinated proteins to met the right specific destiny? Again, we can refer to the diubiquitin paper quoted above.

    How do cells decode this ubiquitin code into proper cellular responses? Recent studies have indicated that members of a protein family, ubiquitin-binding proteins (UBPs), mediate the recognition of ubiquitinated substrates. UBPs contain at least one of 20 ubiquitin-binding domains (UBDs) functioning as a signal adaptor to transmit the signal from ubiquitinated substrates to downstream effectors

    But what are those “interactors” identified by the paper (at least 46 of them)? They are, indeed, complex proteins which recognize specific configurations of the “tag” (the ubiquitin chain), and link the tagged (ubiquinated) protein to other effector proteins which implement its final fate, or anyway contribute in deffrent forms to that final outcome.

    ….sounds real familiar. 🙂 🙂

  36. It reminds me of this… (if I may):

    (from previous writing)

    Pattee recognized that representation and interpretation are two necessary and complementary roles in a very specific physical architecture; an organization where one arrangement of matter serves as a token of memory, while another arrangement of matter determines what is being represented by the arrangement of the token. Pattee further recognized that for this architecture to serve as the basis of open-ended variation, the referent would not be determined by the arrangement of the token. Instead, the referent would be independently specified by a second “interpreting” arrangement of matter, which he described as a non-integrable constraint.

    Symbols do not exist in isolation but are part of a semiotic or linguistic system (Pattee, 1969a). Semiotic systems consist of (1) a discrete set of symbol structures (symbol vehicles) that can exist in a quiescent, rate-independent (non-dynamic) states, as in most memory storage, (2) a set of interpreting structures (non-integrable constraints, codes), and (3) an organism or system in which the symbols have a function (Pattee, 1986). – H.H. Pattee, “The Physics of Symbols: Bridging the Epistemic Cut”. Biosystems. Vol. 60, pp. 5-21. 2001

    In a semiotic system, a non-integrable constraint is an arrangement of matter that physically establishes a trajectory for the system (among alternatives) depending on the individual representation being translated. […] They are referred to as “non-integrable” because a description of the constraints cannot be integrated with a lawful description of the system itself. Thus, the logician’s complementary roles of representation and interpretation are reflected in the physicist’s requirement of complementary descriptions as well — one for the dynamic and another for the symbolic aspects of the system.

  37. Perhaps Art Hunt will stop by and pick up on this excellent OP (and also get back to the unfinished business from your previous OP as well).

    🙂

  38. Gpuccio…

    Thanks for your follow-up and answers. Quickly #30.
    I agree with it being a coordinated, parallel process. And I take note of your thoughts…

    My personal idea is that eukaryotes and metazoa require, just from the beginning, a whole new design approach that is based on many interacting levels of regulation networks.

    Agree with “require… whole new design approach”

    And new codes. Codes that must be interpreted, recognized
    and functional.

    Hardly amenable to random mutations. The amount of just-so happenstance for multiple proteins, signals and systems to simultaneously coordinate seem to be insurmountable.

  39. Good to see you UB. Was responding on GPuccio’s other point.

    #30 in his line below…

    And the key word is: semiosis and regulation.

    Yes, and look forward to hearing from UB on semiosis.

    ———————-
    🙂 ha! Great timing UB. I almost posted then saw your comments.

    So funny as I found your post form March 2016 just minutes earlier and was reading it and about Marcello Barbieri.

    Here is his autobiography…

    Marchello Barbieri Autobiography

    .

  40. #37 UB

    Arthur Hunt, one can hope.

    I thought he might provide some good insight on evolution of the Spliceosome via some of the comments he put forward.

    But nothing since then.

  41. From Barbieri’s autobiography page, interesting to note…

    Many other organic codes have been discovered. Among them, the sequence codes (Trifonov 1987, 1989, 1999), the adhesive code (Redies and Takeichi 1996; Shapiro and Colman 1999), the splicing codes (Barbieri 2003; Fu 2004; Matlin et al. 2005; Pertea et al. 2007; Wang and Burge 2008; Barash et al. 2010; Dhir et al. 2010), the signal transduction codes (Barbieri 2003), the histone code (Strahl and Allis 2000; Jenuwein and Allis 2001; Turner 2000, 2002, 2007; Kühn and Hofmeyr 2014), the sugar code (Gabius 2000, 2009), the compartment codes (Barbieri 2003), the cytoskeleton codes (Barbieri 2003; Gimona 2008), the tubulin code (Verhey and Gaertig 2007), the nuclear signalling code(Maraldi 2008), the apoptosis code (Basañez and Hardwick 2008; Füllgrabe et al. 2010), the ubiquitin code (Komander and Rape 2012), the bioelectric code (Tseng and Levin 2013; Levin 2014), the glycomic code (Buckeridge and De Souza 2014) and the acoustic codes (Farina and Pieretti 2014).

    How many other codes not included or to be found in future?
    Layer upon layer.

    These discoveries have extraordinary consequences because they change our reconstruction and our understanding of the history of life. Any time that a new organic code came into being, something totally new appeared in Nature, something that had never existed before.

    I think Gpuccio is on target re: semiosis and regulation.

    .

  42. Yes, Barbieri is an interesting fellow, and I surely appreciate his work. But there is a real problem — setting aside for a moment the fact that he denies any room for design in biology. I can mentally work around that issue, but he also seams to find some difficulty is reconciling the encoding of biological information and the Peircean concept of interpretation. He sees them in some strange conflict with one another.

    I truly don’t get his issue there. As far as I can see, Pattee put that issue completely to bed decades ago in the late 1960s. The actual physics of the system shot the deciding point. There is no such thing as a code without interpretation.

    – – – – – – – –

    anyway, not taking GP’s excellent OP off course

  43. I think Gpuccio is on target re: semiosis and regulation.

    No question about it.

  44. Pattee, 1969

    How do we tell when there is communication in living systems? Most workers in the field probably do not worry too much about defining the idea of communication since so many concrete, experimental questions about developmental control do not depend on what communication means. But I am interested in the origin of life, and I am convinced that the problem of the origin of life cannot even be formulated without a better understanding of how molecules can function symbolically, that is, as records, codes, and signals. Or as I imply in my title, to understand origins, we need to know how a molecule becomes a message.

  45. Upright BiPed (and DATCG):

    Welcome! 🙂 🙂

    It was really a beautiful surprise to awaken and find your many comments here!

    And your beautiful discussions with DATCG too…

    I was certain that you would like the semiosis angle. 🙂

    OK, our interlocutors, as usual, are nowhere to be seen, but at least I have some true friends! So thanks be given to you, DATCG, Dionisio. 🙂

    I am in a hurry now, so I will be back as soon as possible for some comments. I just wanted to thank all of you.

  46. Upright Biped:

    “sounds real familiar”.

    And it is!

    There are many similarities with the best known example of semiotic system in biology: the genetic code.

    I would like to make a few reflections on the similarities and differences between the two systems:

    Nature of the code:

    The genetic code is in some measure more clear-cut. While redunndant, it is certainly strict and context independent. Codons have one unequivocal meaning.

    The ubiquitin code, like many other biologcial codes, seems to be nore fluid, and probably more context-dependent. In a sense, these seem to be higher forms of code, serving a more complex function.

    However, one specific feature of the ubiquitin code is that it relies practically on one single molecule, ubiquitin (and a few minor variants), and the main nature of the alphabet depends on the variety of chains that can be realized using the 8 internal switches. In that sense, there is an homogeneity of the tool which remind the genetic code, while other symbolic systems (like signal receptor systems) have greater variation and specificity in the nature of the signal.

    Writing the code:

    This is probably the gratest difference between the two systems. The information content coded by the genetic code is pre-existent. It is not written by the cell, it is simply inherited. In a sense, the information is written by the designer (ID point of view) or by the RV + NS process (neo-darwinist point of view). Indeed, one of the main objections against “third way speculations” is that there is no known mechanism in the cell that allows a flow of information from the phenotype to the genome (protein coding genes): IOWs, the cell does not know how to write new information (existing in the phenotype) at the level of protein coding genes.

    On the other hand, the information coded by the ubiquitin code is written by the cell: it is essentially epigenetic information, not heritable genetic information. Of course, the writing is probably guided in some measure by the genome, and probably in some other measure by the epigenome, but the writing itself is implemented by the individual cell.

    More in next post.

  47. Upright Biped:

    Let’s go on with the comparison between the Ubiquitin Code and the Genetic code.

    Reading the code:

    The genetic code, as we well know, is read by the 20 Aminoacyl tRNA synthetases, through the correctly charged tRNAs. This is where the code is really understood.

    The Ubiquitin code is similarly read by a number of specific proteins (maybe 40 – 100) which share a number of ubiquitin-binding domains (UBDs): about 20.

    So, the reading system seems to be a little more complex here, but essentially comparable to the reading system in the genetic code.

    The final effector:

    Here, too, the genetic code system seems to be relatively simpler: the final effector is the ribosome, and translation. However complex this is, it is still a rather homogeneous task.

    Not so in the Ubiquitin System: while there is one major effector (proteasomal degradation), we have seen that it is not the only one: indeed, a lot of important effects, in almost all cell systems, are mediated by specific effects on different protein – protein interactions and protein configurations, without involving proteasomal degradation in any way.

    More in next post.

  48. Upright Biped (and DATCG):

    And, finally:

    The complexity:

    While all the coding systems we have considered (and I would say all those quoted by DATCC at #41 from Barbieri’s autobiography page) are certainly hugely complex from any functional point of view, still the Ubiquitin System has a very special feature: the very high number of individual proteins involved.

    Indeed, with about 1000 proteins involved specifically in the system, it is probably the cellular process with the greatest number of protein coding genes in the genome. We must remember that 1000 protein coding genes out of 20000 is almost 5% of the whole protein coding genome.

    The only other example which comes to mind is olfactory receptors, which corrspond to about 1000 in the mammalian genome (but only 400 active in the human genome).

    But olfactory receptors are a class of not too long proteins which share a great sequence homology in many cases (50% or more), and so appear to be a diversification of similar molecules.

    Instead, E3 ligases, which make up the greatest part of proteins involved in the Ubiquitin System (600+), are a set of really distinct proteins, except for the shared domians. But, as already said, the shared domains are in general a minor part of the molecule. The RING domain, for example, which characterizes most of E3 ligases, is only 40 – 60 AAs long: in a class of proteins which has a mean length of about 500 AAs, that is certainly a minor part of the sequence information.

    The rest of the sequence is certainly involved in the specific function of each E3 ligase: to recognize the correct target protein, in the correct context. And each of the 600+ E3 ligases is completely different from each other in that respect.

    That makes a real lot of specific functional information in the system.

    Even from a proteomic point of view, the system represents a huge part of cell resources, its proteins amounting to about 1.3% of the total cell proteins, as stated here:

    The demographics of the ubiquitin system.

    http://www.cell.com/trends/cel.....15)00054-9

    Many cell resources are used in many systems to generate variety. But while in some systems, like the immune system, variety is created using controlled variation on a limited number of genes, in the ubiquitin system (like in the olfactory system) variety is mostly paid for at genomic level. So, a lot of resources in the genome and the proteome are committed to that, an most of the sequence functional information is directly coded in the genome.

  49. Just adding to our list of cell processes finely regulated by the Ubiquitin System. What are we missing?

    The central nervous system? Neurons? Synapses?

    Neuronal ubiquitin homeostasis.

    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3758786/

    Abstract
    Neurons have highly specialized intracellular compartments that facilitate the development and activity of the nervous system. Ubiquitination is a post-translational modification that controls many aspects of neuronal function by regulating protein abundance. Disruption of this signaling pathway has been demonstrated in neurological disorders such as Parkinson’s disease, Amyotrophic Lateral Sclerosis and Angleman Syndrome. Since many neurological disorders exhibit ubiquitinated protein aggregates, the loss of neuronal ubiquitin homeostasis may be an important contributor of disease. This review discusses the mechanisms utilized by neurons to control the free pool of ubiquitin necessary for normal nervous system development and function as well as new roles of protein ubiquitination in regulating synaptic activity.

    Following the de novo generation of ubiquitin in the cell body, ubiquitin is transported from the soma to distant locals like axons and dendrites. A single study in the literature indicates that ubiquitin is trafficked via slow axonal transport down the rat optic nerve [6]. This transport proceeds at a rate of approximately 3 mm/day, indicating that the length of time required for newly generated ubiquitin to reach synaptic terminals is on the order of days, or even weeks, in some neurons.

    Since the original description of the ubiquitin proteasome system by Ciechanover, Hershko, and Rose in the late 1970’s and early 1980’s, numerous studies have linked ubiquitin to neuronal function [51]. While the contribution of protein ubiquitination in regulating synaptic protein abundance is well documented, there are hints in the literature of additional roles for ubiquitin at the synapse, which may be independent of protein degradation.

    Nedd4 and Nedd4-2: ubiquitin ligases at work in the neuron.

    Abstract
    Ubiquitination of proteins by the Nedd4 family of ubiquitin ligases is a significant mechanism in protein trafficking and degradation and provides for tight spatiotemporal regulation. Ubiquitination is gaining increasing recognition as a central mechanism underpinning the regulation of neuronal development and homeostasis in the brain. This review will focus on the Nedd4 and Nedd4-2 E3 ubiquitin ligases that are implicated in an increasing number of neuronal protein-protein interactions. Understanding of the contribution of Nedd4 and Nedd4-2 in regulating key functions in the brain is shedding new light on the ubiquitination signal not only in orchestrating degradation events but also in protein trafficking. Furthermore, the description of several novel Nedd4/4-2 targets in neurons is changing the way we conceptualize how neurons maintain normal function and how this is altered in disease.

    Seven in Absentia E3 Ubiquitin Ligases: Central Regulators of Neural Cell Fate and Neuronal Polarity

    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5646344/

    Abstract
    During neural development, neural precursors transition from a proliferative state within their germinal niches to a migratory state as they relocate to their final laminar positions. Transitions across these states are coupled with dynamic alterations in cellular polarity. This key feature can be seen throughout the developing vertebrate brain, in which neural stem cells give rise to multipolar or unpolarized transit-amplifying progenitors. These transit-amplifying progenitors then expand to give rise to mature neuronal lineages that become polarized as they initiate radial migration to their final laminar positions. The conventional understanding of the cellular polarity regulatory program has revolved around signaling cascades and transcriptional networks. In this review, we discuss recent discoveries concerning the role of the Siah2 ubiquitin ligase in initiating neuronal polarity during cerebellar development. Given the unique features of Siah ubiquitin ligases, we highlight some of the key substrates that play important roles in cellular polarity and propose a function for the Siah ubiquitin proteasome pathway in mediating a post-translational regulatory network to control the onset of polarization.

    Spatial Organization of Ubiquitin Ligase Pathways Orchestrates Neuronal Connectivity

    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3622823/

    Abstract
    Recent studies reveal that E3 ubiquitin ligases have essential functions in the establishment of neuronal circuits. Strikingly, a common emerging theme in these studies is that spatial organization of E3 ubiquitin ligases plays a critical role in the control of neuronal morphology and connectivity. E3 ubiquitin ligases localize to the nucleus, centrosome, Golgi apparatus, axon and dendrite cytoskeleton, and synapses in neurons. Localization of ubiquitin ligases within distinct subcellular compartments may facilitate neuronal responses to extrinsic cues as well as the ubiquitination of local substrates. Here, we will review the functions of neuronal E3 ubiquitin ligases at distinct subcellular locales and explore how they regulate neuronal morphology and function in the nervous system.

    See in particular Fig. 2 and Fig. 3.

    And so on, and so on…

  50. On the other hand, the information coded by the ubiquitin code is written by the cell: it is essentially epigenetic information, not heritable genetic information. Of course, the writing is probably guided in some measure by the genome, and probably in some other measure by the epigenome, but the writing itself is implemented by the individual cell.

    Fascinating isn’t it. The cell is telling you what it is; layers of interdependence, and always yet another layer to be understood.

  51. Indeed, with about 1000 proteins involved specifically in the system, it is probably the cellular process with the greatest number of protein coding genes in the genome. We must remember that 1000 protein coding genes out of 20000 is almost 5% of the whole protein coding genome.

    I am interested in how that compares numerically to the prokaryotic system.

  52. #48. I really appreciate the numbers you’ve thought through and placed in your presentation. Understanding context is a real bonus for your readers.

  53. #42-44 UB,

    yes, I can work around it too. I found it a bit amusing to a degree. What he’s advocating is screaming Design.

    The Codes, The Codes! 😉

    It’s why I posted them.

    I’ve not time to read Pattee, which I briefly looked at on your earlier post. Will look again when time permits.

    Pattee is right, your quote sums it up nicely.

    “I am convinced that the problem of the origin of life cannot even be formulated without a better understanding of how molecules can function symbolically, that is, as records, codes, and signals. Or as I imply in my title, to understand origins, we need to know how a molecule becomes a message.

    Indeed how is a correct code understood to be correct?
    After a billion, maybe trillion combinations?

    Hmmmmm. Where and when does time and resources become a factor?

    As more function in “junk” DNA is found, the limits to such random “success” becomes smaller as well.

    Will Dan Graur’s threshold for “junk” DNA hold? Will he be found correct, or ENCODE correct and as Dan said, “evolution is wrong?”

    .

  54. Sorry for this off topic question:

    Can this be considered a code issue too?

    Decoding temporal interpretation of the morphogen bicoid in the early drosophila embryo

    Huang, A & Amourda, C & Zhang, S & Tolwinski, Nicholas & Saunders, Timothy. (2017). eLife. 6. . 10.7554/eLife.26258.

    Morphogen gradients provide essential spatial information during development.

    Not only the local concentration but also duration of morphogen exposure is critical for correct cell fate decisions.

    Yet, how and when cells temporally integrate signals from a morphogen remains unclear.

    Here, we use optogenetic manipulation to switch off Bicoid-dependent transcription in the early Drosophila embryo with high temporal resolution, allowing time-specific and reversible manipulation of morphogen signalling.

    We find that Bicoid transcriptional activity is dispensable for embryonic viability in the first hour after fertilization, but persistently required throughout the rest of the blastoderm stage. Short interruptions of Bicoid activity alter the most anterior cell fate decisions, while prolonged inactivation expands patterning defects from anterior to posterior. Such anterior susceptibility correlates with high reliance of anterior gap gene expression on Bicoid. Therefore, cell fates exposed to higher Bicoid concentration require input for longer duration, demonstrating a previously unknown aspect of Bicoid decoding.

  55. DATCG,

    Barbieri wrote an paper, I think entitled “A Short History of Semiotics”, where he recounts the point in time when all the various factions of semiotic thought came together and sought to work together. He describes two postulates that help forge the agreement to come together. The first postulate (recalling from memory) is that semiosis and life are coextensive, i.e. one does not exist without the other. And if you read the paper, he goes on to fully support that postulate. The second postulate, however, was that they (the group) would have nothing to do with any argument for design in biology. Never again throughout the paper is the design argument even mentioned, other than to kill it up front.

    The fact that this kind of maneuver was seen as necessary says quite a bit about the argument for design.

  56. Gpuccio,

    On your original post you mention Ubiquitin Code and UPS in relation to Autophagy above.

    The paper in that section is behind a paywall. I found some interesting info to add, an Open Access Book –
    Chapter 7 The Role of Ubiquitin System in Autophagy
    from 2016.

    Book Link Here-Chapter 7 Role of Ubiquitin in Autophagy

    Specifically on role of Ubiquitin and DUBS, modifications and Selective role in cargo packages for degradation. Pointing out autophagy “was originally thought to be non-selective” via random process for degradation.

    But as is highlighted here and through chapter 7, Ubiquitin and DUBS play a role in tagging cargo for recognition and selection, not random but induced by stress for example.

    This is something I’d expect from an epigenetic point of view. The higher level code using signal processing, tagging as intervention whether positive or negative.

    Not to get off any central points. But guide us where you would like to focus 🙂

    4. Ubiquitination modification and selective autophagy

    Although autophagy was originally thought to be a non-selective pathway which appears to randomly sequester cytosolic components for lysosomal degradation, it is now recognized that autophagy also acts in selective processes that involves specific receptors to target certain cargos [60, 61]. Accumulating evidence indicates that many intracellular degradation events are processed through selective autophagy, including the turnover of damaged organelles such as mitochondria (mitophagy) [62, 63] and peroxisomes (pexophagy) [64, 65], removal of protein aggregates (aggrephagy) [66], and elimination of intracellular pathogens (xenophagy) [67, 68].

    Upon the induction of selective autophagy, phagophore is enriched with specific cargos in a process dependent on cargo receptors [61]. These cargo receptors can interact with both target proteins and the autophagic vesicle components such as LC3/Atg8 family proteins, which result in the enclosure of selective cargos to the autophagosome and promote the autophagic degradation of cargos. Like nonselective autophagy, selective autophagy also plays an important role in cellular homeostasis and has been associated with a variety of human diseases [63, 69].

    4.1. THE ROLE OF UBIQUITIN IN SELECTIVE AUTOPHAGY

    Ubiquitination has long been recognized as a key regulator to determine protein fate by tagging proteins for proteasomal degradation [60]. Ubiquitination of cargo proteins plays a crucial role in selective autophagy process. In selective autophagy, cargos are ubiquitinated and recognized by ubiquitin-binding receptors to transport cargos for lysosomal degradation [70]. Therefore, ubiquitin acts as a degradation signal for selective autophagy. Protein aggregates, damaged organelles, or pathogens can be tagged and targeted for degradation through the lysosome machinery to maintain cellular homeostasis. In this section, we will illustrate the mechanism and importance of ubiquitination in selective autophagy (Figure 2).

    Recent studies have shown that selective autophagy is responsible for delivering a wide range of cargos to the lysosome for degradation [70–72]; however, the detailed mechanisms of selective degradation by lysosome remain largely unknown. Several types of adaptor proteins such as p62, NDP52, optineurin (OPTN), NBR1, and HDAC6, which contain the ubiquitin-binding motif, have been reported to target ubiquitinated cargos for lysosomal degradation under stress conditions [70, 71]. Besides the ubiquitin-binding motif, these cargo receptors often also contain a LC3-interacting region (LIR) or Atg8 interaction motif to interact with the LC3/Atg8 family members [60, 70]. Therefore, through binding to ubiquitinated cargos and LC3 simultaneously, these receptors can deliver selective cargos to the autophagosome and promotes the autophagic degradation.

    And in conclusion from Chapter 6….

    Protein ubiquitination is considered as one of the most important reversible posttranslational modifications and has been implicates in various cellular signaling processes.

    Increasing evidence indicates that the ubiquitin system plays a pivotal role in the regulation of autophagy pathway. Recent studies have explored and highlighted the important functions of ubiquitin system in the pathogenesis of autophagy-related diseases such as tumorigenesis, neurodegeneration, and pathogen infection.

    Further investigations to identify novel E3 ligases and DUBs involved in autophagy and to determine their underlying mechanisms will not only contribute to our understanding on how autophagy is controlled by the ubiquitin system but also provide a rationale for novel therapeutic interventions in autophagy-related diseases.

  57. #55 UB,

    Thanks, I caught a bit of that in his autobiography, not to such detail.

    Yes, agree. It’s a narrative they must stick to while oppressing any thought other than a materialistic, blind point of view of a “blind” “unguided” process.

    It’s an overall area to address by you and Gpuccio’s other comments he extended really well on Reading, Writing Code and comparisons of Code, a Base, Copying and Higher levels of Code.

  58. Ubiquitin turnover and endocytic trafficking in yeast are regulated by Ser57 phosphorylation of ubiquitin

    Despite its central role in protein degradation little is known about the molecular mechanisms that sense, maintain, and regulate steady state concentration of ubiquitin in the cell.

    Here, we describe a novel mechanism for regulation of ubiquitin homeostasis that is mediated by phosphorylation of ubiquitin at the Ser57 position.

    We find that loss of Ppz phosphatase activity leads to defects in ubiquitin homeostasis that are at least partially attributable to elevated levels of Ser57 phosphorylated ubiquitin.

    Phosphomimetic mutation at the Ser57 position of ubiquitin conferred increased rates of endocytic trafficking and ubiquitin turnover.

    These phenotypes are associated with bypass of recognition by endosome-localized deubiquitylases – including Doa4 which is critical for regulation of ubiquitin recycling.

    Thus, ubiquitin homeostasis is significantly impacted by the rate of ubiquitin flux through the endocytic pathway and by signaling pathways that converge on ubiquitin itself to determine whether it is recycled or degraded in the vacuole.

    Lee, Sora & M Tumolo, Jessica & C Ehlinger, Aaron & K Jernigan, Kristin & J Qualls-Histed, Susan & Hsu, Pi-Chiang & Hayes McDonald, W & Chazin, Walter & A MacGurn, Jason. (2017). Ubiquitin turnover and endocytic trafficking in yeast are regulated by Ser57 phosphorylation of ubiquitin. eLife. 6. . 10.7554/eLife.29176.

  59. In recent years, signaling through ubiquitin has been shown to be of great importance for normal brain development. Indeed, fluctuations in ubiquitin levels and spontaneous mutations in (de)ubiquitination enzymes greatly perturb synapse formation and neuronal transmission. In the brain, expression of lysine (K) 48-linked ubiquitin chains is higher at a developmental stage coincident with synaptogenesis. Nevertheless, no studies have so far delved into the involvement of this type of polyubiquitin chains in synapse formation. We have recently proposed a role for polyubiquitinated conjugates as triggering signals for presynaptic assembly. Herein, we aimed at characterizing the axonal distribution of K48 polyubiquitin and its dynamics throughout the course of presynaptic formation. To accomplish so, we used an ubiquitination-induced fluorescence complementation (UiFC) strategy for the visualization of K48 polyubiquitin in live hippocampal neurons. We first validated its use in neurons by analyzing changing levels of polyubiquitin. UiFC signal is diffusely distributed with distinct aggregates in somas, dendrites and axons, which perfectly colocalize with staining for a K48-specific antibody. Axonal UiFC aggregates are relatively stable and new aggregates are formed as an axon grows. Approximately 65% of UiFC aggregates colocalize with synaptic vesicle clusters and they preferentially appear in the axonal domains of axo-somatodendritic synapses when compared to isolated axons. We then evaluated axonal accumulation of K48 ubiquitinated signals in bead-induced synapses. We observed rapid accumulation of UiFC signal and endogenous K48 ubiquitin at the sites of newly formed presynapses. Lastly, we show by means of a microfluidic platform, for the isolation of axons, that presynaptic clustering on beads is dependent on E1-mediated ubiquitination at the axonal level. Altogether, these results indicate that enrichment of K48 polyubiquitin at the site of nascent presynaptic terminals is an important axon-intrinsic event for presynaptic differentiation.

    Pinto, Joana & Reis Pedro, Joana & Costa, Rui & Almeida, Ramiro. (2016). Visualizing K48 Ubiquitination during Presynaptic Formation By Ubiquitination-Induced Fluorescence Complementation (UiFC). Frontiers in Molecular Neuroscience. 9. . 10.3389/fnmol.2016.00043.

  60. re: #57 in reference to you UB and Gpuccio.

    I was trying to Edit, but to late.

    I thought the area covered by Gpuccio, semiosis and your comments on Barbieri, Codes, etc., symbols and recognition by Pattee, overall selective processes would be interesting to expand upon as well in more detail.

  61. GP, if I understand correctly, exactly how the medium is read is still being discovered. Perhaps I am wrong about that. I’ll have to find the time to study it. But from my own perspective, I would be interested in exactly what distinguishes one referent from another, and exactly how the system (process) acquires that from the medium.

    The Quipus

    For me, it matters how I would categorize the system in comparison to other systems.

    — again, great OP and further comments.

  62. #54 Dionisio,

    How’s this?

    Check out the following PDF paper…

    The emergent design of the neural tube: prepattern, SHH morphogen and GLI code

    Here, we review
    some basic aspects of Shh and Gli function to discuss
    how Shh acts as a neural tube morphogen responsible for
    combinatorial Gli function and whether it is only one of
    several informational inputs that create a morphogenetic
    Gli code.

    Notice where it’s published 😉

    Hint: Barbieri

  63. UB, think you might enjoy the following from the main page at the link I provided Dionisio. You may already read it…

    Two outstanding examples

    The genetic code
    In protein synthesis, a sequence of nucleotides is translated into a sequence of amino acids, and the bridge between them is realized by a third type of molecules, called transfer-RNAs, that act as adaptors and perform two distinct operations: at one site they recognize groups of three nucleotides, called codons, and at another site they receive amino acids from enzymes called aminoacyl-tRNA-synthetases. The key point is that there is no deterministic link between codons and amino acids since it has been shown that any codon can be associated with any amino acid (Schimmel 1987; Schimmel et al. 1993). Hou and Schimmel (1988), for example, introduced two extra nucleotides in a tRNA and found that that the resulting tRNA was carrying a different amino acid. This proved that the number of possible connections between codons and amino acids is potentially unlimited, and only the selection of a small set of adaptors can ensure a specific mapping. This is the genetic code: a fixed set of rules between nucleic acids and amino acids that are implemented by adaptors. In protein synthesis, in conclusion, we find all the three essential components of a code: (1) two independents worlds of molecules (nucleotides and amino acids), (2) a set of adaptors that create a mapping between them, and (3) the proof that the mapping is arbitrary because its rules can be changed.

    .

  64. DATCG,

    Here are some Pattee papers (among others) you might find interesting at some point.

    Bibliography

    The thing I appreciate about Pattee is that he is careful in his thoughts. He doesn’t force conclusions into his descriptions.

    Just imagine the rarity of that particular trait among researchers into the origin of life. There is even a point in one paper where Pattee (after giving a fairly comprehensive description of the issues) simply and briefly says that perhaps there was a time in distant history (paraphrasing) that chemistry acted differently than has at any time since. The man wasn’t trying to be clever; I believe it was a genuine puzzlement from a scientist who isn’t going to give up on (skew) a honest scientific analysis because of his own metaphysics.

    That trait (and the fact that he wrote about these issues for 50 years) is obviously endearing. He retired from writing a couple of summers ago. I’d say the scientific enterprise got their money’s worth.

  65. So much more to cover, not enough time. But Code Biology is Design Biology.

    And I thought what Gpuccio summed up here…

    Many cell resources are used in many systems to generate variety. But while in some systems, like the immune system, variety is created using controlled variation on a limited number of genes, in the ubiquitin system (like in the olfactory system) variety is mostly paid for at genomic level. So, a lot of resources in the genome and the proteome are committed to that, an most of the sequence functional information is directly coded in the genome.

    is important.

    will check in later 🙂

  66. Upright Biped, DATCG, Dionisio (and, of course, butifnot):

    It seems that we are having quite a private party here.

    A little bit staggered, I suppose, because of time zones and probably of my sleeping habits! 🙂

    However, it’s great to find your precious comment in the morning, and to answer them later. After all, do we really need the interventions of our courteous (but a little shy) interlocutors from the other side?

    No problems, the discussion is great, and we are deepening many interesting issues.

    As the song says:

    “Oh I get by with a little help from my friends” 🙂

  67. Upright Biped:

    “I am interested in how that compares numerically to the prokaryotic system.”

    Do you mean to whole prokaryotic genomes?

    Well, E. coli, which is a medium sized bacterium, has about 4000 protein coding genes. Some bacteria have less than 1000 protein coding genes.

    If you mean how the eukaryotic system compares to its supposed ancestors in prokaryotes, you can look at this paper:

    Prokaryotic Ubiquitin-Like Protein Modification

    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4757901/

    It seems that the prokaryotic analogs of ubiquitination, although certainly interesting, are rather different under many aspects, and much simpler.

    For example, the two main prokaryotic ubiquitin-like proteins, Pup and UBact, show no significant sequence homology with the ubiquitin sequence.

    The proteosome exists in prokaryoyes (some bacteria, all archaea), but it is much simpler, too. See also here:

    PROKARYOTIC UBIQUITIN-LIKE PROTEIN, PROTEASOMES, AND PATHOGENESIS

    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3662484/

    “In contrast to eukaryotic proteasomes, core particles from archaea and bacteria are far simpler structures with homo-heptameric rings of catalytic ? subunits flanked by homo-heptameric rings of ? subunits16–21 (Table 1, Fig. 2). To date, only bacteria found in the class Actinomycetes are known to have proteasomes19, 22–24. Multi-subunit regulatory complexes similar to those in eukaryotes have not been identified in prokaryotes, suggesting the mechanisms by which proteins are targeted for degradation are different, or that regulatory complex interactions with cores are transient or weak.”

  68. DATCG:

    “Indeed how is a correct code understood to be correct?
    After a billion, maybe trillion combinations?”

    That’s exactly the point. Codes cannot really emerge without design, because they essence is a symbolic relationship. There is absolute no way of getting them in other ways. There must be an understanding of meaning, just from the beginning.

    Codes are structures where something means something else, by arbitrary choices. They scream design and conscious understanding, even in the simplest forms.

    That’s why noe-darwinists have tried in all possible ways to derive the genetic code from ancient biochemical affinities, of course without succeeding. Their only hope is to demonstrate that the origin of the code is not symbolic, and that the mapping is not arbitrary.

    But that is simply not true, and they will never succeed.

  69. Dionisio:

    “Can this be considered a code issue too?”

    This is a difficult question. In brief, what I think is as follows:

    The bicoid gradient is certainly a signal, a signla that could bo cosncidered as a wave of concetrations in space (cell locations) and time (time windows).

    My only problem in considering it a coded signal is that bicoid apparently acts as a transcription factor, directly activationg other genes to get the desired results.

    I am more at ease with the concept of symbolic coding when the system includes some independent actor which reads and inteprets the signal, like ubiquitin-binding proteins, for example. TFs are more similar to final effectors.

    So, I would say that it is certainly a signal, but I would be cautious in stating that it is a coded signal.

  70. Two words: “information jump”

    🙂 🙂

  71. Upright BiPed:

    Our personalities are strangely merging one with the other! 🙂

  72. Upright Biped:

    “The second postulate, however, was that they (the group) would have nothing to do with any argument for design in biology. Never again throughout the paper is the design argument even mentioned, other than to kill it up front.”

    This is very sad.

  73. DATC at #56:

    Wonderful contribution.

    Autophagy by lysosomes is an important alternative degradation pathway, with different specificities compared to the proteasome pathway. It is mentioned in the table in the OP, as connected usually to K63 ubiquitinations, but it is absolutely appropriate that you have given more detail about this other fascinating outcome.

    I agree with you that, even if the Ubiquitin System seems to regulate, start and terminate the whole process (as detailed in Fig. 1), the most interesting part seems to be the role in selective autophagy, where the cargo is recognized and transported in a very specific way (as detailed in Fig. 2).

    A specially interesting issue is certainly mitophagy, the degradation of mitochondria.

    A central role in that process seems to be implemented by Parkin, an elusive E3 ligase whose mutations are known to cause a familial form of Parkinson’s disease known as autosomal recessive juvenile Parkinson’s disease (AR-JP).

    Parkin is a member of the RING-between-RING (RBR) family of E3 ligases, the least common class of these proteins. It has quite a lot of functions listed on its Uniprot page, from which I quote this very interesting statement:

    Mediates monoubiquitination as well as ‘Lys-6’, ‘Lys-11’, ‘Lys-48’-linked and ‘Lys-63’-linked polyubiquitination of substrates depending on the context (PubMed:19229105, PubMed:20889974, PubMed:25621951).

    Emphasis mine.

    It seems that context dependency is the rule, here. These processes are wonderful examples of Context-oriented Programming and Object-oriented Programming of the best kind! 🙂

  74. gpuccio @69:

    I see your point, which makes sense. Thanks.

    The final effect of the morphogen gradients, which are spatiotemporal signaling profiles, depends on many factors, like the location of the sources, the type of signaling molecules, the source activation/deactivation timing, the secretion rate, the transporting of the signaling molecules to their destination, their concentration distribution, the duration of the exposure of the target cells to the spatial concentrations of signaling molecules. But none of that seems associated with codes.
    Interestingly the control of such a fascinating component of morphogenesis is done directly though signals.

  75. DATCG, UB, gpuccio,

    Biosemiotics doesn’t seem to apply directly to spatiotemporal signaling profiles (a.k.a. morphogen gradients), which are important choreographic components of developmental biology and evo-devo.

    Is this correct?

    hanks.

  76. @75 correction

    Thanks

  77. Dio,

    I can’t answer your question just now. Using the word “signal” does indeed seem to indicate semiosis — I.e. a signal is a semiotic element within a system. But I would have to study the system in detail in order to answer the question with any sense of certainty on my part. But I have my hands full right now, and just cannot do that at this time. GP likely already knows the system and can tell you.

    ….my apologies

  78. Upright BiPed:

    Thank you for the Quipus! 🙂

    You are right, the way the ubiquitin signal is read in different cases is often only partially understood.

    For example, even in the case of the proteasome, which is certainly the most studied scenario, many things are stil elusive.

    See for example this very recent paper:

    Recognition of Client Proteins by the Proteasome.

    https://www.annualreviews.org/doi/full/10.1146/annurev-biophys-070816-033719#f1

    Abstract
    The ubiquitin proteasome system controls the concentrations of regulatory proteins and removes damaged and misfolded proteins from cells. Proteins are targeted to the protease at the center of this system, the proteasome, by ubiquitin tags, but ubiquitin is also used as a signal in other cellular processes. Specificity is conferred by the size and structure of the ubiquitin tags, which are recognized by receptors associated with the different cellular processes. However, the ubiquitin code remains ambiguous, and the same ubiquitin tag can target different proteins to different fates. After binding substrate protein at the ubiquitin tag, the proteasome initiates degradation at a disordered region in the substrate. The proteasome has pronounced preferences for the initiation site, and its recognition represents a second component of the degradation signal.

    (Paywall)

    Or this other one:

    Ubiquitin recognition by the proteasome.

    J Biochem. 2017 Feb 1;161(2):113-124. doi: 10.1093/jb/mvw091.

    Abstract
    The 26S proteasome is a 2.5-MDa complex responsible for the selective, ATP-dependent degradation of ubiquitylated proteins in eukaryotic cells. Substrates in hundreds cellular pathways are timely ubiquitylated and converged to the proteasome by direct recognition or by multiple shuttle factors. Engagement of substrate protein triggers conformational changes of the proteasome, which drive substrate unfolding, deubiquitylation and translocation of substrates to proteolytic sites. Recent studies have challenged the previous paradigm that Lys48-linked tetraubiquitin is a minimal degradation signal: in addition, monoubiquitylation or multiple short ubiquitylations can serve as the targeting signal for proteasomal degradation. In this review, I highlight recent advances in our understanding of the proteasome structure, the ubiquitin topology in proteasome targeting, and the cellular factors that regulate proteasomal degradation.

    Much is still to be understood.

  79. UB @77,

    gpuccio covered that @69. However, let’s leave this off topic issue which I mentioned for curiosity but it’s a distracting digression from the main discussion in this thread.

    Thanks.

  80. Is this just my perception?
    After this OP + follow up discussion, now the term “ubiquitin” seems to pop up in many places out there.
    Perhaps this thread has increased our sensitivity to detect that term?
    Or is it that the term is really appearing more often lately?
    Maybe both?
    Blame it on gpuccio!
    🙂

  81. Background
    Parkin (PARK2) is an E3 ubiquitin ligase that is commonly mutated in Familial Parkinson’s Disease (PD). In cell culture models, Parkin is recruited to acutely depolarised mitochondria by PINK1. PINK1 activates Parkin activity leading to ubiquitination of multiple proteins, which in turn promotes clearance of mitochondria by mitophagy. Many substrates have been identified using cell culture models in combination with depolarising drugs or proteasome inhibitors, but not in more physiological settings.

    Methods
    Here we utilized the recently introduced BioUb strategy to isolate ubiquitinated proteins in flies. Following Parkin Wild-Type (WT) and Parkin Ligase dead (LD) expression we analysed by mass spectrometry and stringent bioinformatics analysis those proteins differentially ubiquitinated to provide the first survey of steady state Parkin substrates using an in vivo model. We further used an in vivo ubiquitination assay to validate one of those substrates in SH-SY5Y cells.

    Results
    We identified 35 proteins that are more prominently ubiquitinated following Parkin over-expression. These include several mitochondrial proteins and a number of endosomal trafficking regulators such as v-ATPase sub-units, Syx5/STX5, ALiX/PDCD6IP and Vps4. We also identified the retromer component, Vps35, another PD-associated gene that has recently been shown to interact genetically with parkin. Importantly, we validated Parkin-dependent ubiquitination of VPS35 in human neuroblastoma cells.

    Conclusions
    Collectively our results provide new leads to the possible physiological functions of Parkin activity that are not overtly biased by acute mitochondrial depolarisation.

    Martinez Zarate, Aitor & Lectez, Benoit & Ramirez, Juanma & Popp, Oliver & D Sutherland, James & Urbé, Sylvie & Dittmar, Gunnar & J Clague, Michael & Mayor, Ugo. (2017). Quantitative proteomic analysis of Parkin substrates in Drosophila neurons. Molecular Neurodegeneration. 12. . 10.1186/s13024-017-0170-3.

  82. Dionisio:

    Seems quite a lot of work for one protein!

    Thank you for the quote. 🙂

  83. DATCG:

    Interesting paper. And the world “design” in the title is precious: is the neo-darwinist surveillance becoming more distracted? 🙂

    These three GLI proteins are interesting. They appear in vertebrates (while SHH is older). An interesting pattern of morphogens – TFs interacting.

  84. Checking in for updates and reading before heading out 🙂

    Thanks for all the expanded info Gpuccio. UB and Dio as well. Not sure when I’ll have time to review Pattee, but looking forward to it.

    #5 Gpuccio,
    Yes it raised eyebrows for me when I first read it. And the video clarified some more.

    A review of “unstable” in terms of thermodynamic stability and protein folding? Or reversible? For steady-state levels to be efficient and fast. Maybe if you have time.

    Question, is there a standard steady-state efficiency rate to maintain and how efficient must it be? If it’s even considered to be something important to track.

    We tend to see high efficiency in many functions, like ATP or photosynthesis, but a balancing act is a bit different so would we expect more flexibility too in this area? Or, am I covering the same area you already mentioned Gpuccio?

    If the balance between synthesis and degradation is not maintained, then accumulation of unstable proteins increase.

    I’m curious what happens at different levels of low and high production synthesis vs degradation and at what point does the balance tip over.

    Which suggest a next question, is there a master monitor reporting and/or more regulators of this balancing act.

    I’m getting off subject, but it reminds me of the intricacy of each element, “cargo,” tagging, signals and systems coordinating together.

  85. again from your original #5 post Gpuccio…

    At timepoint 2:39 of the first video there is already a very important point which deserves discussion: why proteins, and especially regulator proteins, need to be unstable so that adjustments in their steady state can be achieved quickly.

    It means that biological systems invest a lot of resources to achieve flexibility and quick (and very intelligent) adaptation to changing conditions.

    YES, So this “flexibility” must be “maintained” as such. Yet another area important for Monitoring, Alerts-Signals, Repair and critical function.

    So somehow “junk” dna and or genetic mutations are supposed to mutate stable and unstable proteins? Step by step that is for interaction among all these systems.

    I suppose that can be achieved because one of life’s features is to depend so critically on far from equilibrium processes.

    Food for thought.

    Really like how you summed it up in last sentence.

    Makes me think I need to replenish with healthy foods to maintain good balance, yet far from equilibrium 😉

    I came across open access Book CMMP – Condensed Matter and Materials Physics, a Chapter with interesting history and how physics has a role in Biology, a bit dated from 2007…

    What is the Physics of Life?>

    quote…

    “One of the entral problems faced by any organism is to transmit information reliably at the molecular level. This problem was phrased beautifully by Schrodinger in “What is Life?,” a series of lectures given(1943) in the wake of the discovery of () genetic information.”

    () edited

    Not recommending reading chapter. But there’s more on error processing I found in agreement.

    Life requires all the sciences and a multi-disciplinary approach to understand it far from-equilibrium.

    Also found this in Far From-Equilibrium Chapter…

    The Next Decade

    “… far from-equilibrium behavior is not rare but ‘ubiquitous’, occuring from the nanometer scale on up…”

    Could not resist 😉

    Haha, have a good day guys!

    .

  86. #83 Gpuccio,

    Haha, I thought so re: Design. But was interesting finding
    another code too. And noticed the Code Biology page includes it, along with another code discovered in 2014-2015.

    I’ve not looked at morphogens at all. Another day 🙂

  87. Dionisio at #80:

    So, ubiquitin is becoming ubiquitous. There is some logic in that.

    RV + NS can certainly explain that all! 🙂

  88. Dionisio, DATCG, Upright Biped:

    What about immunity, both innate and adaptive? What about T and B cells?

    Innate immunity:

    Multifaceted roles of TRIM38 in innate immune and inflammatory responses

    https://www.nature.com/articles/cmi201666

    (Paywall)

    The tripartite motif-containing (TRIM) proteins represent the largest E3 ubiquitin ligase family. The multifaceted roles of TRIM38 in innate immunity and inflammation have been intensively investigated in recent years. TRIM38 is essential for cytosolic RNA or DNA sensor-mediated innate immune responses to both RNA and DNA viruses, while negatively regulating TLR3/4- and TNF/IL-1?-triggered inflammatory responses. In these processes, TRIM38 acts as an E3 ubiquitin or SUMO ligase, which targets key cellular signaling components, or as an enzymatic activity-independent regulator. This review summarizes recent advances that highlight the critical roles of TRIM38 in the regulation of proper innate immune and inflammatory responses.

    TRIM38 is a RING E3 ligase, whose sequence is specially engineered in mammals.

    See also here:

    TRIM Family Proteins: Roles in Autophagy, Immunity, and Carcinogenesis.

    Abstract:
    Tripartite motif (TRIM) family proteins, most of which have E3 ubiquitin ligase activities, have various functions in cellular processes including intracellular signaling, development, apoptosis, protein quality control, innate immunity, autophagy, and carcinogenesis. The ubiquitin system is one of the systems for post-translational modifications, which play crucial roles not only as markers for degradation of target proteins by the proteasome but also as regulators of protein–protein interactions and of the activation of enzymes. Accumulating evidence has shown that TRIM family proteins have unique, important roles and that their dysregulation causes several diseases classified as cancer, immunological disease, or developmental disorders. In this review we focus on recent emerging topics on TRIM proteins in the regulation of autophagy, innate immunity, and carcinogenesis.

    Trends:

    Ubiquitination is catalyzed by the E1 ubiquitin-activating enzyme, E2 ubiquitin-conjugating enzymes, and E3 ubiquitin ligases, which are a critical component directly responsible for substrate recognition.

    Tripartite motif (TRIM) proteins, more than 80 being present in humans, are defined as a subfamily of the RING-type E3 ubiquitin ligase family.

    In autophagy several TRIM proteins function as platforms for the assembly of autophagy regulators and recognize targets such as ubiquitinated cargos via sequestosome-1-like receptors.

    Some TRIM proteins positively or negatively control many regulators including pattern recognition receptors, intracellular signal transducers and transcription factors in innate immunity.

    TRIM proteins are involved in a broad range of oncogenic processes including transcriptional regulation, cell proliferation, apoptosis, DNA repair, and metastasis.

    In plants?

    Conventional and unconventional ubiquitination in plant immunity.

    Abstract:
    Ubiquitination is one of the most abundant types of protein post-translational modification (PTM) in plant cells. The importance of ubiquitination in the regulation of many aspects of plant immunity has been increasingly appreciated in recent years. Most of the studies linking ubiquitination to the plant immune system, however, have been focused on the E3 ubiquitin ligases and the conventional ubiquitination that leads to the degradation of the substrate proteins by the 26S proteasome. By contrast, our knowledge about the role of unconventional ubiquitination that often serves as non-degradative, regulatory signal remains a significant gap. We discuss, in this review, the recent advances in our understanding of ubiquitination in the modulation of plant immunity, with a particular focus on the E3 ubiquitin ligases. We approach the topic from a perspective of two broadly defined types of ubiquitination in an attempt to highlight the importance, yet current scarcity, in our knowledge about the regulation of plant immunity by unconventional ubiquitination.

    More in next post.

  89. Dionisio, DATCG, Upright Biped:

    Adaptive immunity?

    Regulation of T helper cell differentiation by E3 ubiquitin ligases and deubiquitinating enzymes.

    (Paywall)

    Abstract:
    CD4 T cells are essential components of adaptive immunity and play a critical role in anti-pathogenic or anti-tumor responses as well as autoimmune and allergic diseases. Naive CD4 T cells differentiate into distinct subsets of T helper (Th) cells by various signals including TCR, costimulatory and cytokine signals. Accumulating evidence suggests that these signaling pathways are critically regulated by ubiquitination and deubiquitination, two reversible posttranslational modifications mediated by E3 ubiquitin ligases and deubiquitinating enzymes (DUBs), respectively. In this review, we briefly introduce the signaling pathways that control the differentiation of Th cells and then focused on the roles of E3s- and DUBs-mediated ubiquitin modification or demodification in regulating Th cell differentiation.

    See also here:

    The ubiquitin system in immune regulation.

    Abstract
    The ubiquitin system plays a pivotal role in the regulation of immune responses. This system includes a large family of E3 ubiquitin ligases of over 700 proteins and about 100 deubiquitinating enzymes, with the majority of their biological functions remaining unknown. Over the last decade, through a combination of genetic, biochemical, and molecular approaches, tremendous progress has been made in our understanding of how the process of protein ubiquitination and its reversal deubiquitination controls the basic aspect of the immune system including lymphocyte development, differentiation, activation, and tolerance induction and regulates the pathophysiological abnormalities such as autoimmunity, allergy, and malignant formation. In this review, we selected some of the published literature to discuss the roles of protein-ubiquitin conjugation and deubiquitination in T-cell activation and anergy, regulatory T-cell and T-helper cell differentiation, regulation of NF-?B signaling, and hematopoiesis in both normal and dysregulated conditions. A comprehensive understanding of the relationship between the ubiquitin system and immunity will provide insight into the molecular mechanisms of immune regulation and at the same time will advance new therapeutic intervention for human immunological diseases.

    And this is a very good and very recent review:

    Ubiquitin enzymes in the regulation of immune responses

    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5490640/

    (Public access)

    Abstract
    Ubiquitination plays a central role in the regulation of various biological functions including immune responses. Ubiquitination is induced by a cascade of enzymatic reactions by E1 ubiquitin activating enzyme, E2 ubiquitin conjugating enzyme, and E3 ubiquitin ligase, and reversed by deubiquitinases. Depending on the enzymes, specific linkage types of ubiquitin chains are generated or hydrolyzed. Because different linkage types of ubiquitin chains control the fate of the substrate, understanding the regulatory mechanisms of ubiquitin enzymes is central. In this review, we highlight the most recent knowledge of ubiquitination in the immune signaling cascades including the T cell and B cell signaling cascades as well as the TNF signaling cascade regulated by various ubiquitin enzymes. Furthermore, we highlight the TRIM ubiquitin ligase family as one of the examples of critical E3 ubiquitin ligases in the regulation of immune responses.

    Always for lovers of simplicity, I recommend Fig. 3, which covers Ubiquitin enzymes in the T Cell Receptor signaling pathway.

    The B cell pathway is also covered in Fig. 5.

    Fig. 6 is, again, about the NK-kB pathway, which is central in many immunity related processes.

    And Fig. 7 is about our old friend, TRIM.

  90. Dionisio, DATCG, Upright Biped:

    So, here is a brief list of the cellular processes we have touched in the OP and discussion:

    a) Proteasome degradation

    b) Autophagic degradation

    c) Mitophagy

    d) Cell signaling and transmission pathways

    e) Double-Strand Breaks DNA repair

    f) Neuronal regulation

    g) Regulation of innate immunity

    h) Regulation of adaptive immunity

    i) Regulation of T cell and B cell differentiation

    It’s a lot of stuff, I would say.

    And in each of those processes ubquitination has a major role.

    And in each of those processes different proteins, especially different 3 ligases, are involved, often many different ones at a time, each of them complex, each of them usually involved in multiple scenarios.

    Fascinating.

  91. Dionisio, DATCG, Upright Biped:

    Ah, I was forgetting. Our party is still very private.

    And contributions from the other side? OK, let’s me count them…

    Zero?

  92. Dionisio, DATCG, Upright Biped::

    Just a curiosity. Look at this 2016 paper:

    Identification of Top-ranked Proteins within a Directional Protein Interaction Network using the PageRank Algorithm: Applications in Humans and Plants

    http://www.caister.com/cimb/v/v20/13.pdf

    (Public access)

    I quote:

    High-ranking proteins in the human protein network

    The ubiquitin-protein ligase Casitas Blineage lymphoma (CBL) is ranked No. 1 in the reverse ranking, which means that CBL is the top-ranked regulator in the cell. This ligase is responsible for protein ubiquitination and is known to be involved in at least nine pathways, including the insulin signaling pathway (Figure 2). Mutation in the CBL gene has been implicated in a number of human cancers, including acute myeloid leukemia (Naramura et al., 2011).

    Eight proteins are among the top50-ranked proteins in all three ranking approaches (forward, reverse, and nondirectional) (Table 2). Those proteins are ACVR1, CDC42, RAC1, RAF1, RHOA, TGFBR1, TRAF2, and TRAF6 (see Table 2 for names). Each of these eight top-ranked proteins is involved in many pathways. Mutations of these eight proteins are implicated in many types of cancers and severe diseases.

    So, No 1 human protein (according to this classification, of course) is a RING E3 ubiquitin ligase.

    And look at the 8 proteins which are included in the top 50 in all three ranking approaches. Some are membrane receptors, or kinases. But two of them, TRAF2 and TRAF6 (TNF receptor-associated factors) are RING E3 ubiquitin ligases, too.

    In particular, they are both old friends: see the NF-kB section in the OP.

  93. #80 Gpuccio,

    I’m beginning to think these post deserve UD reserve space precisely because of the loaded content in your original post and how you lay it out for readers to observe and contemplate in the comments.

    That list is a a great example you put together. All can be expanded into deeper branches and sub-list.

    You included Neuronal Regulation above and a paper earlier. I briefly searched ubiquitin along with FAS – Fetal Alcohol Syndrome and Ethanol.

    After the tragedy involving Parkland, Florida students at Marjory Stoneman Douglas High School in Broward County and the shooter Nikolas Cruz, neural disorders were of interest.

    Some news reports speculated Cruz suffered from FAS. I don’t know if it’s been confirmed.

    I came across an interesting paper, but waited to post since there’s so much to read.

    Note: TLR4, ubiquitin-proteasome-pathway, autophagy-lysosome-pathway,

    Here’s link and a few quotes Gpuccio as it ties in with your points on 1st Video by Deshaies, starting at 2:39 mark.

    Indeed, many neurodegenerative diseases are characterized by protein misfolding and an abnormal accumulation and aggregation of specific proteins, which can result for deficient clearance systems, such as the ubiquitin-proteasome system (UPS) and the autophagy-lysosome pathway (ALP). Examples of these disorders are Alzheimer’s or Parkinson’s disease, which present an accumulation of aberrant proteins that produce a toxic neuronal effect, also called proteotoxicity.6

    takeaway from above quote:
    Delicate Balance, epigenetics, mutations harm, not enhance, resulting in “abnormal accumulation and aggregation of specific proteins.”

    Here we report that in vivo chronic ethanol treatment alters both UPS and ALP, leading to an accumulation of ubiquitinated proteins in the mice cerebral cortex. However, alcohol upregulates the immunoproteasome by activating the neuroimmune system. Consistently, we provide evidence that the effects of ethanol on proteolytic processes are mediated by innate immune receptor TLR4 signaling, as minimal changes in protein degradation pathways were observed in the cerebral cortex of ethanol-treated TLR4-knockout (KO) mice. These findings provide new insights into the mechanisms underlying ethanol-induced brain damage.

    Effects mediated by TLR4 knockout. But look at the accumulation again and delicate balance.

    Likewise, an immunohistochemical analysis of ubiquitinated proteins (Figure 1b) further demonstrated that chronic ethanol treatment promotes the accumulation of ubiquitinated proteins in the cerebral cortex and that this event is associated with the TLR4 function.

    off-balance

    In summary, the above results suggest that ethanol treatment reduces the 20S constitutive proteasome expression in the cerebral cortex and promotes the accumulation of poly-ubiquitinated proteins, while it also stimulates the production of proinflammatory cytokines, probably through TLR4 signaling, which, in turn, induce the activation of the immunoproteasome.

    Paper Link, Open Access 2014…
    TLR4 mediates the impairment of ubiquitin-proteasome and autophagy-lysosome pathways induced by ethanol treatment in brain

    .

  94. GP,

    I am going to have limited access for a few days, but there’s nothing substantial I can add to your excellent explanation of the Ubiquitin system. I rank this thread as one of the best to appear on UD in recent memory, and I haven’t even the slightest question as to why your opponents are absent from the conversation. Congratulations.

    Reading this OP and your comments, I think about all the layers of control. Control systems have a necessary dual role to play; where they first make variables physically possible, and then they specify variables among the alternatives. And of course, all of this is established by a simultaneous coordination among the constraints that bring the system into being. One doesn’t exist without the other. The constraints can’t persist without the coordination.

    I also think about your discussions on “information jumps”, and of how those jumps (from their very start) have had discoverable physical consequences. I’d like to pass on an extended quote (sorry about the length). This is 45 years old, and on substance, it stands up today and will be standing up tomorrow as well:

    To state my argument briefly, I would say that living matter is distinguished from non-living matter by its evolution in the course of time, and that this evolution depends on a degree of constraint in a physical system that enables records of past events to control its future behavior. I argue that the very concept of a record is classical in the same sense that a measurement is classical, both depending on dissipative constraints which reduce the number of alternative types of behavior available to the system.

    […]

    Life Depends on Coordinated Constraints

    In addition to the use of records, there is a second universal property of life which I regard as fundamental to our interpretation of physical laws, and that is the coordination of all biological activities by hierarchical controls. Many biologists do not regard the origin of coordinated or functional behavior in matter as a physical problem since they accept the theory of evolution in the form of survival of the fittest as a sufficient explanation. However, this evades the question of the origin of any level of coordinated activity where new functions appear. Specifically it evades the problem of the origin of life, that is, the origin of a minimal set of coordinated constraints which write and read records. This course of recorded evolution has continued to generate level upon level of coordinated, hierarchical constraints from the rules of the genetic code to the rules of the languages of man, and yet we have almost no evidence and hardly any theory of how even one of these control levels arose. For this reason it appears to me that the significant activities of matter and of the mind are separated by level upon level of integrated control hierarchies with the gulfs between each level still hidden by inscrutable mysteries.
    If we are to make any progress at all in confronting basic physical principles with the behavior of such hierarchical organisms, then we must begin at the lowest possible level. I have chosen the concept of decision-making to characterize the basic function of a hierarchical control process. I want to consider the simplest examples of decision¬making in physical terms in order to see what problems arise. Decision-making is, of course, the principle function of the brain, but that does not mean that the essential physics of the brain’s function is best studied by looking at such a complex structure.

    […]

    The Language Problem

    I cannot imagine any answer to this type of question about the meaning or interpretation of symbolic processes without presupposing some form of generalized language structure. I am thinking now of language in the broadest possible sense, including not only the highly evolved abstract languages of man and the much more primitive genetic code, but any coordinated set of constraints or rules by which classes of physical structures are trans¬formed into specific actions or events. The essential condition for a language is the coordination of its rules, not in the choice of particular rules which generally appear arbitrary. The concept of meaning and interpretation for symbols does not make any sense when applied to single structures, but only to the relations between structures. For this reason I would say that a single decision or record isolated from a set of constraints which can transform classes of such decisions or records into a coordinated activity does not have a meaning or interpretation.

    […]

    The Origin Problem

    We are led then to our second problem — the physical basis of coordinated constraints which read and write records. Such a coordinated set of constraints we call a generalized language structure. The most universal example of such a language structure is the genetic writing and reading system in which the genetic coding constraints provide the essential read out transformations. What is perhaps most striking about this highly coordinated set of constraints is that it forms the basis for all levels of biological evolution over as long a time span as we can find data, and yet there is no evidence that this set of constraints has itself undergone any significant change. We therefore have theories of biological evolution based on the preexistence of this genotype-phenotype code, but no idea of how this coordinated set of constraints came into existence (e.g., Crick, 1968; Orgel, 1968).

    This mysterious origin problem is not limited to the genetic code, but is characteristic of all new levels of hierarchical control where a new set of coordinated constraints forms a new language structure which can make decisions about the alternative behavior of the level below. The problem of the origin and nature of coordinated constraints which effectively interpret records and make decisions is therefore a universal problem of all life. Alternatively, we could say that the most fundamental function of coordination in biology is to establish generalized language constraints which allow structure at one level to be interpreted as descriptions and executed as decisions at the higher level. It is in this sense we say that a language system is necessary to establish and execute hierarchical controls (Pattee, 1971 b).

    H.H. Pattee, 1973

  95. DATCG at #85:

    Protein stability and protein half-life seem to be a very complex issue.

    Portein half life in human cells seems to vary a lot. Here is a global proteome analysis:

    A Quantitative Spatial Proteomics Analysis of Proteome Turnover in Human Cells

    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3316722/

    Fig. 3 sums up some important results. In the 10% proteins with fastest turnover, half-life seems to vary from almost 0 to about 8 hours. A gross distribution of protein classes in the two extreme groups (slowest and fastest 10%) shows interesting differences. I quote from the paper:

    Functional annotation clustering of gene ontology terms for the fastest and slowest turnover rates showed specific enrichments of proteins with similar functions or characteristics (26, 27) (Fig. 3). The slowest turnover proteins have a wide range of functions. However, most are either present in large, abundant and stable protein complexes, such as ribosome and spliceosome subunits, RNA polymerase II, the nuclear pore, the exosome and the proteasome, or else are mitochondrial (Fig. 3, top). In contrast, many proteins with a faster than average turnover are involved in either mitosis, or other aspects of cell cycle regulation (Fig. 3, bottom). This includes protein components of the centromere, proteins with microtubule motor activity, proteins involved in cytoskeleton reorganization and proteins involved in chromatin assembly and condensation.

    This study was performed on HeLa cells, so things could be different in normal cells in vivo.

    An interesting paper is the following:

    Intrinsically Disordered Segments Affect Protein Half-Life in the Cell and during Evolution

    https://www.sciencedirect.com/science/article/pii/S2211124714006391

    Summary
    Precise control of protein turnover is essential for cellular homeostasis. The ubiquitin-proteasome system is well established as a major regulator of protein degradation, but an understanding of how inherent structural features influence the lifetimes of proteins is lacking. We report that yeast, mouse, and human proteins with terminal or internal intrinsically disordered segments have significantly shorter half-lives than proteins without these features. The lengths of the disordered segments that affect protein half-life are compatible with the structure of the proteasome. Divergence in terminal and internal disordered segments in yeast proteins originating from gene duplication leads to significantly altered half-life. Many paralogs that are affected by such changes participate in signaling, where altered protein half-life will directly impact cellular processes and function. Thus, natural variation in the length and position of disordered segments may affect protein half-life and could serve as an underappreciated source of genetic variation with important phenotypic consequences.

    This is very interesting, because if we (reasonably) assume that a shorter half-life is often related to critical regulatory functions, then it seems that intrinsically disordered segments, in particular at the N-terminus, could be also related to regulatory function.

    Which is an idea that I would very happily endorse! 🙂

  96. UB @94,

    Interesting assessment of GP’s latest OP and follow-up comments.

    Thanks.

  97. DATCG @93:

    That’s an interesting paper related to the effects of human behavior on the complex ubiquitin proteasome that GP has presented to us here.

    Thanks.

  98. DATCG @93:

    Another potential association of the ubiquitin proteasome with serious health-related issues:

    The cellular quality control system degrades abnormal or misfolded proteins and consists of three different mechanisms: the ubiquitin proteasomal system (UPS), autophagy and molecular chaperones. Any disturbance in this system causes proteins to accumulate, resulting in neurodegenerative diseases such as amyotrophic lateral sclerosis, Alzheimer’s disease (AD), Parkinson’s disease, Huntington’s disease and prion or polyglutamine diseases. Alzheimer’s disease is currently one of the most common age-related neurodegenerative diseases. However, its exact cause and pathogenesis are unknown. Currently approved medications for AD provide symptomatic relief; however, they fail to influence disease progression. Moreover, the components of the cellular quality control system represent an important focus for the development of targeted and potent therapies for managing AD. This review aims to evaluate whether existing evidence supports the hypothesis that UPS impairment causes the early pathogenesis of neurodegenerative disorders. The first part presents basic information about the UPS and its molecular components. The next part explains how the UPS is involved in neurodegenerative disorders. Finally, we emphasize how the UPS influences the management of AD. This review may help in the design of future UPS-related therapies for AD.

    Gadhave, Kundlik & Bolshette, Nityanand & Ahire, Ashutosh & Pardeshi, Rohit & Thakur, Krishan & Trandafir, Cristiana & Istrate, Alexandru & Ahmed, Sahabuddin & Lahkar, Mangala & Muresanu, Dafin & Balea, Maria. (2016). The ubiquitin proteasomal system: A potential target for the management of Alzheimer’s disease. Journal of Cellular and Molecular Medicine. 20. 1392-407. 10.1111/jcmm.12817.

  99. DATCG @93:

    Another potential association of the ubiquitin proteasome with serious health-related issues:

    Neurodegenerative diseases (NDDs) such as Alzheimer’s disease, Parkinson’s disease and Huntington’s disease (HD), amyotrophic lateral sclerosis, and prion diseases are all characterized by the accumulation of protein aggregates (amyloids) into inclusions and/or plaques. The ubiquitous presence of amyloids in NDDs suggests the involvement of disturbed protein homeostasis (proteostasis) in the underlying pathomechanisms. This review summarizes specific mechanisms that maintain proteostasis, including molecular chaperons, the ubiquitin-proteasome system (UPS), endoplasmic reticulum associated degradation (ERAD), and different autophagic pathways (chaperon mediated-, micro-, and macro-autophagy). The role of heat shock proteins (Hsps) in cellular quality control and degradation of pathogenic proteins is reviewed. Finally, putative therapeutic strategies for efficient removal of cytotoxic proteins from neurons and design of new therapeutic targets against the progression of NDDs are discussed.

    Penke, Botond & Bogár, Ferenc & Crul, Tim & Sántha, Miklós & Tóth, Melinda & Vígh, László. (2018). Heat Shock Proteins and Autophagy Pathways in Neuroprotection: From Molecular Bases to Pharmacological Interventions. International Journal of Molecular Sciences. 19. 325. 10.3390/ijms19010325.

    http://www.mdpi.com/1422-0067/19/1/325/pdf

    [emphasis added]

  100. gpuccio @95:

    This is very interesting, because if we (reasonably) assume that a shorter half-life is often related to critical regulatory functions, then it seems that intrinsically disordered segments, in particular at the N-terminus, could be also related to regulatory function.

    Which is an idea that I would very happily endorse!

    Who wouldn’t?

    🙂

  101. #94 UB,

    Here Hear, Gpuccio’s done another great post.

    Thanks for your comments and additional sources as well on
    Pattee. I’ve enjoyed reading the snippets and quotes you’ve
    posted. And look more to reading about and from him in
    future.

  102. Dionisio,
    thanks, the amount of health issues is as
    endless as ubiquitin appears to be in it’s arrangements
    and coordinations.

    I’ve saved ncbi searches on this for now to go back and
    review.

  103. #95 Gpuccio, thank you, will follow up on reading these
    as well!

    Please do not make another major post for another few weeks
    😉 haha, as I need time to consume this vast information
    ubiquitin proteosome system, DUBs and more intricate
    details.

  104. DATCG,
    Let me repeat the comment that was posted 5 days ago @21:

    “This thread has too much bad news for the ‘modern synthesis’ and the ‘third way’ clubs.”

  105. DATCG @103,

    I can chew the information poured into this thread, but can’t digest it completely, not even close. There’s much more than I can handle, considering that practically everyday new data are thrown in the bucket. This is insane. And we ain’t see nothin’ yet. The most fascinating discoveries are still ahead.

  106. gpuccio @95:

    This is very interesting, because if we (reasonably) assume that a shorter half-life is often related to critical regulatory functions, then it seems that intrinsically disordered segments, in particular at the N-terminus, could be also related to regulatory function.
    Which is an idea that I would very happily endorse!

    Apparently you’re right on this:

    Late embryogenesis abundant (LEA) proteins are related to cellular dehydration tolerance. Most LEA proteins are predicted to have no stable secondary structure in solution, i.e., to be intrinsically disordered proteins (IDPs), but they may acquire ?-helical structure upon drying. In the model plant Arabidopsis thaliana, the LEA proteins COR15A and COR15B are highly induced upon cold treatment and are necessary for the plants to attain full freezing tolerance. Freezing leads to increased intracellular crowding due to dehydration by extracellular ice crystals. In vitro, crowding by high glycerol concentrations induced partial folding of COR15 proteins. Here, we have extended these investigations to two related proteins, LEA11 and LEA25. LEA25 is much longer than LEA11 and COR15A, but shares a conserved central sequence domain with the other two proteins. We have created two truncated versions of LEA25 (2H and 4H) to elucidate the structural and functional significance of this domain. Light scattering and CD spectroscopy showed that all five proteins were largely unstructured and monomeric in dilute solution. They folded in the presence of increasing concentrations of trifluoroethanol and glycerol. Additional folding was observed in the presence of glycerol and membranes. Fourier transform infra red spectroscopy revealed an interaction of the LEA proteins with membranes in the dry state leading to a depression in the gel to liquid-crystalline phase transition temperature. Liposome stability assays revealed a cryoprotective function of the proteins. The C- and N-terminal extensions of LEA25 were important in cryoprotection, as the central domain itself (2H, 4H) only provided a low level of protection.

    Folding of intrinsically disordered plant LEA proteins is driven by glycerol-induced crowding and the presence of membranes.
    Bremer A1, Wolff M2, Thalhammer A2, Hincha DK
    FEBS J. 2017 Mar;284(6):919-936.
    doi: 10.1111/febs.14023.

  107. gpuccio @95:

    This is very interesting, because if we (reasonably) assume that a shorter half-life is often related to critical regulatory functions, then it seems that intrinsically disordered segments, in particular at the N-terminus, could be also related to regulatory function.
    Which is an idea that I would very happily endorse!

    Apparently you’re right on this:

    The emergence of intrinsically disordered proteins (IDPs) has challenged the classical protein structure-function paradigm by introducing a new paradigm of “coupled binding and folding”. This paradigm suggests that IDPs fold upon binding to their partners. Further studies, however, revealed a novel and previously unrecognized phenomenon of “uncoupled binding and folding” suggesting that IDPs do not necessarily fold upon interaction with their lipid and protein partners. The complex and often unusual biophysics of IDPs makes structural characterization of these proteins and their complexes not only challenging but often resulting in opposite conclusions. For this reason, some crucial questions in this field remain unsolved for well over a decade. Considering an important role of IDPs in cellular regulation, signaling and control in health and disease, more efforts are needed to solve these mysteries. Here, I focus on two long-standing contradictions in the literature concerning dimerization and membrane-binding activities of IDPs. Molecular explanation of these discrepancies is provided. I also demonstrate how resolution of these critical issues in the field of IDPs results in our expanded understanding of cell function and has multiple applications in biology and medicine.

    Structural biology of intrinsically disordered proteins: Revisiting unsolved mysteries.
    Sigalov AB1.
    Biochimie. 2016 Jun;125:112-8.
    doi: 10.1016/j.biochi.2016.03.006.

  108. gpuccio @95:

    This is very interesting, because if we (reasonably) assume that a shorter half-life is often related to critical regulatory functions, then it seems that intrinsically disordered segments, in particular at the N-terminus, could be also related to regulatory function.
    Which is an idea that I would very happily endorse!

    Apparently you’re right on this:

    Intrinsically disordered proteins (IDPs) and intrinsically disordered protein regions (IDPRs) are important constituents of many protein complexes, playing various structural, functional, and regulatory roles. In such disorder-based protein complexes, functional disorder is used both internally (for assembly, movement, and functional regulation of the different parts of a given complex) and externally (for interactions of a complex with its external regulators). In complex assembly, IDPs/IDPRs serve as the molecular glue that cements complexes or as highly flexible scaffolds. Disorder defines the order of complex assembly and the ability of a protein to be involved in polyvalent interactions. It is at the heart of various binding mechanisms and interaction modes ascribed to IDPs. Disorder in protein complexes is related to multifarious applications of induced folding and induced functional unfolding, or defines the entropic chain activities, such as stochastic machines and binding rheostats. This review opens a FEBS Letters Special Issue on Dynamics, Flexibility, and Intrinsic Disorder in protein assemblies and represents a brief overview of intricate roles played by IDPs and IDPRs in various aspects of protein complexes.

    The multifaceted roles of intrinsic disorder in protein complexes.
    Uversky VN1
    FEBS Lett. 2015 Sep 14;589(19 Pt A):2498-506.
    doi: 10.1016/j.febslet.2015.06.004.

  109. gpuccio @95:

    This is very interesting, because if we (reasonably) assume that a shorter half-life is often related to critical regulatory functions, then it seems that intrinsically disordered segments, in particular at the N-terminus, could be also related to regulatory function.
    Which is an idea that I would very happily endorse!

    Apparently you’re right on this:

    Intrinsically disordered proteins (IDPs) are an emerging phenomenon. They may have a high degree of flexibility in their polypeptide chains, which lack a stable 3D structure. Although several biological functions of IDPs have been proposed, their general function is not known. The only finding related to their function is the genetically conserved YSK2 motif present in plant dehydrins. These proteins were shown to be IDPs with the YSK2 motif serving as a core region for the dehydrins’ cryoprotective activity. Here we examined the cryoprotective activity of randomly selected IDPs toward the model enzyme lactate dehydrogenase (LDH). All five IDPs that were examined were in the range of 35–45 amino acid residues in length and were equally potent at a concentration of 50 ?g/mL, whereas folded proteins, the PSD-95/Dlg/ZO-1 (PDZ) domain, and lysozymes had no potency. We further examined their cryoprotective activity toward glutathione S-transferase as an example of the other enzyme, and toward enhanced green fluorescent protein as a non-enzyme protein example. We further examined the lyophilization protective activity of the peptides toward LDH, which revealed that some IDPs showed a higher activity than that of bovine serum albumin (BSA). Based on these observations, we propose that cryoprotection is a general feature of IDPs. Our findings may become a clue to various industrial applications of IDPs in the future.

    Matsuo, Naoki & Goda, Natsuko & Shimizu, Kana & Fukuchi, Satoshi & Ota, Motonori & Hiroaki, Hidekazu. (2018). Discovery of Cryoprotective Activity in Human Genome-Derived Intrinsically Disordered Proteins. International Journal of Molecular Sciences. 19. 401. 10.3390/ijms19020401.

    Maybe a topic for a future OP?

  110. Gpuccio, re: IDPs IDS’s, IDRs…

    After reviwing your post and comments, I found a video on “Intrinsically Disordered Proteins and “unstructural biology” aquiring momentum over last decade. In it, the presenter shows “protein structure-function paradigm” over last 65 years has solved over 100,000 structures.

    That’s great progress!

    But, enter IDPs, gaining momentum. I refer back to your comments in #10…

    “The theme of intrinsically disordered regions in proteins is becoming ever more relevant.”

    and at #95

    “This is very interesting, because if we (reasonably) assume that a shorter half-life is often related to critical regulatory functions, then it seems that intrinsically disordered segments, in particular at the N-terminus, could be also related to regulatory function.”

    Thanks for your patience Gpuccio as I catch up and work through this.
    If I’m not on right track, let me know. Glad for insight or corrections.

    Old Paradigm – What was and still is:
    Protein Seq A -> Function A
    Protein Seq B -> Function B
    Protein Seq C -> Function C …

    vs new research emphasis on Dynamic Model:

    /—————————————-> Function A… xyz
    Protein Sequence —> Dynamic Object —> Function B… xyz
    —————————————-> Function C… xyz

    This may be do to requirements for analog response? Not just digital. Like Heat Shock(or threshold stress levels)? That Dionioso quoted from another paper.

    From a Design perspective – flexibility in a controlled format.

    As feedback Dials up, Dials down, or rapid-response, quick thresholds, etc., signals and changing informational content carried from input processes of
    internal or external demands to a real-time decision tree process.

    Otherwise, as the presenter raised, how could it be handled?

    How many rigid objects would escalate in number?

    Because system demands for rapid interactions require response to survive
    or not be negativealy impacted.

    W/o Dynamic Objects how large or how inefficient is cost of maintenance?
    Just basic production and navigation alone?

    Strict rules importantly require strict enforcement. If Object is not

    well defined the “compiler” or translation, transcription will not accept it

    for specific uses.

    I brought up a Rules Based logic some time ago utilizing Conditional
    Processing. Can’t remember how far back, but as a programmer, I recognized
    it immediately. Rules based works wonderfully with Variable Information
    Reformatting btw, allowing flexibility like IDP’s, etc., including
    reading in of real-time changes dependent upon known or unknown input
    variables.

    There is a purpose for strict, one-way ticket rule that must be
    invoked, or rejected if not right.

    But, how many Protein Sequences for Life if only route is a strict
    Protein Sequence -> Function? W/o Dynamic Objects?

    As an observation, only recently in 2010 did Microsoft introduce Dynamic

    Object function in C# as a comparitive analogy. Not sure it’s a perfect
    analogy in all substantiative cases.

    But, by Design after all these years Microsoft added greater flexibility for coders at Run-Time. Enabling less code for more dynamic responses.
    More efficient for Coders.

    Note: Not verified efficiency of memory and processing run times. Obviously
    there are trade-offs on management of size, efficiency, response time.
    Design must take these into consideration. Materialist evolution only
    cannot. It creates vast amounts of “junk” DNA, right?

    I want to also recognize Dionisio’s efforts in past to highlight

    Object-Oriented Code reference frame.

    As you, UB and Dionioso have emphasized; semiosis, symbols and Code allow such verstailtiy.

    This allows independent response w/o need for building another protein
    sequence. Keeping proteins at resonable number.

    Hmmmm… I’m tempted to use a well known innovators in shipping and receiving, not just FedEx but in product distribution. Amazon.

    Input -> Dynamic Object -> Signal Out -> A million possible outcomes with final location and method for specific time-dependent delivery.

    Which brings us back to Gpuccio’s discussion and UB’s highlighting of

    “Information Jump”

    Under archaea and bacteria IDPs and IDRs are below 5%

    In Eukaryotes – IDP jumps to 20%, 30% for IDR
    In humans – IDR jumps to 35%

    .

  111. Dionioso @105, no kidding, I need a NCBI wireless jack installed 😉

    It’s a bit like The Matrix as Neo sees all the Data explosion of information but has not quite grasped how to use it. Before he mind bends the matrix, lol.

    Upload Ubiquitin Code – Flash! Upload IDP,IDR,IDS! Flash!

  112. DATCG,

    as result of reductionist reverse-engineering approach to research, what we’re seeing in the scientific literature is like the cacophony produced by the orchestra musicians tuning their individual instruments separately… the actual symphony hasn’t started yet… the biological ballet choreography, with all its colorful splendor, hasn’t been presented yet… it’s all ahead. Just get ready to see more fascinating things in the future. Biology-related research is moving at a very accelerated pace.

  113. The innate antiviral response is integral in protecting the host against virus infection. Many proteins regulate these signaling pathways including ubiquitin enzymes. The ubiquitin-activating (E1), -conjugating (E2), and -ligating (E3) enzymes work together to link ubiquitin, a small protein, onto other ubiquitin molecules or target proteins to mediate various effector functions. The tripartite motif (TRIM) protein family is a group of E3 ligases implicated in the regulation of a variety of cellular functions including cell cycle progression, autophagy, and innate immunity. Many antiviral signaling pathways, including type-I interferon and NF-?B, are TRIM-regulated, thus influencing the course of infection. Additionally, several TRIMs directly restrict viral replication either through proteasome-mediated degradation of viral proteins or by interfering with different steps of the viral replication cycle. In addition, new studies suggest that TRIMs can exert their effector functions via the synthesis of unconventional polyubiquitin chains, including unanchored (non-covalently attached) polyubiquitin chains. TRIM-conferred viral inhibition has selected for viruses that encode direct and indirect TRIM antagonists. Furthermore, new evidence suggests that the same antagonists encoded by viruses may hijack TRIM proteins to directly promote virus replication. Here, we describe numerous virus-TRIM interactions and novel roles of TRIMs during virus infections.

    Tol, Sarah & Hage, Adam & Isabel Giraldo, Maria & Bharaj, Preeti & Rajsbaum, Ricardo. (2017). The TRIMendous role of TRIMs in virus-host interactions. Vaccines. 5. 23. 10.3390/vaccines5030023.

  114. Sparrer, Konstantin & Gack, Michaela. (2018). TRIM proteins: New players in virus-induced autophagy. PLOS Pathogens. 14. e1006787. 10.1371/journal.ppat.1006787.

    http://journals.plos.org/plosp.....=printable

  115. The ubiquitin-proteasome system (UPS) ensures regulation of the protein pool in the cell by ubiquitination of proteins followed by their degradation by the proteasome. It plays a central role in the cell under normal physiological conditions as well as during viral infections. On the one hand, the UPS can be used by the cell to degrade viral proteins, thereby restricting the viral infection. On the other hand, it can also be subverted by the virus to its own advantage, notably to induce degradation of cellular restriction factors. This makes the UPS a central player in viral restriction and counter-restriction. In this respect, the human immunodeficiency viruses (HIV-1 and 2) represent excellent examples. Indeed, many steps of the HIV life cycle are restricted by cellular proteins, some of which are themselves components of the UPS. However, HIV itself hijacks the UPS to mediate defense against several cellular restriction factors. For example, the HIV auxiliary proteins Vif, Vpx and Vpu counteract specific restriction factors by the recruitment of cellular UPS components. In this review, we describe the interplay between HIV and the UPS to illustrate its role in the restriction of viral infections and its hijacking by viral proteins for counter-restriction.

    Seissler, Tanja & Marquet, Roland & Paillart, Jean-Christophe. (2017). Hijacking of the Ubiquitin/Proteasome Pathway by the HIV Auxiliary Proteins. Viruses. 9. 10.3390/v9110322.

  116. .

  117. grrr, having network issues. Where’s ubiguitin flexibility and IDP when you need it! 😉

    Dionisio,
    Yes, agree. I came across another video, actually a
    commercial for products, but found it interesting highlight
    of the direction we’re heading.

    I think often times real life business undercuts ideological differences by neo-Darwinist…

    https://www.youtube.com/watch?v=ICz49GY24mI

    can skip to minute 1:14

    Look how fast the field and publishing is growing since first discovery up to 2014, time of video.

    Also, nitpicking, but I hate the naming conventions,

    Disorder?

    There’s no disorder here and it’s confusing nomenclature in my opinion.

    Flexible, Differential sure, but not disorder. It’s almost like calling unknown gemonic data – “junk” which is no longer junk.

    Should be renamed I think. I get that they were naming it as an opposite function to “ordered” or Fixed 3D structures.

    But really? Disordered? Why not call them IFPs?

    Intrinsic Flexible Proteins, or Conditional Proteins, or Relaxed Proteins.

    No, instead, unsuspected flexible, conditional order is called disorder.

    Sigh. The naming conventions in biology, genetics, etc., have long suffered. It’s bothered me since my first biology class and only seems to be more mired today in same illogical naming conventions.

    LOL, maybe too nitpicky, but I think half the problem with it is making logical naming conventions, also open it up to more understanding and people in general.

    .

  118. Gpuccio #95, thank you for this…

    Distribution of protein turnover

    “Fig 3 Proteins were sorted on the x axis from fastest to slowest turnover and represented as a scatter plot with the 50% protein turnover value on the y axis. Approximately 60% (blue lines) of the HeLa proteins show a 50% turnover”

    .

  119. Most TRIM proteins contain E3 ubiquitin ligase activity, a class of enzymes which catalyze the final step (E3 step) in the ubiquitination cascade to form an ubiquitin covalent bond with a substrate lysine.

    […] how zinc impacts MG53 E3-ubiquitin ligase activity and the identities of the E3-ligase substrates involved in MG53-mediated wound healing remain elusive and requires further investigation.

    Wound care is a major healthcare expenditure. Treatment of burns, surgical and trauma wounds, diabetic lower limb ulcers and skin wounds is a major medical challenge with current therapies largely focused on supportive care measures. Successful wound repair requires a series of tightly coordinated steps including coagulation, inflammation, angiogenesis, new tissue formation and extracellular matrix remodelling. Zinc is an essential trace element (micronutrient) which plays important roles in human physiology. Zinc is a cofactor for many metalloenzymes required for cell membrane repair, cell proliferation, growth and immune system function. The pathological effects of zinc deficiency include the occurrence of skin lesions, growth retardation, impaired immune function and compromised would healing. Here, we discuss investigations on the cellular and molecular mechanisms of zinc in modulating the wound healing process. Knowledge gained from this body of research will help to translate these findings into future clinical management of wound healing.

    Lin, Peihui & Sermersheim, Matthew & Li, Haichang & Lee, Peter & Steinberg, Steven & Ma, Jianjie. (2017). Zinc in Wound Healing Modulation. Nutrients. 10. 16. 10.3390/nu10010016.

  120. DATCG,

    Thanks for the link to the interesting video.

    Here’s another video:

    https://www.youtube.com/embed/wqdlET1_SQM

  121. Background
    TRIM25 is a novel RNA-binding protein and a member of the Tripartite Motif (TRIM) family of E3 ubiquitin ligases, which plays a pivotal role in the innate immune response. However, there is scarce knowledge about its RNA-related roles in cell biology. Furthermore, its RNA-binding domain has not been characterized.

    Results
    Here, we reveal that the RNA-binding activity of TRIM25 is mediated by its PRY/SPRY domain, which we postulate to be a novel RNA-binding domain. Using CLIP-seq and SILAC-based co-immunoprecipitation assays, we uncover TRIM25’s endogenous RNA targets and protein binding partners. We demonstrate that TRIM25 controls the levels of Zinc Finger Antiviral Protein (ZAP). Finally, we show that the RNA-binding activity of TRIM25 is important for its ubiquitin ligase activity towards itself (autoubiquitination) and its physiologically relevant target ZAP.

    Conclusions
    Our results suggest that many other proteins with the PRY/SPRY domain could have yet uncharacterized RNA-binding potential. Together, our data reveal new insights into the molecular roles and characteristics of RNA-binding E3 ubiquitin ligases and demonstrate that RNA could be an essential factor in their enzymatic activity.

    Electronic supplementary material
    The online version of this article (doi:10.1186/s12915-017-0444-9) contains supplementary material, which is available to authorized users.

    Choudhury, Nila & Heikel, Gregory & Trubitsyna, Maryia & Kubik, Peter & Nowak, Jakub & Webb, Shaun & Granneman, Sander & Spanos, Christos & Rappsilber, Juri & Castello, Alfredo & Michlewski, Gracjan. (2017). RNA-binding activity of TRIM25 is mediated by its PRY/SPRY domain and is required for ubiquitination. BMC Biology. 15. 10.1186/s12915-017-0444-9.

  122. RNA-binding proteins (RBPs) are typically thought of as proteins that bind RNA through one or multiple globular RNA-binding domains (RBDs) and change the fate or function of the bound RNAs. Several hundred such RBPs have been discovered and investigated over the years. Recent proteome-wide studies have more than doubled the number of proteins implicated in RNA binding and uncovered hundreds of additional RBPs lacking conventional RBDs. In this Review, we discuss these new RBPs and the emerging understanding of their unexpected modes of RNA binding, which can be mediated by intrinsically disordered regions, protein–protein interaction interfaces and enzymatic cores, among others. We also discuss the RNA targets and molecular and cellular functions of the new RBPs, as well as the possibility that some RBPs may be regulated by RNA rather than regulate RNA.

    W. Hentze, Matthias & Castello, Alfredo & Schwarzl, Thomas & Preiss, Thomas. (2018). A brave new world of RNA-binding proteins. Nature Reviews Molecular Cell Biology. 10.1038/nrm.2017.130.

  123. DATCG at #117:

    Beautiful video, than you.

    OK, I suppose we must accept the nomenclature as it is, but your proposals are very good. I would definitely like “Intrinsically Flexible Proteins”! 🙂

    My interest for Intrinsically Disordered, Proteins, and in particular Intrinsically Disordered regions, started some time ago with my interest in Transcription Factors.

    Indeed, testing TFs for their evolutionary conservation, I realized that most of them can easily be considered as the sum of two components:

    1) One (or more) DNA binding region: this is the domain part, which is almost always highly conserved, and shared between many different TFs.

    2) One (or more) region, often much longer than the DNA binding region, where no recognizable domains can usally be found. This part has variable evolutionary conservation.

    IOWs, this is more or less the same pattern we observe in E3 ligases, and in many regulatory proteins, while in traditional “effector” proteins we can usually observe a more striking prevalence of recognizable domains.

    In TFs, moreover, this pattern is remarkably constant.

    So, it is perfectly reasonable to assume that the “non domain” part is the one important in regulation issues, and in particular in flexible protein-protein interactions.

    This was the idea behind my first OPs about information jumps, where I focused on Prickle1 protein, showing that the the “non domain” part was the one with the big information jump in vertebrates:

    https://uncommondescent.com/intelligent-design/homologies-differences-and-information-jumps/

    and that the “non domain part” was highly but differentially conserved in different groups of animals, as consistent with different and specific funations:

    https://uncommondescent.com/intelligent-design/information-jumps-again-some-more-facts-and-thoughts-about-prickle-1-and-taxonomically-restricted-genes/

    It seems that my ideas about “non domain” parts, which of course are usually called now “Intrinsically Disordered Regions”, has become rather fashionable.

    For example, this recent enough paper seems to be exactly about that concept as applied to TFs:

    Targeting protein-protein interactions (PPIs) of transcription factors: Challenges of intrinsically disordered proteins (IDPs) and regions (IDRs).

    https://www.sciencedirect.com/science/article/pii/S0079610715000796?via%3Dihub

    Unfortunately, it is not public access, and I could not access the full text, but the brief abstract is already telling:

    Abstract:

    In this review we discuss recent progress in targeting the protein–protein interactions made by oncogenic transcription factors. We particularly focus on the challenges posed by the prevalence of intrinsically disordered regions in this class of protein and the strategies being used to overcome them.

  124. Dionisio, DATCG, Upright Biped:

    I have just found this very, very interesting and very recent paper. It seems to add beautifully to the ideas expressed in my previous comment at #123:

    Evidence for a Strong Correlation Between Transcription Factor Protein Disorder and Organismic Complexity

    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5434936/

    And it is public access! 🙂

    Abstract:

    Studies of diverse phylogenetic lineages reveal that protein disorder increases in concert with organismic complexity but that differences nevertheless exist among lineages. To gain insight into this phenomenology, we analyzed all of the transcription factor (TF) families for which sequences are known for 17 species spanning bacteria, yeast, algae, land plants, and animals and for which the number of different cell types has been reported in the primary literature. Although the fraction of disordered residues in TF sequences is often moderately or poorly correlated with organismic complexity as gauged by cell-type number (r2?<?0.5), an unbiased and phylogenetically broad analysis shows that organismic complexity is positively and strongly correlated with the total number of TFs, the number of their spliced variants and their total disordered residues content (r2?>?0.8). Furthermore, the correlation between the fraction of disordered residues and cell-type number becomes stronger when confined to the TF families participating in cell cycle, cell size, cell division, cell differentiation, or cell proliferation, and other important developmental processes. The data also indicate that evolutionarily simpler organisms allow for the detection of subtle differences in the conserved IDRs of TFs as well as changes in variable IDRs, which can influence the DNA recognition and multifunctionality of TFs through direct or indirect mechanisms. Although strong correlations cannot be taken as evidence for cause-and-effect relationships, we interpret our data to indicate that increasing TF disorder likely was an important factor contributing to the evolution of organismic complexity and not merely a concurrent unrelated effect of increasing organismic complexity.

    (Emphasis mine)

    Beautiful, isn’t it? 🙂

    I have not time now, but I will come back on that later.

  125. Dionisio, DATCG, Upright Biped:

    This is one of the first important papers about the relationship between TFs and IDR:

    Intrinsic disorder in transcription factors

    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2538555/#FN2

    (Public access)

    Abstract
    Intrinsic disorder (ID) is highly abundant in eukaryotes, which reflect the greater need for disorder-associated signaling and transcriptional regulation in nucleated cells. Although several well-characterized examples of intrinsically disordered proteins in transcriptional regulation have been reported, no systematic analysis has been reported so far. To test for a general prevalence of intrinsic disorder in transcriptional regulation, we used the Predictor Of Natural Disorder Regions (PONDR) to analyze the abundance of intrinsic disorder in three transcription factor datasets and two control sets. This analysis revealed that from 94.13% to 82.63% of transcription factors posses extended regions of intrinsic disorder, relative to 54.51% and 18.64% of the proteins in two control datasets, which indicates the significant prevalence of intrinsic disorder in transcription factors. This propensity of transcription factors for intrinsic disorder was confirmed by cumulative distribution function analysis and charge-hydropathy plots. The amino acid composition analysis showed that all three transcription factor datasets were substantially depleted in order-promoting residues, and significantly enriched in disorder-promoting residues. Our analysis of the distribution of disorder within the transcription factor datasets revealed that: (a) The AT-hooks and basic regions of transcription factor DNA-binding domains are highly disordered; (b) The degree of disorder in transcription factor activation regions is much higher than that in DNA-binding domains; (c) The degree of disorder is significantly higher in eukaryotic transcription factors than in prokaryotic transcription factors; (d) The level of alpha-MoRFs (molecular recognition feature) prediction is much higher in transcription factors. Overall, our data reflected the fact that the eukaryotes with well-developed gene transcription machinery require transcription factor flexibility to be more efficient.

  126. Dionisio, DATCG, Upright Biped:

    Here is the Wikipedia page about IDPs:

    https://en.wikipedia.org/wiki/Intrinsically_disordered_proteins

    A few brief quotes:

    The discovery of IDPs has challenged the traditional protein structure paradigm, that protein function depends on a fixed three-dimensional structure. This dogma has been challenged over the 2000s and 2010s by increasing evidence from various branches of structural biology, suggesting that protein dynamics may be highly relevant for such systems. Despite their lack of stable structure, IDPs are a very large and functionally important class of proteins. In some cases, IDPs can adopt a fixed three-dimensional structure after binding to other macromolecules. Overall, IDPs are different from structured proteins in many ways and tend to have distinct properties in terms of function, structure, sequence, interactions, evolution and regulation

    Intrinsic disorder is particularly enriched in proteins implicated in cell signaling, transcription and chromatin remodeling functions.

    Disordered proteins have a low content of predicted secondary structure

    Many key tumour suppressors have large intrinsically unstructured regions, for example p53 and BRCA1. These regions of the proteins are responsible for mediating many of their interactions.

    Let’s remember that BRCA1 is a RING E3 ligase 1863 AAs long. The RING domain is located at the N terminus, and is abou 50 AAs long. Near the C terminus two BRCT domains, about 70 AAs each, can be recognized, and in the middle BLAST identifies a serine rich BRCT associated domain of about 150 AAs.

    All the rest is “domainless”. We have a shorter N terminus sequence of almost 300 AAs, and a very long C terminus sequence of about 1150 AAs. No recognized domains in those two sequences!

    Now, while a traditional approach to protein function would mainly emphasize the role of the domains, which is certainly very important, the new approach based on recognition of IDRs would probably try to understand the possible relational roles of the two longer sequences.

  127. Here’s a list of papers referenced in this OP:

    Burroughs, A. Maxwell, Lakshminarayan M. Iyer, and L. Aravind. “Structure and Evolution of Ubiquitin and Ubiquitin-Related Domains.” In Ubiquitin Family Modifiers and the Proteasome, edited by R. Jürgen Dohmen and Martin Scheffner, 832:15–63. Totowa, NJ: Humana Press, 2012. https://doi.org/10.1007/978-1-61779-474-2_2.

    Chen, N., S. Balasenthil, J. Reuther, A. Frayna, Y. Wang, D. S. Chandler, L. V. Abruzzo, et al. “DEAR1 Is a Chromosome 1p35 Tumor Suppressor and Master Regulator of TGF- -Driven Epithelial-Mesenchymal Transition.” Cancer Discovery 3, no. 10 (October 1, 2013): 1172–89. https://doi.org/10.1158/2159-8290.CD-12-0499.

    Chen, Zhijian J. “Ubiquitin Signalling in the NF-?B Pathway.” Nature Cell Biology 7, no. 8 (August 2005): 758–65. https://doi.org/10.1038/ncb0805-758.

    Chen, Zhijian J., and Lijun J. Sun. “Nonproteolytic Functions of Ubiquitin in Cell Signaling.” Molecular Cell 33, no. 3 (February 2009): 275–86. https://doi.org/10.1016/j.molcel.2009.01.014.

    Collins, Patricia, Izaskun Mitxitorena, and Ruaidhrí Carmody. “The Ubiquitination of NF-?B Subunits in the Control of Transcription.” Cells 5, no. 4 (May 12, 2016): 23. https://doi.org/10.3390/cells5020023.

    Finley, D., H. D. Ulrich, T. Sommer, and P. Kaiser. “The Ubiquitin-Proteasome System of Saccharomyces Cerevisiae.” Genetics 192, no. 2 (October 1, 2012): 319–60. https://doi.org/10.1534/genetics.112.140467.

    Komander, David, and Michael Rape. “The Ubiquitin Code.” Annual Review of Biochemistry 81, no. 1 (July 7, 2012): 203–29. https://doi.org/10.1146/annurev-biochem-060310-170328.

    Kwon, Yong Tae, and Aaron Ciechanover. “The Ubiquitin Code in the Ubiquitin-Proteasome System and Autophagy.” Trends in Biochemical Sciences 42, no. 11 (November 2017): 873–86. https://doi.org/10.1016/j.tibs.2017.09.002.

    Morreale, Francesca Ester, and Helen Walden. “Types of Ubiquitin Ligases.” Cell 165, no. 1 (March 2016): 248–248.e1. https://doi.org/10.1016/j.cell.2016.03.003.

    Stewart, Mikaela D, Tobias Ritterhoff, Rachel E Klevit, and Peter S Brzovic. “E2 Enzymes: More than Just Middle Men.” Cell Research 26, no. 4 (April 2016): 423–40. https://doi.org/10.1038/cr.2016.35.

    Sun, Shao-Cong. “Non-Canonical NF-?B Signaling Pathway.” Cell Research 21, no. 1 (January 2011): 71–85. https://doi.org/10.1038/cr.2010.177.

    Uchil, P. D., A. Hinz, S. Siegel, A. Coenen-Stass, T. Pertel, J. Luban, and W. Mothes. “TRIM Protein-Mediated Regulation of Inflammatory and Innate Immune Signaling and Its Association with Antiretroviral Activity.” Journal of Virology 87, no. 1 (January 1, 2013): 257–72. https://doi.org/10.1128/JVI.01804-12.

    Veen, Annemarthe G. van der, and Hidde L. Ploegh. “Ubiquitin-Like Proteins.” Annual Review of Biochemistry 81, no. 1 (July 7, 2012): 323–57. https://doi.org/10.1146/annurev-biochem-093010-153308.

    Zhang, Xiaofei, Arne H. Smits, Gabrielle B.A. van Tilburg, Pascal W.T.C. Jansen, Matthew M. Makowski, Huib Ovaa, and Michiel Vermeulen. “An Interaction Landscape of Ubiquitin Signaling.” Molecular Cell 65, no. 5 (March 2017): 941–955.e8. https://doi.org/10.1016/j.molcel.2017.01.004.

    Please, correct any inaccuracies or omissions in the above list. Thanks.

    Please, note that the above list does not include papers referenced in the comments.

  128. Dionisio, DATCG, Upright Biped:

    I would like to spend a few words about a concept which, while implicit in the many things we have said, has not been explicitly touched, neither in the OP nor in the discussion: the irreducible complexity of the Ubiquitin System.

    Now, I think that it is an irreducible complexity of a very particular form.

    Indeed, while we could certainly affirm an irreducible complexity of the whole system, I want to focus here on a very interesting aspect:

    The Ubiquitin System, as we have seen, can be considered as the sum of hundreds, maybe thousands, of specific sub-systems. And each one of them is irreducibly complex.

    I will try to be more clear.

    Each functional sub-system can be considered as formed, at least, by:

    1) Ubiquitin, or some ubiquitin-like protein

    2) An E1 enzyme

    3) An E2 enzyme

    4) An E3 ligase

    5) A target protein (the substrate of ubiquitination)

    6) Usually, one or more deubiquitinating enzyme

    7) Some ubiquitin interactor, recognizing the signal

    8) Some final effector system, or protein, implementing the desired outcome

    These components are more or less always present in each sub-system, even if sometimes some of them can be joined in one structure or protein. For example, in the proteasome the signal recognition and the effector are both part of the same structure, but of different parts of it. And there are rare cases of proteins that are at the same time E2 and E3 enzymes.

    Now, it is rather clear that some parts are shared between some or all the subsystems.

    Ubiquitin, for example, is the same in almost all of them (except when its role is implemented by ubiquitin-like proteins).

    And the E1 enzyme is almost always the same (there are only a couple of forms of it).

    The E2 enzymes are already more varied (about 40 of them), but for the moment we can consider them as mainly shared, for the sake of simplicity.

    So, what is it that defines each individual sub-system?

    The answer is easy enough: it’s the individual substrate, and its finale outcome.

    Because, if we want to really define the function, we have to define it as follows:

    The function of this system is to interact with this specific substrate X and direct it to the specific final outcome Y.

    This function is different for each couple of X and Y.

    But we know that there are thousands of different substrates which are ubiquinated, and we also know that at least some of them can be marked for different outcomes by the ubiquitin system.

    So, it is not an overstatement to say that there are thousands of individual sub-systems in the ubiquitin system, each of them with a specific function, different from the function of all the others, because defined by a different couple of X and Y.

    Of course, much of that diversification (but not all) is implemented by the fundamental role of E3 ligases, of which we have more than 600 different forms.

    So, even if we reason only in terms of the characterizing E3 ligase (which is certainly reductive, but at least is simple), we have hundreds of different functional sub-systems, each of them defined by:

    a) A specific substrate (which indeed is the object on which the system works, rather than a part of it)

    b) A specific E3 ligase

    c) A specific symbolic signal (generated by the interaction of a and b)

    d) A specific outcome

    OK? Now comes the interesting part:

    Each of those sub-systems, even if it shares parts with the other sub-systems (for example the E1 enzyme or the proteasome), is in itself irreducibly complex.

    Why?

    Because it would have no function at all if all the parts we have listed were not present, and it would have no function at all if even one of the specific parts, in particular the specific E3 ligase, were not present.

    IOWs, the system is irreducibly complex because no single part of it can be discarded, in the core form we have described. And that irreducible complexity is different for each subsystem, because at least one part (the E3 ligase) is absolutely specific, and cannot be discarded at all. Because it’s the E3 ligase that recognizes the specific Y, and knows what signal is appropriate for it.

    Of course, many other factors can contribute to the unicity of each sub-system. For example, even if one E3 ligase can deal with many different substrates, it is perfectly reasonable that a specific combination of E1, E2, E3, signal, deubiquinating enzyme and outcome is usually unique for most substrates. That would make thousands of unique irreducibly complex sub-systems, rather than “only” hundreds.

    So, while the Ubiquitin System is, as stated in the OP and analyzed in detail in the discussion, a wonderful example where functional complexity and semiosis are joined, it is equally true that we can extend that statement to the third outstanding feature of complex designed systems, and say that:

    The Ubiquitin System is a wonderful example of a system where functional complexity, semiosis and irreducible complexity are joined together.

  129. Dionisio at #121 and 122:

    This is another completely new aspect, and discovered very recently, it seems. Thank you for finding that! You are, as usual, a very good digger. 🙂

    Of course, RNA could not be missing form the many-faceted world of ubiquitin regulation. In the end, RNA is probably the key crossroad where all regulatory networks do join. I suppose that Arthur Hunt would agree on that! 🙂

    TRIM25 seems to be a very busy actor too. From Uniprot:

    Functions as a ubiquitin E3 ligase and as an ISG15 E3 ligase. Involved in innate immune defense against viruses by mediating ubiquitination of DDX58. Mediates ‘Lys-63’-linked polyubiquitination of the DDX58 N-terminal CARD-like region which is crucial for triggering the cytosolic signal transduction that leads to the production of interferons in response to viral infection. Promotes ISGylation of 14-3-3 sigma (SFN), an adapter protein implicated in the regulation of a large spectrum signaling pathway. Mediates estrogen action in various target organs. Mediates the ubiquitination and subsequent proteasomal degradation of ZFHX3

  130. gpuccio @129:

    TRIM25 seems to be a very busy actor too.

    Actors in Hollywood would dream of having at least a fraction of the roles TRIM25 seems to perform according to the information you quoted. 🙂

    Ubiquitin could collect Oscars in a warehouse. 🙂

    However, ironically ubiquitin is not a celebrity according to the mainstream media. Not yet.

    BTW, you mentioned professor Arthur Hunt. Any news from him? Did he ever come back to continue the discussion on the spliceosome?

  131. gpuccio,

    FYI – The list @127 was produced using Zotero.

    However, if I were to stick to the neo-Darwinian style, I would have to say that it was produced by Zotero, which implies that the software decided how, where and when to produce that list.

    🙂

  132. Dionisio:

    No, no news from Arthur Hunt.

    However, I would say that his “presence” seems to linger among us! 🙂

    It seems to be a good strategy: gives just a little of yourself, and you will be sorely missed. 🙂

  133. Dionisio:

    Zotero seems an useful tool. I will try it.

    Do you think it is a designed object? 🙂

  134. Dionisio, DATCG, Upright Biped:

    Hey, we have almost forgotten apoptosis! 🙂

    Fortunately, here is a brand new paper on that subject:

    Delineating Crosstalk Mechanisms of the Ubiquitin Proteasome System That Regulate Apoptosis

    https://www.frontiersin.org/articles/10.3389/fcell.2018.00011/full

    (Public access)

    Abstract:

    Regulatory functions of the ubiquitin-proteasome system (UPS) are exercised mainly by the ubiquitin ligases and deubiquitinating enzymes. Degradation of apoptotic proteins by UPS is central to the maintenance of cell health, and deregulation of this process is associated with several diseases including tumors, neurodegenerative disorders, diabetes, and inflammation. Therefore, it is the view that interrogating protein turnover in cells can offer a strategy for delineating disease-causing mechanistic perturbations and facilitate identification of drug targets. In this review, we are summarizing an overview to elucidate the updated knowledge on the molecular interplay between the apoptosis and UPS pathways. We have condensed around 100 enzymes of UPS machinery from the literature that ubiquitinates or deubiquitinates the apoptotic proteins and regulates the cell fate. We have also provided a detailed insight into how the UPS proteins are able to fine-tune the intrinsic, extrinsic, and p53-mediated apoptotic pathways to regulate cell survival or cell death. This review provides a comprehensive overview of the potential of UPS players as a drug target for cancer and other human disorders.

    Emphasis mine.

    “Around 100 enzymes” from the Ubiquitn System, just to regulate Apoptosis? Not bad, I would say.

    “Fine tune”? You bet!

    By the way, Figures 3,4, 5 and 6 (and their respective legends) are some more fun for the lovers of simplicity! 🙂

    And Tables 1, 2, 3, 4, 5, 6, 7 and 8 are a very good list of the ubiquinitating and deubiquinating enzymes involved in this “simple” regulation network.

    But, of course, RV and NS can easily explain all that. No need to take part in such a trivial discussion…

  135. Dionisio, DATCG, Upright Biped:

    Just a quick update of the list at #90:

    A brief list of the cellular processes we have touched in the OP and discussion, all of them deeply connected to the Ubiquitin System network:

    a) Proteasome degradation

    b) Autophagic degradation

    c) Mitophagy

    d) Cell signaling and transmission pathways

    e) Double-Strand Breaks DNA repair

    f) Neuronal regulation

    g) Regulation of innate immunity

    h) Regulation of adaptive immunity

    i) Regulation of T cell and B cell differentiation

    j) RNA interactions

    k) Apoptosis

    OK, I am sure we are going to add new ones, if we keep digging. 🙂

  136. Dionisio, DATCG, Upright Biped:

    We have seen the role of ubiquitin system in histone ubiquitination, especially in DNA repair.

    But what about DNA methylation, probably the first and foremost epigenetic mechanism?

    Another brand new paper:

    Structural and mechanistic insights into UHRF1-mediated DNMT1 activation in the maintenance DNA

    https://academic.oup.com/nar/advance-article/doi/10.1093/nar/gky104/4870013

    Abstract:

    UHRF1 plays multiple roles in regulating DNMT1-mediated DNA methylation maintenance during DNA replication. The UHRF1 C-terminal RING finger functions as an ubiquitin E3 ligase to establish histone H3 ubiquitination at Lys18 and/or Lys23, which is subsequently recognized by DNMT1 to promote its localization onto replication foci. Here, we present the crystal structure of DNMT1 RFTS domain in complex with ubiquitin and highlight a unique ubiquitin binding mode for the RFTS domain. We provide evidence that UHRF1 N-terminal ubiquitin-like domain (UBL) also binds directly to DNMT1. Despite sharing a high degree of structural similarity, UHRF1 UBL and ubiquitin bind to DNMT1 in a very distinct fashion and exert different impacts on DNMT1 enzymatic activity. We further show that the UHRF1 UBL-mediated interaction between UHRF1 and DNMT1, and the binding of DNMT1 to ubiquitinated histone H3 that is catalyzed by UHRF1 RING domain are critical for the proper subnuclear localization of DNMT1 and maintenance of DNA methylation. Collectively, our study adds another layer of complexity to the regulatory mechanism of DNMT1 activation by UHRF1 and supports that individual domains of UHRF1 participate and act in concert to maintain DNA methylation patterns.

    DNMT1, DNA (cytosine-5)-methyltransferase 1, is a key enzyme in DNA methylation, and many other things. From Uniprot:

    Methylates CpG residues. Preferentially methylates hemimethylated DNA. Associates with DNA replication sites in S phase maintaining the methylation pattern in the newly synthesized strand, that is essential for epigenetic inheritance. Associates with chromatin during G2 and M phases to maintain DNA methylation independently of replication. It is responsible for maintaining methylation patterns established in development. DNA methylation is coordinated with methylation of histones. Mediates transcriptional repression by direct binding to HDAC2. In association with DNMT3B and via the recruitment of CTCFL/BORIS, involved in activation of BAG1 gene expression by modulating dimethylation of promoter histone H3 at H3K4 and H3K9. Probably forms a corepressor complex required for activated KRAS-mediated promoter hypermethylation and transcriptional silencing of tumor suppressor genes (TSGs) or other tumor-related genes in colorectal cancer (CRC) cells (PubMed:24623306). Also required to maintain a transcriptionally repressive state of genes in undifferentiated embryonic stem cells (ESCs) (PubMed:24623306). Associates at promoter regions of tumor suppressor genes (TSGs) leading to their gene silencing (PubMed:24623306). Promotes tumor growth.

  137. Dionisio, DATCG, Upright Biped:

    What about early embryo development?

    Embryonic lethality in mice lacking Trim59 due to impaired gastrulation development.

    https://www.nature.com/articles/s41419-018-0370-y

    (Public access)

    Abstract:

    TRIM family members have been implicated in a variety of biological processes such as differentiation and development. We here found that Trim59 plays a critical role in early embryo development from blastocyst stage to gastrula. There existed delayed development and empty yolk sacs from embryonic day (E) 8.5 in Trim59-/- embryos. No viable Trim59-/- embryos were observed beyond E9.5. Trim59 deficiency affected primary germ layer formation at the beginning of gastrulation. At E6.5 and E7.5, the expression of primary germ layer formation-associated genes including Brachyury, lefty2, Cer1, Otx2, Wnt3, and BMP4 was reduced in Trim59-/- embryos. Homozygous mutant embryonic epiblasts were contracted and the mesoderm was absent. Trim59 could interact with actin- and myosin-associated proteins. Its deficiency disturbed F-actin polymerization during inner cell mass differentiation. Trim59-mediated polymerization of F-actin was via WASH K63-linked ubiquitination. Thus, Trim59 may be a critical regulator for early embryo development from blastocyst stage to gastrula through modulating F-actin assembly.

    Emphasis mine.

    TRIM59 is a RING E3 ligase. This is apparently a new functional specificity, because the Uniprot page function section states simply:

    “May serve as a multifunctional regulator for innate immune signaling pathways.”

  138. Dionisio, DATCG, Upright Biped:

    Embryonic stem cells are certainly an extremely hot subject.

    This too is brand new:

    Cellular functions of stem cell factors mediated by the ubiquitin–proteasome system

    https://link.springer.com/article/10.1007%2Fs00018-018-2770-7

    (Paywall)

    Abstract:

    Stem cells undergo partitioning through mitosis and separate into specific cells of each of the three embryonic germ layers: endoderm, mesoderm, and ectoderm. Pluripotency, reprogramming, and self-renewal are essential elements of embryonic stem cells (ESCs), and it is becoming evident that regulation of protein degradation mediated by the ubiquitin-proteasome system (UPS) is one of the key cellular mechanisms in ESCs. Although the framework of that mechanism may seem simple, it involves complicated proteolytic machinery. The UPS controls cell development, survival, differentiation, lineage commitment, migration, and homing processes. This review is centered on the connection between stem cell factors NANOG, OCT-3/4, SOX2, KLF4, C-MYC, LIN28, FAK, and telomerase and the UPS. Herein, we summarize recent findings and discuss potential UPS mechanisms involved in pluripotency, reprogramming, differentiation, and self-renewal. Interactions between the UPS and stem cell transcription factors can apply to various human diseases which can be treated by generating more efficient iPSCs. Such complexes may permit the design of novel therapeutics and the establishment of biomarkers that may be used in diagnosis and prognosis development. Therefore, the UPS is an important target for stem cell therapeutic product research.

  139. #123 Gpuccio,

    Oh joy 🙂 haha, more reading! I briefly looked at your first link OP on Information Jump that started all of this. Briefly looked through it and read the epic take down in your comment at #138 of Alicia’s selective blast. Ha!

    OK, looks like more OPs to read. Thanks for sharing those links. I’ll catch up over time to gain more perspective.

  140. #112 Dionisio

    Agree, and a belief in blind, unguided orthodoxy.

    It may be why Systems Biologist and Systems based approaches in molecular sciences have recognized the lack of answers and are more willing to open up discussions on failure of neo-Darwinism to address known problems.

    Eventually science must move ahead, be it Darwin dead or neo-Darwinism being pushed aside too as a weak explanation.

  141. GP,

    10 days and 140 comments, not a critic in sight. Congratulations.

    You should be feeling good about that.

    🙂

  142. #123 Gpuccio,

    1st, thanks for playing along with my nomenclature mini-rant
    😉 Alas, no renaming of IDP, IDR.

    I guess I see IDPs as flexible-ordered 3D states. A bit like a rubic cube but in a combinatorial process that recognizes a match or is stimulated and folds upon binding sites.

    Or another analogy, a universal socket or universal wrench that adjust based upon conditional sizes of bolts.

    In case of IDPs, their flexibly adjustable so-to-speak to a binding site of multiple substrates

    Disorder represents a randomness to it, even if it’s used in contrast to normal folding or “normal” regions. Whatever is normal per say in a random world of natural selection?

    If by Design, then there’s a reason for this open pattern, 3D free structure and so-called disordered regions.

    Came across another paper re: IDPs, IDRs and folding, partial-fold, conditional-folds, etc.

    Mosaic nature of the protein structure–function space: functional foldons, semi-foldons, inducible foldons, nonfoldons and unfoldons

    On the other hand, studies of the past decade and a half have clearly showed that these models provide a suitable functional description of only a part of a protein universe because not all proteins are structured throughout their entire lengths, with many biologically active proteins instead being highly flexible or intrinsically disordered [5-15]. Indeed, bioinformatics studies indicate that intrinsically disordered proteins (IDPs) and hybrid proteins with ordered domains and intrinsically disordered protein regions (IDPRs) are highly abundant in nature [16-20], with ~ 25–30% of eukaryotic proteins being mostly disordered [18], more than half of eukaryotic proteins having long regions of disorder [16-18] and > 70% of signaling proteins possessing long disordered regions [21]. Functions of IDPs and IDPRs are typically complementary to functions of ordered proteins and domains [5, 6, 8, 9, 15].

    and more…


    Some functions of IDPRs: functionality of inducible foldons, semi-foldons and nonfoldons

    A recent review [32] emphasized that the distribution of intrinsic disorder within the protein sequence is nonhomogeneous, and protein tails are typically more disordered than residues in the middle of the protein chain [33]. Furthermore, although tails can be engaged in the majority of functions ascribed to IDPRs in general, there are some disorder-based functions that are tail-specific [32]. Accordingly, the functions of the intrinsically disordered protein tails (IDPTs) were systemized according to the basic molecular mechanism of their action (e.g. interaction or binding, entropic activities, chaperone functions) and based on the conditions resulting from their action (e.g. inhibition, activation, protection, stabilization, etc.) [32]. Because such activities of protein tails are often associated with at least the partial folding of at least some parts of these IDPTs, the aforementioned functions are attributed to semi-foldons and inducible foldons. Among the interaction-based functions of IDPTs are specific activities, where disordered regions act as inhibitors (including autoinhibitors and external inhibitors), competitors, activators, affinity tuners, signal carriers, DNA sliders and brachiation domains, DNA/RNA benders and twisters, engagers, protectors, stabilizers, assemblers, modification displays, degrons, intertwinders, penetrators, domain swappers, chameleons, hijackers, anchors, switches, recyclers, grabbers, multirods, recruiters, and capasitators [32]. Among the entropic chain functions attributed to the disordered tails are entropic bristles, solubilizers and entropic clocks. Also, chaperone-related functions are assigned to the IDPTs of some protein and RNA chaperones, as well as (for some proteins) to the disordered tails acting as intramolecular chaperones [32]. Obviously, these entropic chain activities are functions of nonfoldons (i.e. protein parts that remain mostly unfolded in their functional states).

    There’s a reason for nonhomogeneous distinction and long length of disordered regions. And tails being more disordered as well. What yet I do not know, but if by design there is.

    This paper notes that IDPs IDPTs IDRs are complimentary to “ordered” proteins.

    and more… environmental factors 🙂


    Some functions of the transiently disordered regions: unfoldons in action

    The basic difference in the functionality of the transiently disordered regions (unfoldons) with respect to the functionality of IDPRs is the crucial dependence of the conditionally disordered proteins/regions on their unfolding. Therefore, functions of unfoldons can be classified based on the molecular mechanisms responsible for the induction of such functional unfolding [31]. Figure 2 shows that cryptic disorder can be awoken by a wide spectrum of factors, which were grouped into two major classes: passive and active [31]. Passive factors were defined as factors originated from some global changes in protein environment, such as changes in temperature, pH, mechanical force, the redox potential or light exposure. These passive environmental factors were independent of the specific interaction between the protein and its partners [31]. Active factors were defined as those related to some specific interactions of a protein with its environment. Among such active factors were interactions with membranes, ligands, other proteins and nucleic acids, as well as the different means responsible for the release of autoinhibition and various PTMs causing local protein unfolding [31].

    Who knew disorder could be functional? 😉

  143. oops, wrong link in #142, Should be…


    Mosaic nature of the protein structure–function space: functional foldons, semi-foldons, inducible foldons, nonfoldons and unfoldons…

    http://www.pnas.org/content/111/43/15420.full

    .

  144. I wonder if his graphic is entirely accurate though…

    Obviously, all these pieces of the protein structural mosaic might have well-defined and specific functions (Fig. 1). Below, some of the characteristic disorder-based functions of semi-foldons, inducible foldons, nonfoldons and unfoldons are considered.

    Fig 1 Functional roles of transiently and intrinsically disordered regions within proteins Circle of

    Figure 1. Schematic representation of the mosaic nature of the protein structure–function space. It should be noted that ‘Dormant disorder’ is different from the other ‘outer-ring’ functional grouping because the corresponding segment does not describe a particular functional group but rather represents the means by which the functionality is achieved.

    .

  145. Try this one last time, for link at #142…

    Yay…

    Functional roles of transiently and intrinsically disordered regions within proteins

    http://onlinelibrary.wiley.com.....13202/full

  146. Gpuccio @126
    re: BRCA1

    Enjoying reading FEBS Journal by FEBSPress on the subject
    you expanded upon. Don’t remember coming across this journal in the past.

    re: your comments…

    All the rest is “domainless”. We have a shorter N terminus sequence of almost 300 AAs, and a very long C terminus sequence of about 1150 AAs. No recognized domains in those two sequences!

    Yep, long sequences.

    Now, while a traditional approach to protein function would mainly emphasize the role of the domains, which is certainly very important, the new approach based on recognition of IDRs would probably try to understand the possible relational roles of the two longer sequences.

    That’s interesting, 1481AAs from paper below in 2014
    on BRCA1 …

    BRCA1, A ‘Complex’ Protein Involved in the Maintenance of Genomic Stability – 2014

    ~1481AAs = “…no Domain structure and is predicted to be intrinsically disordered…”

    The central region of the BRCA1 protein is largely encoded by a single exon (exon 11) that is present in vertebrates but not in lower eukaryotes. This region, covering amino acids 170–1649 contains no known domain structures and is predicted to be intrinsically disordered by both in silico and experimental analysis [19, 20]. This type of protein structure (i.e. a disordered region flanked by structural domains) is typical of scaffold or ‘hub’ proteins.

    So it is a “type of protein structure” Just not rigid folding angles of “noraml” proteins so they named it disordered, instead of the Flexible aspect, or Conditional activity of folding. Or Induced, etc. Though I agree Gpuccio, Flexible includes all folds and activity.

    But can they make up their minds? “Type of Structure,” no
    structure? Unstructural? 😉

    I digress, onward…

    Disordered regions allow binding promiscuity as well as *structural flexibility* between ordered domains, thereby facilitating binding diversity and allowing the conformational flexibility to facilitate the formation of multiple different macromolecular complexes [20].

    Indeed, this region(disordered) of BRCA1 has been shown to interact with numerous different proteins and contains a number of DNA damage-induced phosphorylation sites that are considered to mediate the formation of various BRCA1 protein complexes [7].

    * , () emphasis mine

    There’s some info on N-Terminus…

    At its N-terminus, BRCA1 contains a series of eight conserved Cys3-His-Cys4 motif repeats known as the RING domain, a catalytic domain involved in protein ubiquitination and protein–protein interactions [7]. The most well characterized RING domain interaction occurs with the BRCA1 associated RING domain protein 1 (BARD1), forming the BRCA1/BARD1 heterodimer, a known E3 ubiquitin ligase complex [8]. It is now well accepted that BRCA1 and BARD1 are obligate heterodimers, with each protein required for the stability of the other [9].

    .

  147. And kind of funny, came across Douglas Axe posting on Conditional Folding! 🙂

    I’m not the only one stuck on meaningful nomanclature. Plus great stuff on IDPs! And good response to Venema.

    Conditional Folding Is Still Folding
    On the first point, Venema surely knows he’s misleading his readers. With respect to proteins, folding refers to the process by which initially floppy protein chains lock into well-defined three-dimensional structures that perform specific functions within cells. Venema cites a good review paper on so-called “intrinsically disordered proteins,” claiming the existence of this class of proteins shows that protein function doesn’t actually require folding. However, if Venema read the paper, he knows it has a section titled “Coupled folding and binding,” referring to the “mechanism by which disordered interaction motifs associate with and fold upon binding to their targets” (emphasis added). In other words, the term “intrinsically disordered proteins” is a misnomer (whoever coined the term evidently didn’t know what the word intrinsic means).

    “A better term would be conditionally folded proteins.”

    Moreover, anyone who reads this review paper with open eyes will see that conditional folding is in fact a remarkable design feature. As the authors say, “An exciting recent finding is that many proteins containing low-complexity or prion-like sequences can promote phase separation to form membrane-less organelles within the cytoplasm or nucleoplasm, thus contributing to their compartmentalization in a regulated manner.” Speaking of conditionally folded proteins in general, the authors note that the levels of these proteins within cells are “tightly regulated to ensure precise signaling in time and space, and mutations in [them] or changes in their cellular abundance are associated with disease.”

    So, if Venema pictures these conditional folders as being easy evolutionary onramps for mutation and selection to make unconditionally folded proteins, he’s badly mistaken. Both kinds of proteins are at work in cells in a highly orchestrated way, both requiring just the right amino-acid sequences to perform their component functions, each of which serves the high-level function of the whole organism

    That’s great stuff by Axe! And a good take down of Venema’s lazy reading of a research paper.

    Axe’s response to an upcoming critique and response to his Book …
    Losing the Forest by Fixating on the Trees — A Response to Venema’s Critique of Undeniable

    .

  148. Upright Biped:

    “10 days and 140 comments, not a critic in sight. Congratulations.

    You should be feeling good about that.”

    Maybe I should. But I miss the fight! 🙂

    DATCG just mentioned Alicia Cartelli, and, believe it or not, I felt a pang of nostalgia! 🙂

  149. Understood.

    …but still.

    🙂 🙂 🙂

  150. DATCG:

    “Who knew disorder could be functional?”

    Thank you for the link.

    “Functional roles of transiently and intrinsically disordered regions within proteins”

    Very interesting indeed.

    So, folding and unfolding can be considered as a continuum, a space of functional potentialities which is differently structured in different proteins and protein regions. Fascinating.

    I thin the concept here is not so much of “disorder”, but rather of “absence of a rigid, or prevailing, modality of folding”.

    This is a very interesting point. Indeed, functional specification has little to do with order. Functional sequences are often more pseudo-random than pseudo-ordered. That’s one important reason why functional complexity can never arise from necessity laws, which by definition are defined by regularities.

    Secondary structure is probably more tractable and more similar to some form of “order”, but tertiary structure can scarcely be predicted with our resources.

    A functional sequence which eludes both secondary structures and rigid tertiary structures is even more interesting, because it seems to be pure function with really negligible “order”. In that sense, it is an even more amazing object of design.

    A scenario of incredible functional flexibility and complexity is daily emerging from the new biology. It’s not surprising that out kind interlocutors don’t seem too keen to discuss it! 🙂

  151. DATCG at #147:

    Axe is one of the best! 🙂

    By the way, I am reading his “Undeniable”. I will probably write something about it when I am finished.

    He says:

    “Moreover, anyone who reads this review paper with open eyes will see that conditional folding is in fact a remarkable design feature.”

    Of course, he is absolutely right! The function of IDPs and IDRs (however we call them) screams design even more than the traditional function of well understood domain proteins!

  152. Gpuccio #150

    “So, folding and unfolding can be considered as a continuum, a space of functional potentialities which is differently structured in different proteins and protein regions. Fascinating.”

    Yes! Thanks for summing it up nicely. 🙂

    Functional Potential, conditional, induced or not. The “Flexibility” of the structure allows it.

    There’s a place for both Rigid enforcement of angles and flexible IDPs based on Systems
    Requirement, Surroundings or function specificity – Purpose.

    This is a very interesting point. Indeed, functional specification has little to do with order. Functional sequences are often more pseudo-random than pseudo-ordered. That’s one important reason why functional complexity can never arise from necessity laws, which by definition are defined by regularities.

    Yes 🙂 Again, thanks for simplifying with a good summary! It’s common sense.

    But it’s hard to use common sense from a Darwin perspective. They must use convoluted reasoning, avoidance of certain terms to get around, jump-over and slide-under Design in Life to embrace unguided blind random events.

    What’s funny is even if they carefully self-govern or by incompetence use language that keeps out Design principles, it inevitably creeps back in because even the most ardent Darwinist have a slip of the tongue. A rotor’s a rotor and not by accident.

    We know there’s differences between Random, Ordered and Organized Function(see Abel, Trevors;
    Three Subsets of Sequence Complexity: Random, Ordered and Functional… Theorectical Biology)

    Where: Functional = Organized Code

    Which brings us back to your point on Semiosis and UB’s contributions on the subject.
    What is information? How is it represented and communicated? Different Types and Category?

    This is where Darwinist usually dismiss these areas based on definitions and inability to measure precisely Organized, Sybmolic, Informational Content, etc. There’s been many good post on this here at UD.

    But they continue to miss the entire point. It’s Code! 🙂 And Code is well known, common sense sign of Design. Not of random process, certainly not ordered patterns.

    Secondary structure is probably more tractable and more similar to some form of “order”, but tertiary structure can scarcely be predicted with our resources.

    Hopefully new technology will allow scientist to look deeper and through time as flexible folds are captured during binding or unfolding and release. I thought the NMR
    video captured the difficulty you mention quite well.

    A functional sequence which eludes both secondary structures and rigid tertiary structures is even more interesting, because it seems to be pure function with really negligible “order”. In that sense, it is an even more amazing object of design.

    Organized Functionality 🙂 Sequences and structure may appear “disordered” but they’re highly organized, not random. With intent of flexible functionality. Screams Design.

    And as the paper by Abel, Trevors points out, “Functional Sequence Complexity is invariably associated with all forms of complex biofunction…,”

    Whereas “No empirical evidence exist of either Random Sequences or Ordered Sequences having produced a single instance of sophisticated biological organization.”

    “Organization… manifest Functional Sequence Complexity, rather than Random Events(RSC) or low-informational self-ordering phenomena(OSC).”

    Random does not, Ordered does not. Symbolic Organization does via Design.

    A scenario of incredible functional flexibility and complexity is daily emerging from the new biology. It’s not surprising that out kind interlocutors don’t seem too keen to discuss it!

    right! 🙂 You’ve put together another well thought out post. Thanks for your patience as well in these discussions.

    Really have enjoyed these post.

    .

  153. #152 Gpuccio

    re: Axe

    As Axe stated, most people get Design intuitively. That was a
    great read, his article that is 🙂 He mops up the floor with Venema.

    I’ve not purchased Undeniable yet, but will. Have read Meyers
    Signals in the Cell which I thoroughly enjoyed. Still need to get through Darwin’s Doubt. Look forward to Axe’s book.

    How do you like it so far? Not to give away your Book Review, , but what is a highlight of the book to you?

    .

  154. Just to mention once again Gpuccio.

    So, I’d kinda went on a mini-rant about terms. But had not read Axe. Then, when I came across Axe’s comments in that article. it made me realize I was not alone in my frustration and if an expert scientist like him sees this, it made me feel like I was on the right track.

    Terms matter and Darwinist are always choosing which are standardized into our vocabulary of genetics, molecular biology, etc. Then children are indoctrinated to it. If I could, I’d rewrite so much of it, lol.

    Like an entire Architectural Design Initiative to free minds of the blind, and open their eyes to Design.

    It would be a massive undertaking of course. But what if, instead of Latin, instead of indoctrinated language of blind, unguided process, we could rewrite Life’s processes by Design Code Logic and nomemclature?

    Just a wild thought. In the approach of an Open Code of Life, so to speak. There’s closed platforms like Apple and Microsoft, then there is Linux and solutions like WordPress, etc, many open architectures where developers around the world contribute.

    Kids and young people pick up on it fast, very fast.

    I think sometimes, the past holds us back. By restricting thought to ancient languages and antiquated ideas(Darwinism), the specialist keep a closed platform. Many of these specialist are great documenters, collectors and categorizers, but many are not innovators or creatives. Not to say we do not need such specializations, but often times I think the barriers are artificially high due to past historical norms.

    We need to open it up. Open up the Codes of Life to innovators and creative people.

  155. For readers, Video and explanations of Protein Structures…

    Primary, secondary and tertiary…
    https://www.khanacademy.org/science/biology/macromolecules/proteins-and-amino-acids/v/tertiary-structure-of-proteins

  156. FYI, has anyone seen Abel’s paper from 2015?

    off-topic: PDF Document…

    Functional Sequence Complexity (FSC) Measured in Fits(Functional bits)

    No empirical evidence exists of either RSC of OSC ever having produced a single instance of sophisticated function or true organization. Algorithmic optimization requires purposeful choices to pursue eventual ideal function. Prescriptive Information (PI), circuit integration, and organization all invariably manifest FSC. Any attempt to deny the need and reality of purposeful choices precludes the production of any sophisticated function. Naturalistic philosophic presuppositions militate against acknowledging the obvious facts of reality. “Chance and Necessity” is a false dichotomy. Reality actually consists of three fundamental categories, not two: Chance, Necessity and Choice. By Choice, we do not mean mere Selection FROM AMONG [evolution]. The kind of Choice clearly observed everyday by everyone as a major component of reality includes Selection FOR (IN PURSUIT OF) not yet existent function.8-11 Inanimate nature cannot exercise or generate such choice with intent. Only agency does. Mere mass/energy interactions have never been shown to produce the slightest hint of agency.

    Functional Bits (Fits) The evolution of amino acid sequence, and its effect on biofunction, can now be quantified in “fits” (functional bits).4 To understand how Functional Sequence Complexity can be measured, we must first understand “Functional Uncertainty (Hf):”

    “Shannon’s original formulation, when applied to biological sequences, does not express variations related to biological functionality such as metabolic utility. Shannon uncertainty, however, can be extended to measure the joint variable (X, F), where X represents the variability of data, and F functionality. This explicitly incorporates empirical knowledge of metabolic function into the measure that is usually important for evaluating sequence complexity. This measure of both the observed data and a conceptual variable of function jointly can be called Functional Uncertainty (Hf),74 and is defined by the equation:

    H(Xf(t)) = – ? P(Xf(t)) logP(Xf(t)) (1) where t = a certain time and Xf denotes the conditional variable of the given sequence data (X) on the described biological function f which is an outcome of the variable (F).”74

    7 In this approach, f might represent the known 3-D structure of a protein family. The entire set of aligned sequences that satisfies that protein’s function, therefore, would constitute the outcomes of Xf. The advantage of using H(Xf(t)) is that evolutionary changes through time in the functionality of sequences can be measured.

    .

  157. Back on topic, Durston, Wong, Chiu, Li paper(2012)…
    Statistical Discovery of Site inter-dependencies in Sub-molecular Hierarchical Protein Structuring

    Partial Background

    To reveal the patterns of associations among individual amino acids or sub-domain components within the structure, we apply ak-modes attribute (aligned site) clustering algorithm to the ubiquitin and transthyretin families in order to discover associations among groups of sites within the multiple sequence alignment. We then observe what these associations imply within the 3D structure of these two protein families.

    Results

    The k-modes site clustering algorithm we developed maximizes the intra-group interdependencies basedon a normalized mutual information measure. The clusters formed correspond to sub-structural components or binding and interface locations. Applying this data-directed method to the
    ubiquitin and transthyretin protein family multiple sequence alignments as a test bed, we located numerous interesting associations of interdependent sites. These clusters were then arranged into cluster tree diagrams which revealed four structural sub-domains within the single domain structure of ubiquitin and a single large sub-domain within transthyretin associated with the interface among transthyretin monomers. In addition, several clusters of mutually interdependent sites were discovered for each protein family, each of which appear to play an important role in the molecular structure and/or function.

    Conclusions:

    Our results demonstrate that the method we present here using a k-modes site clustering algorithm based on interdependency evaluation among sites obtained from a sequence alignment of homologous proteinscan provide significant insights into the complex, hierarchical inter-residue structural relationships within the 3D structure of a protein family

    Section on Classifying the Structures

    To evaluate the validity of this modular view of ubiquitin, it has already been observed that at pH values of 6.8 to 7.5, the chemical shift perturbations are modular between the single ubiquitin molecule and the Lys-48linked dimer form of ubiquitin (Ub2), with four areas of perturbations [41]. The same four areas of perturbationshave also been observed for Lys-63 linked Ub2[40].

    The remarkable correspondence among the four modules obtained by our method and the four areas of chemical shift perturbations is summarized in Table 4, providing validation that these four modules actually represent structural units within ubiquitin.

  158. DATCG at #153:

    I am still at the beginning of Undeniable, too early to say something, except that it seems well written and interesting.

  159. DATCG at #152:

    Thank you for quoting Abel and Trevors, and their fundamental paper.

    I think that Abel and his co-workers have given us some very clear and basic intuitions, that still remains as foundations of ID. I am paticularly in debt to Abel for many important concepts, like the three types if sequence complexity, the difference between descriptive information and prescriptive information, the idea of “configurable switches” and so on.

    Durston is another important contributor to ID thought. He has been the first to propose a real method to measure functional information in proteins:

    Measuring the functional sequence complexity of proteins

    https://tbiomed.biomedcentral.com/articles/10.1186/1742-4682-4-47

    While my approach, based on sequence conservation, is slightly different, the basic idea is the same.

    I did not know Durtson’s paper on ubiquitin, that you quote at #157. I will read it with great interest! 🙂

    The same is true for Abel’s paper from 2015 that you reference at #156. I hope to comment on that too, as soon as possible. 🙂

    Thank you for your precious contributions!

  160. Exosome relation with ubiquitin regulation of platelets.

    Exosome poly-ubiquitin inhibits platelet activation, downregulates CD36 and inhibits pro-atherothombotic cellular functions

    Authors: S. Srikanthan, W. Li, R. L. Silverstein, T. M. McIntyre

    First published: 13 October 2014

    Exosomes of nucleated cells are enriched with ubiquitin [32] and ubiquitinated proteins [23, 32], and are nature’s most conserved carriers of extracellular ubiquitin [1, 33].

    The immunosuppressive and anti-inflammatory properties of exosomes may relate to this ubiquitin [22-24], and free ubiquitin inhibits select immune events [34].

    Platelets also modify their proteins with ubiquitin, and exosomes shed from stimulated platelets were extensively adducted by ubiquitin.

    Platelets contain a functional ubiquitin/proteasome system [35, 36] that controls their high avidity responses to thrombin [37] and, we now find, rapidly modifies and degrades CD36. Ubiquitin availability in platelets may be limiting because exosome accumulation by platelets (Fig. S1) increased ubiquitination of platelet proteins (Fig.S2). MG-132 enhanced this ubiquitination (Fig. S3), indicating exosomal ubiquitin contributes to the ubiquitin cycle. Extracellular ubiquitin can participate in platelet ubiquitin metabolism becauserecombinant poly-ubiquitin also reduced platelet reactivity. Notably, this poly-ubiquitin was unanchored, that is, not conjugated to protein. Protein-free poly-ubiquitin does exist and may have a role in innate immunity [38] as well as kinase activation [39].

  161. Gpuccio #159

    The day I first came across Abel and Trevors paper; Three Subsets…, was eye-opening in understanding the deception of Darwinism not recognizing Organization as Design principle.

    I read through it again and again. And probably need to refresh after so long.

    I agree, they’re contributions are foundational and clear away old Darwinian concepts or deceptions to muddy the water between Order and Organization.

    And yes, Durston 🙂 So I’m curious what you think of Abel’s FITS paper. Look forward to it!

    The Null Hypothesis they set forth I don’t think has been challenged? But it’s been some time.

    We repeat that a single incident of nontrivial algorithmic programming success achieved without selection for fitness at the decision-node programming level would falsify any of these null hypotheses. This renders each of these hypotheses scientifically testable. We offer the prediction that none of these four hypotheses will be falsified.

    I wonder if any of day to day commenters her at UD are willing to give it a go in attempt to falsify Abel and Trevors’ challenge. That would make for an interesting post?

    Have a good day/evening.

  162. DATCG at #160:

    Amazing. So, we can add cell to cell communication to the processes where ubiquitin is directly involved!

  163. Gpuccio #162
    thought you might like that 🙂

  164. DATCG:

    I have read the Durston paper linked at #157, Very interesting.

    Durston is certainly going on finding beautiful ways to analyze protein sequences for specific patterns.

    His guiding principle is, as always, functional information. And therefore he founds his measures upon sequence conservation. And that is great!

    In this paper, he is focusing not on traditional functional information, but on functional information related to aminoacid site interdependency. He has developed a complex and brilliant bioinformatics algorithm to do that.

    The results seem very good, because the algorithm seems capable of identifying functionally related subunits just by analyzing sequences and their conservation, and those results correspond very well to what is known from structural studies.

    This is a wonderful demonstration of how focuisng on functional information can yield important results and be a new sensitive tool to understand the complex world of protein function.

    That’s why the reluctance of the neo-darwinist academy to even consider functional information as a real and tangible and measurable entity is a serious science stopper.

    This is a good example of cognitive bias, and of how a wrong ideological approach can make scientists blind to obvious avenues of research.

    It is not a case, of course, that Durston (and, probably, his co-workers) come from the ID field! 🙂

  165. DATCG:

    By the way, it’s also not a case, I believe, that he is using ubiquitn as the main testing molecule for his method. That certainly confirms not only the huge amount of functional information in its sequence, but also the complex and nuanced aspects of its biophysics profile.

  166. DATCG:

    I have read Abel’s “Fits” paper, referenced by you at #156.

    It is a very good summary of his thought, and of his personal contributions to ID theory.

    I absolutely agree with Abel’s ideas. They are clear and undeniable.

    I would like to explain here the only two points that I usually need to add in my discussions about my personal model of ID. They are in no way in contradiction with Abel’s discourse, but they are not usually explicit in his reasoning, at least as far as I can say.

    1) The first is that the role of consciousness is not made explicit. It is clearly implicit, for example here:

    Thus, FSC arises only out of wise choices at true decision nodes, logic gates, and purposeful configurable switch-settings. The latter can only be set by formalism, not physicality, if sophisticated function is to be realized. In the generation of FSC, not only must each successive choice opportunity be free from physicodynamic determinism, it must be deliberately chosen en route to achieving eventual formal and final function.

    Emphasis mine.

    Or here:

    The kind of Choice clearly observed everyday by everyone as a major component of reality includes Selection FOR (IN PURSUIT OF) not yet existent function. 8-11 Inanimate nature cannot exercise or generate such choice with intent. Only agency does. Mere mass/energy interactions have never been shown to produce the slightest hint of agency.

    We can see here a strange pattern that is often seen in design literature. There is a constant reference to ideas like choice, or agency, but they are not explicitly defined.

    The same is usually true for design itself: strange as it seems, it is usually not defined in ID literature.

    I have always felt that as a rpoblem, so the first OP I have published here was dedicated to:

    Defining design.

    https://uncommondescent.com/intelligent-design/defining-design/

    And here is my definition:

    Design is a process where a conscious agent subjectively represents in his own consciousness some form and then purposefully outputs that form, more or less efficiently, to some material object.

    We call the process “design”. We call the conscious agent who subjectively represents the initial form “designer”. We call the material object, after the process has taken place, “designed object”.

    I think that it is not possible to define “agency” or simply “design” without referring to conscious states.

    The reason is simple: conscious experiences are observable facts, and therefore have intrinsic scientific validity, while “agency”, “choice”, or even “design” are concepts, and need to be defined in terms of observable fatcs to be used in science.

    So, I define design in terms of conscious experiences (experience of meaning and of purpose) that are at the origin of the form outputted into material objects. That is an empirical and universal definition.

    2) The second point is that we need to set a quantitative threshold to functional complexity to use it to infer design.

    The reason is simple: if we want to allow for any definition of function (which I do in all my reasonings), we must acknoledge that very simple functions do exist.

    Now, very simple functions can be implemented by a few bits of information, and a few bits of information can certainly arise by random events.

    IOWs, a cloud can resemble a weasel, if the resemblance is not so strict that a lot of specific bits are involved.

    So, it is true, as Abel says, that:

    No empirical evidence exists of either RSC of OSC ever having produced a single instance of sophisticated function or true organization.

    (Emphasis mine)

    It’s absolutyely true! The only problem is that “sophisticated” is too vague.

    In science, we need an objective and possibly quantitative way to decide what is “sophisticated” and what is not.

    A quantitative approach is to define functional information as “complex” by an appropriate threshold. Usually, 500 bits are more than enough for any physical system in the universe. Realistically, much less is enough.

    A qualitative approach can be to assess semiosis, the presence of symbolic codes. Those structures are “qualitatively” beyond the reach of non conscious systems.

    The beauty of the Ubiquitin Systems is that it includes, in huge amounts, both those features: Functional Complexiy in the order of thousands or millions of bits, and undeniable and rich Semiosis.

    And, as I have argued at #128, Irreducible Complexity too. Tons of it! 🙂

  167. DATCG @160, gpuccio @162:

    Is this related?

    Gupta, Nilaksh & Li, Wei & Mcintyre, Thomas. (2015). Deubiquitinases Modulate Platelet Proteome Ubiquitination, Aggregation, and Thrombosis. Arteriosclerosis, thrombosis, and vascular biology. 35. 10.1161/ATVBAHA.115.306054.

    Objective:
    Platelets express a functional ubiquitin-proteasome system. Mass spectrometry shows that platelets contain several deubiquitinases, but whether these are functional, modulate the proteome, or affect platelet reactivity are unknown.

    Approach and results:
    Platelet lysates contained ubiquitin-protein deubiquitinase activity hydrolyzing both Lys48 and Lys63 polyubiquitin conjugates that was suppressed by the chemically unrelated deubiquitinase inhibitors PYR41 and PR619. These inhibitors acutely and markedly increased monoubiquitination and polyubiquitination of the proteome of resting platelets. PYR41 (intravenous, 15 minutes) significantly impaired occlusive thrombosis in FeCl3-damaged carotid arteries, and deubiquitinase inhibition reduced platelet adhesion and retention during high shear flow of whole blood through microfluidic chambers coated with collagen. Total internal reflection microscopy showed that adhesion and spreading in the absence of flow were strongly curtailed by these inhibitors with failure of stable process extension and reduced the retraction of formed clots. Deubiquitinase inhibition also sharply reduced homotypic platelet aggregation in response to not only the incomplete agonists ADP and collagen acting through glycoprotein VI but also to the complete agonist thrombin. Suppressed aggregation was accompanied by curtailed procaspase activating compound-1 binding to activated IIb/IIIa and inhibition of P-selectin translocation to the platelet surface. Deubiquitinase inhibition abolished the agonist-induced spike in intracellular calcium, suppressed Akt phosphorylation, and reduced agonist-stimulated phosphatase and tensin homolog phosphatase phosphorylation. Platelets express the proteasome-associated deubiquitinases USP14 and UCHL5, and selective inhibition of these enzymes by b-AP15 reproduced the inhibitory effect of the general deubiquitinase inhibitors on ex vivo platelet function.

    Conclusions:
    Remodeling of the ubiquitinated platelet proteome by deubiquitinases promotes agonist-stimulated intracellular signal transduction and platelet responsiveness.

  168. Mukherjee, Rukmini & Das, Aneesha & Chakrabarti, Saikat & Chakrabarti, Oishee. (2017). Calcium dependent regulation of protein ubiquitination – Interplay between E3 ligases and calcium binding proteins. Biochimica et Biophysica Acta (BBA) – Molecular Cell Research. 1864. 10.1016/j.bbamcr.2017.03.001.

    The ubiquitination status of proteins and intracellular calcium levels are two factors which keep changing inside any living cell. These two events appear to be independent of each other but recent experimental evidences show that ubiquitination of cellular proteins are influenced by calcium, Calmodulin, Calmodulin-dependent kinase II and other proteins of calcium dependent pathways. E3 ligases like Nedd4, SCF complex, APC, GP78 and ITCH are important regulators of calcium mediated processes. A bioinformatics analysis to inspect sequences and interacting partners of 242 candidate E3 ligases show the presence of calcium and/or Calmodulin binding motifs/domains within their sequences. Building a protein-protein interaction (PPI) network of human E3 ligase proteins identifies Ca2 + related proteins as direct interacting partners of E3 ligases. Review of literature, analysis of E3 ligase sequences and their interactome suggests an interconnectivity between calcium signaling and the overall UPS system, especially emphasizing that a subset of E3 ligases have importance in physiological pathways modulated by calcium.

  169. Ubiquitin conjugation probed by inflammation in MDSC extracellular vesicles

    R. Adams, Katherine & Chauhan, Sitara & B. Patel, Divya & K. Clements, Virginia & Wang, Yan & Jay, Steven & Edwards, Nathan & Ostrand-Rosenberg, S & Fenselau, Catherine. (2017). Ubiquitin conjugation probed by inflammation in MDSC extracellular vesicles. Journal of Proteome Research. 17. 10.1021/acs.jproteome.7b00585.

    Ubiquitinated proteins carried by the extracellular vesicles (EV) released by myeloid-derived suppressor cells (MDSC) have been investigated using proteomic strategies to examine the effect of tumor-associated inflammation. EV were collected from MDSC directly following isolation from tumor-bearing mice with low and high inflammation. Among 1092 proteins (high inflammation) and 925 proteins (low inflammation) identified, more than 50 % were observed as ubiquitinated proteoforms. More than three ubiquitin-attachment sites were characterized per ubiquitinated protein, on average. Multiple ubiquitination sites were identified in the pro-inflammatory proteins S100 A8 and S100 A9 characteristic of MDSC, and in histones and transcription regulators, among other proteins. Spectral counting and pathway analysis suggests that ubiquitination occurs independently of inflammation. Some ubiquitinated proteins were shown to cause migration of MDSC, which has been previously connected with immune suppression and tumor progression. Finally, MDSC EV are found collectively to carry all the enzymes required to catalyze ubiquitination, and the hypothesis is presented that a portion of the ubiquitinated proteins are produced in situ. Spectral counting and pathway analysis applied to the total protein content of inflammatory and conventional MDSC EV reveals significant enrichment in the proteins of the neutrophil degranulation pathway with induced inflammation.

  170. Dionisio @ 169: Please give me the simplified version of your comments 167 through 169. What is the main point you are making?

  171. TWSYF @170:

    Good question. Thanks for asking.

    Those referenced papers add more evidences for this argument:

    The known facts -not the unknown- definitely point to complex functionally specified informational complexity.

  172. Dionisio at #167:

    Paltelets are strange things. they are not really cells, but rather cell fragments. And yet, these small packages of cytoplasm are central in a lot of complex regulations involving the whole organism, especially coagulation and inflammation.

    I am not surprised at all that ubiquitin is well represented in the platelet repertoire, and that it could have important functions both in the platelet itself, and in cell to cell communication.

    Hematology has always been one of my favourite subjects in medicine! 🙂

  173. Dionisio at #168:

    Here again, no surprise. Calcium is one of the most important second messengers in cell signaling. It has fundamental roles in almost everything.

    We have already seen that the ubiquitin system and cell signaling are as connected as one can imagine. 🙂

  174. Truth Will Set You Free:

    I think the main point could be:

    We started with the idea that ubiquitin is really ubiquitous, but by digging and digging I would say that reality has greatly overcome our wildest imagination! 🙂

  175. Dionisio #167-168 and 169

    oh yes, good ones!

    I agree with Gpuccio’s comments. Thanks for the papers.

    And in posting the Journal of Immunology. Been a while since I looked directly at it. I usually stay buried at NCBI for searchers. Also Research Gate.

    Great additions!

  176. Gpuccio @164

    That’s why the reluctance of the neo-darwinist academy to even consider functional information as a real and tangible and measurable entity is a serious science stopper.

    This is a good example of cognitive bias, and of how a wrong ideological approach can make scientists blind to obvious avenues of research.

    Right, Agreed! The constant complaint by neo-darwinist is Design is a “science stopper,” but truth is Design is a great Science Starter.

    How do we know this? By all the scientist before us who sought design and purpose of function in the past.

    And modern day arguments of “junk” DNA. Design Theorist said we should find function in “junk” DNA. They did not predict all of it would be, but certainly more than neo-Darwinist who had written off “junk” DNA as leftover trash.

    Design is the better heuristic going forward in the age of ENCODE and molecular processing systems. It takes systems analysis of the whole to reverse engineer the myriad of organized, synchronized components that dance together in the cell, across cells, across networks, organs and forms.

    re: Durston, yes yes yes 🙂 It’s been years since I’ve ran Differential Equations through any kind of coding efforts, so I’m rusty on it. But I read his papers as well and was thrilled to see him tackle Function. I lurk here a lot, don’t usually comment to often at times. But always read anything from Durston, et al., and obviously post like yours 🙂

  177. Gpuccio #165…

    Durston’s, et al., Figure 2 Ubiquitin findings…

    Cluster tree for ubiquitin.

    The attribute clusters discovered from the aligned sequence data for the Ubiquitin family are shown above and organized vertically according to their order (the number of interdependent sites they contain). The organized clusters form primary branches, numbered 1 to 14 across the top of the figure. In each branch, the attribute cluster with the highest internal interdependency (highest SR(mode) value) was chosen as the representative cluster for that branch and is labeled according to its branch number. Two secondary clusters, discussed in the text, are labeled 12 s and 13 s. From this cluster tree, new insights can be gained into details of folding and structure.

  178. Durston, continued…

    Ubiquitin (A) 3D Structure using 1UBQ Solved Structure, (B) Module 1 Clusters, (C) Cluster 12 s, discovered by K-modes Algorithm

    Ubiquitin:

    (A) The 3D structure of ubiquitin, using the 1UBQ solved structure. Part of cluster 13 s is also shown within the ubiquitin molecule. The three sites are all within van der Waals interaction distance of each other and may have an important role in the stability of the overall structure.

    (B) Module 1. This module contains two H-bonded clusters, which may play a role both in folding and then maintaining structural stability once folding is complete, especially cluster 1 with six H-bonds anchoring the loop.

    (C) Cluster 12 s, a strong example of a van der Waals cluster discovered by the k-modes algorithm.

    .

  179. Durston, Wong, Li Chui, continued…

    Graphic representation…

    Fig 4 Secondary structure of ubiquitin with locations of modules and major clusters.

    The location of the major clusters associated with the four major modules is shown, for clarity, on two identical secondary structure ribbons, taken from model 1UBQ.

    An example of smaller clusters nested within larger clusters can be seen in the clusters associated with Module 2.

    The attribute (site) clusters associated with Modules 1 and 2 were found to be compact, with only a small region of interlacing involving sites 26, 28, and 31.

    Of particular interest are the two clusters associated with Module 4 that contain some sites that are quite distant in the primary structure. The key sites in these very extended clusters are sites 30, 43, and 69 which, although widely separated in the primary sequence, are actually in Van der Waals contact, as shown in Figure 3.

  180. BTW re: David Abel,

    While I’d followed most of his publshed papers, I did not know he had the following site:

    Peer-reviewed publications of David L. Abel

    More reading! 😉

  181. Dionisio, DATCG, Upright Biped:

    This is brand new (14 February 2018) about the possible connections between E3 ligases mutations and human neurological disorders:

    A Comprehensive Atlas of E3 Ubiquitin Ligase Mutations in Neurological Disorders

    Abstract:

    Protein ubiquitination is a posttranslational modification that plays an integral part in mediating diverse cellular functions. The process of protein ubiquitination requires an enzymatic cascade that consists of a ubiquitin activating enzyme (E1), ubiquitin conjugating enzyme (E2) and an E3 ubiquitin ligase (E3). There are an estimated 600-700 E3 ligase genes representing ~5% of the human genome. Not surprisingly, mutations in E3 ligase genes have been observed in multiple neurological conditions. We constructed a comprehensive atlas of disrupted E3 ligase genes in common (CND) and rare neurological diseases (RND). Of the predicted and known human E3 ligase genes, we found ~13% were mutated in a neurological disorder with 83 total genes representing 70 different types of neurological diseases. Of the E3 ligase genes identified, 51 were associated with an RND. Here, we provide an updated list of neurological disorders associated with E3 ligase gene disruption. We further highlight research in these neurological disorders and discuss the advanced technologies used to support these findings.

  182. gpuccio @182:

    That paper just came out of the printing press!
    The ink hasn’t dried out yet.

    This reminds me of something you wrote @87:

    So, ubiquitin is becoming ubiquitous. There is some logic in that.
    RV + NS can certainly explain that all!

  183. Gpuccio @182,

    oooooo… followed your paper link and found another by
    same arthor, who responded to a question.

    She recommended this link: UbiNet Not to be confused
    with SkyNet 😉

    UbiNet the Next Net frontier in Ubiquitin Services

    Haha, seriously though. A site full of resources. Had not seen this.

    from the site…

    UbiNet is a knowledge-based system that could provide potential E3 ligases for ubiquitinated proteins by the information of protein-protein interactions and substrate site specificities. In eukaryotes, protein ubiquitination catalyzed by E3 ubiquitin ligases plays crucial roles in regulating a variety of biological processes. With the high-throughput of mass spectrometry-based proteomics, a number of methods have been developed to experimentally determine the ubiquitination sites, leading to an increasing of large-scale ubiquitinome data. However, at the moment, there exists no resource designed to explore the regulatory networks of E3 ligases for ubiquitinated proteins in humans. Therefore, the UbiNet was designed to provide a full investigation of protein ubiquitination networks by integrating all available ubiquitinome datasets, experimentally verified E3 ligases, and protein-protein interactions. Moreover, UbiNet could provide potential E3 ligases for ubiquitinated proteins by the information of protein-protein interactions and substrate site specificities. The UbiNet could help users to identify E3 ligase-mediated ubiquitination networks and their roles in biological processes.

    never know what you will find down the wabbit hole…

    It has a lovely FUNCTION button to click on that leads you
    to this page…

    Functions Galore of Ubiquitin and Cool Categories to Choose from, including Cellular Process

    Organ Development is one of them. Had we already added that to the UBQ List?

    Also, MAP3K

    Enjoy! 🙂

    And thanks for the link on Neurological Disorders.

    70 different types of neurological disorders. Wow!

    I wonder in the future, how many diseases and disorders across the spectrum of the Genome is due to Epigenetic mutations? Which are deleterious to active configurations, passed down as a result of environmental queues or bad actions of the parent, for example like Fetal Alcohol Syndrome.

    Or how the old saying goes, You Are What You Eat or Drink

    .

  184. #182 Dionisio,

    Good day 🙂 Check out the UbiNet Link I posted above, then click Cellular Process Tab at top and voila…

    organ morphogenesis

    surely you jest, RM+NS solves everything! 😉

  185. TWSYF @170:
    Valid request. Thanks.

    The best comments related to your valid request were written by GP @173-175 & DATCG @176.

  186. DATCG @185:

    Thank you for such a good information.
    I’m glad you found that resource and shared it with us here right away. It seems like you’ve discovered a rich mine. Well done!
    Have a good weekend.

  187. This excellent 100% scientific thread -started and maintained by gpuccio with very helpful contributions by DATCG and other folks- seems to confirm the Big Data issue associated with Biology research lately. The overwhelming avalanche of data coming out of wet and dry labs seems to demand more multidisciplinary research teams working really hard to try figuring out how to understand all that information.
    We ain’t seen nothin’ yet. The most fascinating discoveries are still ahead.

  188. You too Dionisio!

    Yeah, it really ties Ubiquitin and it’s impact altogether
    in a neat, easy format.

    I wonder if they’re using Durston’s, Wong, et al., K-mode algorithm. Hmmmm…

  189. Dionisio @188,

    And how much of the former “junk” has been fully explored yet?

    I want to keep reminding people of Dan Graur’s epic meltdown…

    “If ENCODE is right, evolution is wrong!”

    I’m curious if anyone is tracking the artificial threshold he set on Epigenetic Function vs “JUNK”

    Because anytime researchers find new function in previous areas thought to be junk, it opens new doors for more research in those new areas.

    You’re so right… the future’s so bright, the Darwinist will have to wear Shades… on top of their Blinders.

  190. DATCG @184:

    The interesting resource you have found seems to be produced by this university:

    UbiNet’s Team: Van-Nui Nguyen, Kai-Yao Huang, Prof. Tzong-Yi Lee, and Prof. K. Robert Lai: Department of Computer Science and Engineering, Yuan Ze University.
    https://www.yzu.edu.tw/

    Thanks for sharing your discovery right away.

    I certainly can use it.

  191. gpuccio,

    I would like to read your assessment of the interesting resource found by DATCG.
    No rush. Take your time.

    Thanks.

  192. TWSYF @170:

    You may enjoy reading what DATCG just wrote @190:

    “[…] the future’s so bright, the Darwinist will have to wear Shades… on top of their Blinders”

    Really funny. DATCG has a healthy sense of humor.

  193. DATCG at #184:

    UbiNet! Great resource, thank you! 🙂

    I am avidly exploring it.

    It was great ofnyou to find it. I had looked for U3 ligases databases, but did not find it.

    As usual, I like the general statistics a lot:

    Ubiquitinated proteins 14,692
    – Ubiquitination sites 43,948
    E1 (activating enzyme) 2
    E2 (conjugating enzyme) 46
    E3 (ubiquitin ligase) 499
    Protein-protein interaction 430,530
    Domain-domain interaction 286,758
    E3-associated functional category 2,143
    Literatures 44,184

    Emphasis mine.

    So, it seems that our idea that most proteins undergo some ubiquitination, at some moment in their history, is not too wild!

  194. Dionisio:

    “That paper just came out of the printing press!
    The ink hasn’t dried out yet.”

    Then look at this one (1 March 2018):

    Degradation for better survival? Role of ubiquitination in epithelial morphogenesis.

    http://onlinelibrary.wiley.com.....4/abstract

    ABSTRACT:

    As a prevalent post-translational modification, ubiquitination is essential for many developmental processes. Once covalently attached to the small and conserved polypeptide ubiquitin (Ub), a substrate protein can be directed to perform specific biological functions via its Ub-modified form. Three sequential catalytic reactions contribute to this process, among which E3 ligases serve to identify target substrates and promote the activated Ub to conjugate to substrate proteins. Ubiquitination has great plasticity, with diverse numbers, topologies and modifications of Ub chains conjugated at different substrate residues adding a layer of complexity that facilitates a huge range of cellular functions. Herein, we highlight key advances in the understanding of ubiquitination in epithelial morphogenesis, with an emphasis on the latest insights into its roles in cellular events involved in polarized epithelial tissue, including cell adhesion, asymmetric localization of polarity determinants and cytoskeletal organization. In addition, the physiological roles of ubiquitination are discussed for typical examples of epithelial morphogenesis, such as lung branching, vascular development and synaptic formation and plasticity. Our increased understanding of ubiquitination in epithelial morphogenesis may provide novel insights into the molecular mechanisms underlying epithelial regeneration and maintenance.

    (Paywall)

    The paper is very good.

    I would like to quote this passage which sums up some of the combinatorial potentialities of the ubiquitin system:

    From a mathematical point of view, with the availability of a few types of E1 ligase, dozens of E2 ligases, and hundreds of E3 ligases, even if each is used in only one reaction, there are probably over 50000 different permutations that can each potentially generate a unique Ub code. Similarly, for
    a Ub chain length of between 1 and at least 10, with seven lysine residues and an N-terminus serving as linkage points, and the possibility of forming single or multiple, linear or branched chains adopting open or closed configurations, there are theoretically millions of different permutations for Ub codes. This variety of Ub modification, together with other post-translational modifications, might partially
    compensate for relative genomic simplicity (e.g. human
    DNA encodes only 25000 genes) to allow the functional
    complexity of higher organisms.

    So, in brief, we have a potential of:

    a) Over 50000 different permutations for the writing system

    b) Millions of different permutations for the code itself

    That’s quite a potential! 🙂

  195. Dionisio:

    Knowing your interests, I though you could like this part of the “Contents” section of the paper referenced at #195:

    III. Ubiquitination in cellular events of epithelial morphogenesis

    1) Cell adhesion
    a) Cell–ECM interaction
    b) Cell–cell interaction
    2) Asymmetric localization of polarity complexes
    a) Planar polarity components
    b) Apico-basal polarity components
    3) Cytoskeleton organization
    a) Microtubules
    b) Actinfilaments
    c) Intermediate filaments

  196. gpuccio @195,

    You pulled that paper out of the printing press and got still-wet ink spilled all over! 🙂

    And Wow! that’s quite a rich code!

    Now, what determines which of those combinations is used where and when and for what so that things fit nicely?

    Please, note that the tricky word ‘exactly’ is not in the above question so that it qualifies as honest. 🙂

    Thanks.

  197. gpuccio @196:

    Yes, you thought it right: that’s a very interesting topic for me. Thank you for pointing to that paper.

  198. Gpuccio,

    Curious, which group of Proteins are NOT degraded, or tagged for destruction by Ubiquitin tagging?

    Even as you pointed out, Apoptosis is at some point impacted by Ubiquitination processes.

    Even if not by direct interaction, it appears 2nd, 3rd levels along the path can involve different forms of mono or poly-ubiquitin chains that guide or modify the pathway to eventual degradation or recyling, etc.

    Am I going to far in contemplating, all Proteins may have some form of interaction? Either direct or at a distance with the Ubiquitin Code?

    .

  199. gpuccio @89:

    Always for lovers of simplicity, I recommend Fig. 3, which covers Ubiquitin enzymes in the T Cell Receptor signaling pathway.
    The B cell pathway is also covered in Fig. 5.
    Fig. 6 is, again, about the NK-kB pathway, which is central in many immunity related processes.
    And Fig. 7 is about our old friend, TRIM.

    For lovers of simplicity?

    Yes, quite simple indeed:

    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5490640/figure/F0003/

    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5490640/figure/F0005/

    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5490640/figure/F0006/

    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5490640/figure/F0007/

  200. UB @36, @42, @44, @94:

    (Attn. DATCG @101 too)

    It seems like both Barbieri and Pattee made some interesting and valid affirmations, but overall they still went off the tangent and drew the wrong conclusions.

    I assume UB referred to this person:
    http://binghamton.academia.edu/HowardPattee

  201. Let me repeat a comment that was posted @21:
    “This thread has too much bad news for the ‘modern synthesis’ and the ‘third way’ clubs.”

  202. Dionisio:

    I agree: interesting arguments, interesting discussions, and wrong conclusions.

    Why?

    That’s the only thing that can happen when intelligent and honest people start from some acritical assumption, which can never be questioned, even if it is not based on any evidence: so, all wrong consclusions in his reasonings simply derive from accepting as a dogma that evolution happened by the imaginary mechanisms of the neo-darwinian theory.

    A wrong assumption is not only a science stopper: it is a thought stopper!

  203. Dionisio:

    “This thread has too much bad news for the ‘modern synthesis’ and the ‘third way’ clubs.”

    Maybe. So, it seems they are definitely recurring to some Ostrich policy! 🙂

    https://en.wikipedia.org/wiki/Ostrich_policy

  204. gpuccio @133:

    No, zotero is not a designed object.

    That software resulted from a bunch of pieces of code accidentally thrown in by many different programmers from around the world, without any goal in mind, completely unguided. We were lucky that it somehow turned out useful. It’s called “emergent functionality”, in case you don’t know it. I see you don’t understand evolution. Go and take some basic biology 101. That should help you next time.

    🙂

    PS. my cheek hurts badly

  205. gpuccio @204:

    Ostrich policy!

    That explains their conspicuous absence from these discussions!

    Thanks!

  206. gpuccio @203:

    Yes, agree.

  207. Dio at 201

    Yep. Pattee was born in 1921 and graduated from Stanford in 1948. Barbieri was born in 1940 and graduated from Ferrara in 1964.

    They ended up with the only conclusion that was professionally allowed to them. That’s the ideological damage done to science. Beyond saying they were “men of their times”, I do not know how to properly classify them. I know that I am personally grateful to both.

  208. Upright BiPed:

    “I know that I am personally grateful to both.”

    And you are right to be grateful. Intelligent and creative argumentation is always precious, even if it takes place in a wrong context. 🙂

  209. gpuccio @134:

    “By the way, Figures 3,4, 5 and 6 (and their respective legends) are some more fun for the lovers of simplicity!”

    Fig. 1

    Fig. 2

    Fig. 3

    Fig. 4

    Fig. 5

    Fig. 6

    Yes, very simple indeed!

  210. Dionisio, Gpuccio, Upright Biped,

    re: Barbieri and Pattee

    UB said,

    They ended up with the only conclusion that was professionally allowed to them. That’s the ideological damage done to science. Beyond saying they were “men of their times”, I do not know how to properly classify them. I know that I am personally grateful to both.

    So true in being grateful. Seeing Barbieri's Code Biology is refreshing even if he refuses to accept Design. At least he acknowledges signs and signals cannot exist without symbolic representation and meaning. Even if the tries to "naturalize" it.

    Question, although we have places like UD and Discovery Institute today, are the times worse today for scientist?

    As was seen in recent past and often we forget damage done in harsher treatment. For example, locking a scientist out of his office?

    Richard Steinberg’s wretched treatment at Smithsonian

    I lost my office space.
    I was twice forced to move specimens from my office space on short notice for no good reason, my name plate was removed from my office door, and eventually I was deprived of all official office space and forced to use a shared work area as my work location in the Museum.

    With those kind of treatments, discouraging open and free thought by scientist is it any wonder most would not “go there” to Design Theory?

    Moving on…

    Maybe, a modern day version of Babieri and Pattee is James Shapiro? of Third Way and “Natural” Genetic Engineering?

    Since when is engineering a “natural” function without conscious thought? Without Design principles firmly glued to the functional thought process of mindful forethought of an Engineer?

    Only when as in our times, it is not allowed without damaging repercussions to one’s job, livelihood and professional reputation.

    Or, is Shapiro like Darwin? An end-run attempt around well known Design principles? Mainly around Code, Semantics, Semiosis and Engineering principles, but prefaced with “Natural” as an internal edification to his soul because he cannot accept it any other way?

    I’ve not read any of his personal musings on it, but it seems even though we have the power of the Internet and brave people are fighting a good fight in these times. Still scientist are rarely allowed to speak openly without severe consequences and shaming. And most do not do so unless retired, or protected by tenure.

    And who can blame them? Maybe as Dr. Sanford and others, many will quietly publish and “come out” when they can in the future.

    Pattee and Barieri could not but help admit partially the truth before them.

    This, even if they refuse the truth before them still helps our cause. The Thought Minders hate it.

    Viva pensiero libero 🙂

    .

  211. DATCG @156:

    Does that paper have DOI code?

  212. DATCG @157,

    Is that the same paper referenced @178-180 too?

  213. List of papers referenced by GP in comments posted in this thread:

    1. Boisvert, François-Michel, Yasmeen Ahmad, Marek Gierli?ski, Fabien Charrière, Douglas Lamont, Michelle Scott, Geoff Barton, and Angus I. Lamond. “A Quantitative Spatial Proteomics Analysis of Proteome Turnover in Human Cells.” Molecular & Cellular Proteomics 11, no. 3 (March 2012): M111.011429. https://doi.org/10.1074/mcp.M111.011429.

    2. Cao, Jian, and Qin Yan. “Histone Ubiquitination and Deubiquitination in Transcription, DNA Damage Response, and Cancer.” Frontiers in Oncology 2 (2012). https://doi.org/10.3389/fonc.2012.00026.

    3. Cheng, Xiaoxiang, Jun Zheng, Gang Li, Verena Göbel, and Hongjie Zhang. “Degradation for Better Survival? Role of Ubiquitination in Epithelial Morphogenesis: Ubiquitination in Epithelial Morphogenesis.” Biological Reviews, March 1, 2018. https://doi.org/10.1111/brv.12404.

    4. Choi, Jihye, and Kwang-Hyun Baek. “Cellular Functions of Stem Cell Factors Mediated by the Ubiquitin–proteasome System.” Cellular and Molecular Life Sciences, February 8, 2018. https://doi.org/10.1007/s00018-018-2770-7.

    5. Citterio, Elisabetta. “Fine-Tuning the Ubiquitin Code at DNA Double-Strand Breaks: Deubiquitinating Enzymes at Work.” Frontiers in Genetics 6 (September 8, 2015). https://doi.org/10.3389/fgene.2015.00282.

    6. Clague, Michael J., Claire Heride, and Sylvie Urbé. “The Demographics of the Ubiquitin System.” Trends in Cell Biology 25, no. 7 (July 2015): 417–26. https://doi.org/10.1016/j.tcb.2015.03.002.

    7. Darwin, K. Heran. “Prokaryotic Ubiquitin-like Protein (Pup), Proteasomes and Pathogenesis.” Nature Reviews Microbiology 7, no. 7 (July 2009): 485–91. https://doi.org/10.1038/nrmicro2148.

    8. Donovan, Prudence, and Philip Poronnik. “Nedd4 and Nedd4-2: Ubiquitin Ligases at Work in the Neuron.” The International Journal of Biochemistry & Cell Biology 45, no. 3 (March 2013): 706–10. https://doi.org/10.1016/j.biocel.2012.12.006.

    9. Durston, Kirk K, David KY Chiu, David L Abel, and Jack T Trevors. “Measuring the Functional Sequence Complexity of Proteins.” Theoretical Biology and Medical Modelling 4, no. 1 (2007): 47. https://doi.org/10.1186/1742-4682-4-47.

    10. Ebner, Petra, Gijs A. Versteeg, and Fumiyo Ikeda. “Ubiquitin Enzymes in the Regulation of Immune Responses.” Critical Reviews in Biochemistry and Molecular Biology 52, no. 4 (July 4, 2017): 425–60. https://doi.org/10.1080/10409238.2017.1325829.

    11. Gao, Si-Fa, Bo Zhong, and Dandan Lin. “Regulation of T Helper Cell Differentiation by E3 Ubiquitin Ligases and Deubiquitinating Enzymes.” International Immunopharmacology 42 (January 2017): 150–56. https://doi.org/10.1016/j.intimp.2016.11.013.

    12. George, Arlene J., Yarely C. Hoffiz, Antoinette J. Charles, Ying Zhu, and Angela M. Mabb. “A Comprehensive Atlas of E3 Ubiquitin Ligase Mutations in Neurological Disorders.” Frontiers in Genetics 9 (February 14, 2018). https://doi.org/10.3389/fgene.2018.00029.

    13. Guharoy, Mainak, Pallab Bhowmick, and Peter Tompa. “Design Principles Involving Protein Disorder Facilitate Specific Substrate Selection and Degradation by the Ubiquitin-Proteasome System.” Journal of Biological Chemistry 291, no. 13 (March 25, 2016): 6723–31. https://doi.org/10.1074/jbc.R115.692665.

    14. Gupta, Ishita, Kanika Singh, Nishant K. Varshney, and Sameena Khan. “Delineating Crosstalk Mechanisms of the Ubiquitin Proteasome System That Regulate Apoptosis.” Frontiers in Cell and Developmental Biology 6 (February 9, 2018). https://doi.org/10.3389/fcell.2018.00011.

    15. Hallengren, Jada, Ping-Chung Chen, and Scott M. Wilson. “Neuronal Ubiquitin Homeostasis.” Cell Biochemistry and Biophysics 67, no. 1 (September 2013): 67–73. https://doi.org/10.1007/s12013-013-9634-4.

    16. Hatakeyama, Shigetsugu. “TRIM Family Proteins: Roles in Autophagy, Immunity, and Carcinogenesis.” Trends in Biochemical Sciences 42, no. 4 (April 2017): 297–311. https://doi.org/10.1016/j.tibs.2017.01.002.

    17. Hu, Ming-Ming, and Hong-Bing Shu. “Multifaceted Roles of TRIM38 in Innate Immune and Inflammatory Responses.” Cellular & Molecular Immunology 14, no. 4 (April 2017): 331–38. https://doi.org/10.1038/cmi.2016.66.

    18. “Identification of Top-Ranked Proteins within a Directional Protein Interaction Network Using the PageRank Algorithm: Applications in Humans and Plants.” Current Issues in Molecular Biology, 2016. https://doi.org/10.21775/cimb.020.013.

    19. Lee, Robin van der, Marija Buljan, Benjamin Lang, Robert J. Weatheritt, Gary W. Daughdrill, A. Keith Dunker, Monika Fuxreiter, et al. “Classification of Intrinsically Disordered Regions and Proteins.” Chemical Reviews 114, no. 13 (July 9, 2014): 6589–6631. https://doi.org/10.1021/cr400525m.

    20. Li, Tao, Linsheng Wang, Yongming Du, Si Xie, Xi Yang, Fuming Lian, Zhongjun Zhou, and Chengmin Qian. “Structural and Mechanistic Insights into UHRF1-Mediated DNMT1 Activation in the Maintenance DNA Methylation.” Nucleic Acids Research, February 19, 2018. https://doi.org/10.1093/nar/gky104.

    21. Liu, Jiangang, Narayanan B. Perumal, Christopher J. Oldfield, Eric W. Su, Vladimir N. Uversky, and A. Keith Dunker. “Intrinsic Disorder in Transcription Factors †.” Biochemistry 45, no. 22 (June 2006): 6873–88. https://doi.org/10.1021/bi0602718.

    22. Maupin-Furlow, Julie A. “Prokaryotic Ubiquitin-Like Protein Modification.” Annual Review of Microbiology 68, no. 1 (September 8, 2014): 155–75. https://doi.org/10.1146/annurev-micro-091313-103447.

    23. Ohtake, Fumiaki, Yasushi Saeki, Satoshi Ishido, Jun Kanno, and Keiji Tanaka. “The K48-K63 Branched Ubiquitin Chain Regulates NF-?B Signaling.” Molecular Cell 64, no. 2 (October 2016): 251–66. https://doi.org/10.1016/j.molcel.2016.09.014.

    24. Ong, Taren, and David J. Solecki. “Seven in Absentia E3 Ubiquitin Ligases: Central Regulators of Neural Cell Fate and Neuronal Polarity.” Frontiers in Cellular Neuroscience 11 (October 13, 2017). https://doi.org/10.3389/fncel.2017.00322.

    25. Park, Yoon, Hyung-seung Jin, Daisuke Aki, Jeeho Lee, and Yun-Cai Liu. “The Ubiquitin System in Immune Regulation.” In Advances in Immunology, 124:17–66. Elsevier, 2014. https://doi.org/10.1016/B978-0-12-800147-9.00002-9.

    26. Saeki, Yasushi. “Ubiquitin Recognition by the Proteasome.” Journal of Biochemistry, January 8, 2017, mvw091. https://doi.org/10.1093/jb/mvw091.

    27. Sammak, Susan, and Giovanna Zinzalla. “Targeting Protein–protein Interactions (PPIs) of Transcription Factors: Challenges of Intrinsically Disordered Proteins (IDPs) and Regions (IDRs).” Progress in Biophysics and Molecular Biology 119, no. 1 (October 2015): 41–46. https://doi.org/10.1016/j.pbiomolbio.2015.06.004.

    28. Smeenk, Godelieve, and Niels Mailand. “Writers, Readers, and Erasers of Histone Ubiquitylation in DNA Double-Strand Break Repair.” Frontiers in Genetics 7 (June 28, 2016). https://doi.org/10.3389/fgene.2016.00122.

    29. Su, Xiaomin, Chenglei Wu, Xiaoying Ye, Ming Zeng, Zhujun Zhang, Yongzhe Che, Yuan Zhang, Lin Liu, Yushuang Lin, and Rongcun Yang. “Embryonic Lethality in Mice Lacking Trim59 Due to Impaired Gastrulation Development.” Cell Death & Disease 9, no. 3 (March 2018). https://doi.org/10.1038/s41419-018-0370-y.

    30. van der Lee, Robin, Benjamin Lang, Kai Kruse, Jörg Gsponer, Natalia Sánchez de Groot, Martijn A. Huynen, Andreas Matouschek, Monika Fuxreiter, and M. Madan Babu. “Intrinsically Disordered Segments Affect Protein Half-Life in the Cell and during Evolution.” Cell Reports 8, no. 6 (September 2014): 1832–44. https://doi.org/10.1016/j.celrep.2014.07.055.

    31. Yamada, Tomoko, Yue Yang, and Azad Bonni. “Spatial Organization of Ubiquitin Ligase Pathways Orchestrates Neuronal Connectivity.” Trends in Neurosciences 36, no. 4 (April 2013): 218–26. https://doi.org/10.1016/j.tins.2012.12.004.

    32. Yruela, Inmaculada, Christopher J. Oldfield, Karl J. Niklas, and A. Keith Dunker. “Evidence for a Strong Correlation Between Transcription Factor Protein Disorder and Organismic Complexity.” Genome Biology and Evolution 9, no. 5 (May 2017): 1248–65. https://doi.org/10.1093/gbe/evx073.

    33. Yu, Houqing, and Andreas Matouschek. “Recognition of Client Proteins by the Proteasome.” Annual Review of Biophysics 46, no. 1 (May 22, 2017): 149–73. https://doi.org/10.1146/annurev-biophys-070816-033719.

    34. Zhou, Bangjun, and Lirong Zeng. “Conventional and Unconventional Ubiquitination in Plant Immunity: Ubiquitination in Plant Immunity.” Molecular Plant Pathology 18, no. 9 (December 2017): 1313–30. https://doi.org/10.1111/mpp.12521.

    Please, correct any inaccuracies or omissions in the above list. Thanks.
    Please, note that the above list does not include papers referenced in the OP.

  214. List of papers referenced by GP in comments posted in this thread:

    George et al., “A Comprehensive Atlas of E3 Ubiquitin Ligase Mutations in Neurological Disorders”;

    Boisvert et al., “A Quantitative Spatial Proteomics Analysis of Proteome Turnover in Human Cells”;

    Choi and Baek, “Cellular Functions of Stem Cell Factors Mediated by the Ubiquitin–proteasome System”;

    van der Lee et al., “Classification of Intrinsically Disordered Regions and Proteins”;

    Zhou and Zeng, “Conventional and Unconventional Ubiquitination in Plant Immunity”;

    Cheng et al., “Degradation for Better Survival?”;

    Gupta et al., “Delineating Crosstalk Mechanisms of the Ubiquitin Proteasome System That Regulate Apoptosis”;

    Guharoy, Bhowmick, and Tompa, “Design Principles Involving Protein Disorder Facilitate Specific Substrate Selection and Degradation by the Ubiquitin-Proteasome System”;

    Su et al., “Embryonic Lethality in Mice Lacking Trim59 Due to Impaired Gastrulation Development”;

    Yruela et al., “Evidence for a Strong Correlation Between Transcription Factor Protein Disorder and Organismic Complexity”;

    Citterio, “Fine-Tuning the Ubiquitin Code at DNA Double-Strand Breaks”;

    Cao and Yan, “Histone Ubiquitination and Deubiquitination in Transcription, DNA Damage Response, and Cancer”;

    “Identification of Top-Ranked Proteins within a Directional Protein Interaction Network Using the PageRank Algorithm”;

    Liu et al., “Intrinsic Disorder in Transcription Factors †”;

    van der Lee et al., “Intrinsically Disordered Segments Affect Protein Half-Life in the Cell and during Evolution”;

    Durston et al., “Measuring the Functional Sequence Complexity of Proteins”;

    Hu and Shu, “Multifaceted Roles of TRIM38 in Innate Immune and Inflammatory Responses”;

    Donovan and Poronnik, “Nedd4 and Nedd4-2”;

    Hallengren, Chen, and Wilson, “Neuronal Ubiquitin Homeostasis”;

    Darwin, “Prokaryotic Ubiquitin-like Protein (Pup), Proteasomes and Pathogenesis”;

    Maupin-Furlow, “Prokaryotic Ubiquitin-Like Protein Modification”;

    Yu and Matouschek, “Recognition of Client Proteins by the Proteasome”;

    Gao, Zhong, and Lin, “Regulation of T Helper Cell Differentiation by E3 Ubiquitin Ligases and Deubiquitinating Enzymes”;

    Ong and Solecki, “Seven in Absentia E3 Ubiquitin Ligases”;

    Yamada, Yang, and Bonni, “Spatial Organization of Ubiquitin Ligase Pathways Orchestrates Neuronal Connectivity”;

    Li et al., “Structural and Mechanistic Insights into UHRF1-Mediated DNMT1 Activation in the Maintenance DNA Methylation”;

    Sammak and Zinzalla, “Targeting Protein–protein Interactions (PPIs) of Transcription Factors”;

    Clague, Heride, and Urbé, “The Demographics of the Ubiquitin System”;

    Ohtake et al., “The K48-K63 Branched Ubiquitin Chain Regulates NF-?B Signaling”;

    Park et al., “The Ubiquitin System in Immune Regulation”;

    Hatakeyama, “TRIM Family Proteins”; Ebner, Versteeg, and Ikeda, “Ubiquitin Enzymes in the Regulation of Immune Responses”;

    Saeki, “Ubiquitin Recognition by the Proteasome”;

    Smeenk and Mailand, “Writers, Readers, and Erasers of Histone Ubiquitylation in DNA Double-Strand Break Repair.”

    Please, correct any inaccuracies or omissions in the above list. Thanks.
    Please, note that the above list does not include papers referenced in the OP.

  215. List of papers referenced by DATCG in comments posted in this thread:

    1. Abel, David L., and Jack T. Trevors. “Three Subsets of Sequence Complexity and Their Relevance to Biopolymeric Information.” Theoretical Biology and Medical Modelling 2, no. 1 (August 11, 2005): 29. https://doi.org/10.1186/1742-4682-2-29.

    2. Durston, Kirk K, David KY Chiu, Andrew KC Wong, and Gary CL Li. “Statistical Discovery of Site Inter-Dependencies in Sub-Molecular Hierarchical Protein Structuring.” EURASIP Journal on Bioinformatics and Systems Biology 2012, no. 1 (December 2012). https://doi.org/10.1186/1687-4153-2012-8.

    3. Pla, A, M Pascual, J Renau-Piqueras, and C Guerri. “TLR4 Mediates the Impairment of Ubiquitin-Proteasome and Autophagy-Lysosome Pathways Induced by Ethanol Treatment in Brain.” Cell Death & Disease 5, no. 2 (February 2014): e1066–e1066. https://doi.org/10.1038/cddis.2014.46.

    4. Rogers, J. M., V. Oleinikovas, S. L. Shammas, C. T. Wong, D. De Sancho, C. M. Baker, and J. Clarke. “Interplay between Partner and Ligand Facilitates the Folding and Binding of an Intrinsically Disordered Protein.” Proceedings of the National Academy of Sciences 111, no. 43 (October 28, 2014): 15420–25. https://doi.org/10.1073/pnas.1409122111.

    5. Ruiz i Altaba, Ariel, Vân Nguyên, and Verónica Palma. “The Emergent Design of the Neural Tube: Prepattern, SHH Morphogen and GLI Code.” Current Opinion in Genetics & Development 13, no. 5 (October 2003): 513–21. https://doi.org/10.1016/j.gde.2003.08.005.

    6. Savage, Kienan I., and D. Paul Harkin. “BRCA1, a ‘Complex’ Protein Involved in the Maintenance of Genomic Stability.” The FEBS Journal 282, no. 4 (February 2015): 630–46. https://doi.org/10.1111/febs.13150.

    7. Srikanthan, S., W. Li, R. L. Silverstein, and T. M. McIntyre. “Exosome Poly-Ubiquitin Inhibits Platelet Activation, Downregulates CD36 and Inhibits pro-Atherothombotic Cellular Functions.” Journal of Thrombosis and Haemostasis 12, no. 11 (November 2014): 1906–17. https://doi.org/10.1111/jth.12712.

    8. Uversky, Vladimir N. “Functional Roles of Transiently and Intrinsically Disordered Regions within Proteins.” FEBS Journal 282, no. 7 (April 2015): 1182–89. https://doi.org/10.1111/febs.13202.

    9. Wang, Yi-Ting, and Guang-Chao Chen. “The Role of Ubiquitin System in Autophagy.” In Autophagy in Current Trends in Cellular Physiology and Pathology, edited by Nikolai V. Gorbunov and Marion Schneider. InTech, 2016. https://doi.org/10.5772/64728.

    Please, correct any inaccuracies or omissions in the above list. Thanks.

  216. List of papers referenced by Dionisio in comments posted in this thread:

    1. Adams, Katherine R., Sitara Chauhan, Divya B. Patel, Virginia K. Clements, Yan Wang, Steven M. Jay, Nathan J. Edwards, Suzanne Ostrand-Rosenberg, and Catherine Fenselau. “Ubiquitin Conjugation Probed by Inflammation in Myeloid-Derived Suppressor Cell Extracellular Vesicles.” Journal of Proteome Research 17, no. 1 (January 5, 2018): 315–24. https://doi.org/10.1021/acs.jproteome.7b00585.

    2. Bremer, Anne, Martin Wolff, Anja Thalhammer, and Dirk K. Hincha. “Folding of Intrinsically Disordered Plant LEA Proteins Is Driven by Glycerol-Induced Crowding and the Presence of Membranes.” The FEBS Journal 284, no. 6 (March 2017): 919–36. https://doi.org/10.1111/febs.14023.

    3. Burroughs, A. Maxwell, Lakshminarayan M. Iyer, and L. Aravind. “Structure and Evolution of Ubiquitin and Ubiquitin-Related Domains.” In Ubiquitin Family Modifiers and the Proteasome, edited by R. Jürgen Dohmen and Martin Scheffner, 832:15–63. Totowa, NJ: Humana Press, 2012. https://doi.org/10.1007/978-1-61779-474-2_2.

    4. Chatzidaki-Livanis, Maria, Michael J. Coyne, Kevin G. Roelofs, Rahul R. Gentyala, Jarreth M. Caldwell, and Laurie E. Comstock. “Gut Symbiont Bacteroides Fragilis Secretes a Eukaryotic-Like Ubiquitin Protein That Mediates Intraspecies Antagonism.” Edited by John J. Mekalanos. MBio 8, no. 6 (November 28, 2017): e01902-17. https://doi.org/10.1128/mBio.01902-17.

    5. Choudhury, Nila Roy, Gregory Heikel, Maryia Trubitsyna, Peter Kubik, Jakub Stanislaw Nowak, Shaun Webb, Sander Granneman, et al. “RNA-Binding Activity of TRIM25 Is Mediated by Its PRY/SPRY Domain and Is Required for Ubiquitination.” BMC Biology 15, no. 1 (December 2017). https://doi.org/10.1186/s12915-017-0444-9.

    6. Dubrez, Laurence. “Regulation of E2F1 Transcription Factor by Ubiquitin Conjugation.” International Journal of Molecular Sciences 18, no. 12 (October 19, 2017): 2188. https://doi.org/10.3390/ijms18102188.

    7. Gadhave, Kundlik, Nityanand Bolshette, Ashutosh Ahire, Rohit Pardeshi, Krishan Thakur, Cristiana Trandafir, Alexandru Istrate, et al. “The Ubiquitin Proteasomal System: A Potential Target for the Management of Alzheimer’s Disease.” Journal of Cellular and Molecular Medicine 20, no. 7 (July 2016): 1392–1407. https://doi.org/10.1111/jcmm.12817.

    8. Gilberto, Samuel, and Matthias Peter. “Dynamic Ubiquitin Signaling in Cell Cycle Regulation.” The Journal of Cell Biology 216, no. 8 (August 7, 2017): 2259–71. https://doi.org/10.1083/jcb.201703170.

    9. Gupta, Nilaksh, Wei Li, and Thomas M. McIntyre. “Deubiquitinases Modulate Platelet Proteome Ubiquitination, Aggregation, and ThrombosisSignificance.” Arteriosclerosis, Thrombosis, and Vascular Biology 35, no. 12 (December 2015): 2657–66. https://doi.org/10.1161/ATVBAHA.115.306054.

    10. Hentze, Matthias W., Alfredo Castello, Thomas Schwarzl, and Thomas Preiss. “A Brave New World of RNA-Binding Proteins.” Nature Reviews Molecular Cell Biology, January 17, 2018. https://doi.org/10.1038/nrm.2017.130.

    11. Huang, Anqi, Christopher Amourda, Shaobo Zhang, Nicholas S Tolwinski, and Timothy E Saunders. “Decoding Temporal Interpretation of the Morphogen Bicoid in the Early Drosophila Embryo.” ELife 6 (July 10, 2017). https://doi.org/10.7554/eLife.26258.

    12. Krupina, Ksenia, Charlotte Kleiss, Thibaud Metzger, Sadek Fournane, Stephane Schmucker, Kay Hofmann, Benoit Fischer, et al. “Ubiquitin Receptor Protein UBASH3B Drives Aurora B Recruitment to Mitotic Microtubules.” Developmental Cell 36, no. 1 (January 2016): 63–78. https://doi.org/10.1016/j.devcel.2015.12.017.

    13. Lee, Sora, Jessica M Tumolo, Aaron C Ehlinger, Kristin K Jernigan, Susan J Qualls-Histed, Pi-Chiang Hsu, W Hayes McDonald, Walter J Chazin, and Jason A MacGurn. “Ubiquitin Turnover and Endocytic Trafficking in Yeast Are Regulated by Ser57 Phosphorylation of Ubiquitin.” ELife 6 (November 13, 2017). https://doi.org/10.7554/eLife.29176.

    14. Lin, Pei-Hui, Matthew Sermersheim, Haichang Li, Peter Lee, Steven Steinberg, and Jianjie Ma. “Zinc in Wound Healing Modulation.” Nutrients 10, no. 2 (December 24, 2017): 16. https://doi.org/10.3390/nu10010016.

    15. Martinez, Aitor, Benoit Lectez, Juanma Ramirez, Oliver Popp, James D. Sutherland, Sylvie Urbé, Gunnar Dittmar, Michael J. Clague, and Ugo Mayor. “Quantitative Proteomic Analysis of Parkin Substrates in Drosophila Neurons.” Molecular Neurodegeneration 12, no. 1 (December 2017). https://doi.org/10.1186/s13024-017-0170-3.

    16. Matsuo, Naoki, Natsuko Goda, Kana Shimizu, Satoshi Fukuchi, Motonori Ota, and Hidekazu Hiroaki. “Discovery of Cryoprotective Activity in Human Genome-Derived Intrinsically Disordered Proteins.” International Journal of Molecular Sciences 19, no. 2 (January 30, 2018): 401. https://doi.org/10.3390/ijms19020401.

    17. Mukherjee, Rukmini, Aneesha Das, Saikat Chakrabarti, and Oishee Chakrabarti. “Calcium Dependent Regulation of Protein Ubiquitination – Interplay between E3 Ligases and Calcium Binding Proteins.” Biochimica et Biophysica Acta (BBA) – Molecular Cell Research 1864, no. 7 (July 2017): 1227–35. https://doi.org/10.1016/j.bbamcr.2017.03.001.

    18. Penke, Botond, Ferenc Bogár, Tim Crul, Miklós Sántha, Melinda Tóth, and László Vígh. “Heat Shock Proteins and Autophagy Pathways in Neuroprotection: From Molecular Bases to Pharmacological Interventions.” International Journal of Molecular Sciences 19, no. 2 (January 22, 2018): 325. https://doi.org/10.3390/ijms19010325.

    19. Pinto, Maria J., Joana R. Pedro, Rui O. Costa, and Ramiro D. Almeida. “Visualizing K48 Ubiquitination during Presynaptic Formation By Ubiquitination-Induced Fluorescence Complementation (UiFC).” Frontiers in Molecular Neuroscience 9 (June 10, 2016). https://doi.org/10.3389/fnmol.2016.00043.

    20. Seissler, Tanja, Roland Marquet, and Jean-Christophe Paillart. “Hijacking of the Ubiquitin/Proteasome Pathway by the HIV Auxiliary Proteins.” Viruses 9, no. 12 (October 31, 2017): 322. https://doi.org/10.3390/v9110322.

    21. Sigalov, Alexander B. “Structural Biology of Intrinsically Disordered Proteins: Revisiting Unsolved Mysteries.” Biochimie 125 (June 2016): 112–18. https://doi.org/10.1016/j.biochi.2016.03.006.

    22. Sparrer, Konstantin M. J., and Michaela U. Gack. “TRIM Proteins: New Players in Virus-Induced Autophagy.” Edited by Rebecca Ellis Dutch. PLOS Pathogens 14, no. 2 (February 1, 2018): e1006787. https://doi.org/10.1371/journal.ppat.1006787.

    23. Tol, Sarah van, Adam Hage, Maria Giraldo, Preeti Bharaj, and Ricardo Rajsbaum. “The TRIMendous Role of TRIMs in Virus–Host Interactions.” Vaccines 5, no. 4 (August 22, 2017): 23. https://doi.org/10.3390/vaccines5030023.

    24. Uversky, Vladimir N. “The Multifaceted Roles of Intrinsic Disorder in Protein Complexes.” FEBS Letters 589, no. 19PartA (September 14, 2015): 2498–2506. https://doi.org/10.1016/j.febslet.2015.06.004.

    25. Yan, Kaowen, Li Li, Xiaojian Wang, Ruisha Hong, Ying Zhang, Hua Yang, Ming Lin, et al. “The Deubiquitinating Enzyme Complex BRISC Is Required for Proper Mitotic Spindle Assembly in Mammalian Cells.” The Journal of Cell Biology 210, no. 2 (July 20, 2015): 209–24. https://doi.org/10.1083/jcb.201503039.

    Please, correct any inaccuracies or omissions in the above list. Thanks.

  217. Dio,

    Here’s the DOI for #156
    DOI: 10.1186/1687-4153-2012-8 · Source: PubMed

    re: 157, yes,
    Comments 178-180 reference the Figures which I found
    fascinating of Durston, Wong, et al., in using their K-mode
    algorithm and how they represented it graphically.

  218. Digging a bit deeper into pre-ubiqutionation, E1 phase. Trying to understand signals that kick-off Ubiquitin tagging.

    Interesting paper from 2009, on Signaling Degradation,
    Degrons N-End Pathway, and Ubiquitin

    Was curious how, where, when Ubiquitin was called upon(“signaled”), to respond. Either for Damaged Proteins or normal Rates of Turnover for Proteins.

    So, from the beginning to the end, paper and quotes…

    Degradation Signal Diversity in the Ubiquitin-Proteasome System

    Open Access

    Nat Rev Mol Cell Biol. Author manuscript; available in PMC 2009 Mar 1.
    Published in final edited form as:
    Nat Rev Mol Cell Biol. 2008 Sep; 9(9): 679–690.
    doi: 10.1038/nrm2468

    Introduction

    Intracellular protein degradation has been studied for more than half a century, and it became clear early on that such degradation is highly selective, with individual protein half-lives ranging from minutes to years (for reviews of the early literature, see refs. 1-2). Moreover, much of this degradation was found to be energy-dependent despite the exergonic nature of peptide-bond cleavage. This energy dependence derives from the dual requirements of high substrate specificity and substrate protein unfolding to make the polypeptide backbone fully accessible for proteolytic cleavage.

    The vast majority of regulated protein degradation in eukaryotes is executed by the ubiquitin-proteasome system 3-5. Polyubiquitin tagging of substrates by specific enzymes provides the major source of selectivity in the system (Box 1), whereas the 26S proteasome complex performs the protein unfolding necessary for processive cleavage of the tagged proteins into short peptides (Box 2).
    In addition, ubiquitin ligation can function independently of the proteasome by directing certain -usually membrane- proteins to the lysosome/vacuole for proteolysis. Conversely, proteasomes can degrade some proteins without their prior modification by ubiquitin.

    “Signals” and Targeting

    As will be discussed in detail below, most short-lived proteins are distinguished by localized structure determinants (‘signals’) that target them to the ubiquitin ligase machinery or to the proteasome (or lysosome in some cases). A degradation signal or ‘degron’ 10, is usually defined as a minimal element within a protein that is sufficient for recognition and degradation by a proteolytic apparatus.

    An important property of degrons is that they are transferable. That is, genetically engineered attachment of such sequences confers metabolic instability (a short half-life) on otherwise long-lived proteins 3.

    Degrons can be defined for distinct proteolytic pathways, but we will confine this review to the description of degrons that target proteins to the ubiquitin-proteasome pathway.

    At this point, generalizations in this field are limited to some degree by the sheer diversity of substrates of the system.

    For example, many regulatory proteins are degraded in a temporally and spatially specific manner.

    These proteins are often tightly controlled by other post-translational modifications that are dependent on cell-signalling events.

    On the other hand, quality control of newly synthesized proteins and removal of misfolded proteins is under a very different set of constraints.

    The recognition and destruction of such aberrant proteins is expected to depend on their folding or assembly state. If folding or assembly goes awry, proteins would be expected to expose normally cryptic degrons that exist in many different proteins.

    “Design of different Degrons”

    In this review, we will focus on how different physiological requirements for the degradation of specific proteins dictate the design of different degrons. Distinct determinants comprise a degron, and they have different roles in the degradation pathway.

    Specifically, we consider primary recognition determinants as those sequences or structures within the degron that bind directly to the E3-E2 ubiquitin-ligase complex or its ancillary factors.

    Another determinant in ubiquitin-dependent degrons is the presence of an appropriate acceptor site(s) for attachment of the polyubiquitin chain, such as a lysine residue.

    The (polyubiquitin-modified) substrate must also be able to interact with the proteasome or shuttling factors that deliver it to the proteasome.

    Finally, the degron must be situated within the substrate such that the proteasome can initiate its unfolding and translocate it into the proteasome core. Our emphasis here will be on primary recognition determinants.

    Not all regulation of protein ubiquitylation occurs through substrate changes that activate a degron.

    Although we do not have space to cover this area, E3 and E2 enzymes can themselves be regulated by post-translational modification, such as the phosphorylation of the anaphase-promoting complex E3 11, or by binding to small molecules.

    For instance, dipeptide binding can allosterically modulate the N-recognin E3 (see next section) 12, and the growth-regulating plant indole auxin binds to a specific E3 ligase and forms part of an enlarged protein-binding interface that allows high-affinity interaction with specific protein substrates 13.

    N-degrons and the N-end rule pathway

    The notion that covalent ubiquitin conjugation commits proteins for degradation led the discoverers of ubiquitin-mediated proteolysis to propose that substrate selection takes place mainly at the stage of ubiquitin ligation 14, 15. By adding a variety of proteins to a rabbit reticulocyte lysate, Hershko and colleagues noted an apparent correlation between the presence of a free ?-amino group in the proteins and their ubiquitin-dependent degradation 16. They subsequently isolated a 180 kD protein with E3 ubiquitin ligase activity that appeared to have higher affinity for proteins with a free ?-amino group than those with a blocked N terminus 15.

    This particular E3 distinguishes proteins not only by a free N-terminal ?-amino group but also the side chain of the N-terminal residue.

    Varshavsky and co-workers systematically changed the N-terminal residue in an otherwise identical series of Escherichia coli ?-galactosidase test substrates and expressed them in yeast, where they displayed a remarkable range of degradation rates 17. Half-lives ranged from a few minutes to greater than 20 hours. Thus, an E3 is able to bind protein substrates with very high selectivity, in this case being able to distinguish substrates by recognizing a specific residue at the N terminus of a protein 18. This degradation pathway, termed the ‘N-end rule pathway,’ states that the half-life of a protein is determined by the nature of its N-terminal residue. Peptide sequences within the N-terminal region of the substrate that are sufficient for ubiquitin-dependent turnover constitute the ‘N-degron.’ (Figure 1A).

    Degrons in the ER

    All the degrons discussed so far can be classified as conditional signals for which post-translational modifications at specific sites are necessary to create a functional degron.

    The modified residues, along with neighbouring regions in the polypeptide, comprise the basic structural elements for recognition by a specific ubiquitin-ligase complex.

    However, not all substrate proteins are recognized through prior covalent modifications. Structural features that are revealed when a protein assumes a specific conformation or assembly state can serve as recognition elements in a wide range of degrons.

    Polypeptides that fail to assume their native tertiary or quaternary structures, collectively referred to as protein quality control (PQC) substrates, are often subject to this mode of substrate recognition (see refs. 59, 60).

    A major site for ubiquitin-dependent PQC is the endoplasmic reticulum (ER), where most secretory and integral membrane proteins are folded and assembled before being trafficked to their site of action.

    Proteins unable to fold or assemble properly usually never make it from the ER to the Golgi but instead are extracted back across the bilayer into the cytosol, ubiquitylated and degraded by the cytosolic proteasome (reviewed in refs. 61-64).

    Components of this ER-associated degradation (ERAD) system can also recognize native proteins undergoing transient or induced structural changes, allowing regulation of the levels of specific ER-resident proteins.

    Concluding remarks

    As is apparent from this limited survey, degrons are fundamental elements of protein degradation by the ubiquitin system.

    The foregoing discussion focused on elements within ubiquitin-dependent degrons that function in ubiquitin-ligase binding and ubiquitin-substrate conjugation.

    Once a polyubiquitin chain has been attached to a protein, the protein must still be properly routed to the proteasome, unfolded and then degraded.

    These steps depend on additional features of the degron or the proteolytic substrate.

    For example, several Lys48-linked polyubiquitin chain-modified cellular proteins have also been shown to bind the proteasome without concomitant degradation 137-139.

    .

    Several studies have indicated that in addition to tethering the substrate to the proteasome, the degron must also have a more loosely structured peptide segment that initiates unfolding and insertion into the proteasome 140-142.

    .

    The true range and variety of protein quality-control degrons are poorly defined, including the question of whether soluble and membrane PQC(Protein Quality Control) substrates use related or distinct degrons.

    ( ) emphasis mine

    In summary, what came first, the Signal, the Recognizer, the Tagger, the Receiver, the Cleaver, the Recycler?

    And since when does a blind system know what to recycle? And what NOT to recycle?

    .

  219. Degron Wiki Reference…


    A Degron is…

    … a portion of a protein that is important in regulation of protein degradation rates. Known Degrons include short amino acid sequences,[1] structural motifs[2] and exposed amino acids (often Lysine[3] or Arginine[4]) located anywhere in the protein. In fact, some proteins can even contain multiple degrons.[2][5] Degrons are present in a variety of organisms, from the N-degrons (see N-end Rule) first characterized in yeast[6] to the PEST sequence of mouse ornithine decarboxylase.[7] Degrons have been identified in prokaryotes[8] as well as eukaryotes.

    While there are many types of different degrons, and a high degree of variability even within these groups, Degrons are all similar for their involvement in regulating the rate of a protein’s degradation.[9][10][11] Much like protein degradation (see proteolysis) mechanisms are categorized by their dependence or lack thereof on Ubiquitin, a small protein involved in proteasomal protein degradation,[12][13][14]

    Degrons may also be referred to as “Ubiquitin-dependent”[9] or “Ubiquitin-independent”.[10][11]

    Two-step, Three-step Identification

    In order to identify a portion of a protein as a degron, there are often three steps performed.[2][19][20]

    First, the degron candidate is fused to a stable protein, such as GFP, and protein abundances over time are compared between the unaltered protein and the fusion (as shown in green).[21] If the candidate is in fact a degron, then the abundance of the fusion protein will decrease much faster than that of the unaltered protein.[9][10][11]

    Second, a mutant form of the degron’s protein is designed such that it lacks the degron candidate. Similar to before, the abundance of the mutant protein over time is compared to that of the unaltered protein (as shown in red). If the deleted degron candidate is in fact a degron, then the mutant protein abundance will decrease much slower than that of the unaltered protein.[9][10][11] Recall that degrons are often referred to as “Ubiquitin-dependent” or “Ubiquitin-independent”

    The third step performed is often done after one or both of the previous two steps, because it serves to identify the Ubiquitin dependence or lack thereof of a previously identified degron. In this step, protein A and A’ (identical in every way except the presence of degron in A’) will be examined. Note that mutation or fusion procedures could be performed here, so either A is a protein like GFP and A’ is a fusion of GFP with the degron (as shown in green) or A’ is the degron’s protein and A is a mutant form without the degron (as shown in Red.) The amount of Ubiquitin bound to A and to A’ will be measured.[2][7][20] A significant increase in the amount of Ubiquitin in A’ as compared to A will suggest that the degron is Ubiquitin-dependent.[2][9]

    See referenced diagram at link under Identification heading.

  220. Note on Degrons.

    Gpuccio posted papers in Original Post, noting use of Degrons, one behind a paywall.

    And another at Comment #10 interestingly enough titled…


    “Design Principles
    Involving Protein Disorder Facilitate Specific Substrate Selection and Degradation by the Ubiquitin-Proteasome System”

    🙂

    Part of the text Gpuccio highlighted…

    Next we review well studied substrates and discuss that substrate elements (degrons) recognized by E3 ligases are highly disordered: short linear motifs recognized by many E3s constitute an important class of degrons, and these are almost always present in disordered regions.

    Disordered as we remember being Flexible, Conditional and Context dependent.

    .

  221. Upright Biped, Dionisio, Gpuccio,

    Martello Barbieri’s inspired Biosemetics Journal has this
    in it’s first paragraph…

    “Biosemiotics is dedicated to building a bridge between biology, philosophy, linguistics and the communication sciences. If it is true that biosemiotics is “the study of signs, of communication and of information in living organisms” (Oxford Dictionary of Biochemistry and Molecular Biology, 1997, p. 72), it is also true that, in time, it has acquired a more general scope. Today, its main challenge is the attempt to naturalize biological meaning, in the belief that signs are fundamental, constitutive components of the living world. Biosemiotics has triggered revision of fundamentals of both biology and semiotics: biology needs to recognize the semiotic nature of life and reformulate its theories accordingly, and semiotics has to accept the existence of signs in animals, plants, and even individual cells. Biosemiotics has become in this way the leading edge of the research on the fundamentals of life, and is a young exciting field on the move.”

    “… attempt to naturalize biological meaning”

    is an interesting way to put it and a “challenge” indeed.

  222. DATCG,

    Apparently not all the complex biological processes dealing with functionally specified information can be easily described through biosemiotics principles. For example, as we discussed earlier in this thread [ @54 & @69 ], spatiotemporal signal concentration profiles (a.k.a. morphogen gradients) are not easily associated with biosemiotics or codes, because -unlike translation- the effectors seem embedded in the process without discontinuity.

  223. DATCG @219:

    Thanks for the information.

  224. DATCG,

    Regarding the comment @224, see also @74-77.

    That particular issue was briefly discussed @54, @69, and @74 through 77.

  225. DATCG at #220:

    Very interesting paper about degrons.

    Indeed, this is of course the set of “signals” that the E3-E2 complex must be able to recognize. That adds to the semiosis of the system.

    So, the E3 ligase(complexed to its E2) must be able to:

    a) Recognize its specific protein substrate.

    b) Recognize some specific degron signal in that substarte (at least for protein degradation which, as we know, is not the only target od ubiquitination).

    c) Be able to apply the correct ubiquitination signal to that specific target with that specific degron signal.

    This extreme semiotic complexity is very interesting, and of course must have very good motivations. In theory, we could think of some much simpler system, which can recognize a few degron signal on any protein, and aplly some standard degradation signal to them all.

    But that’s not the case. In the ubiquitin system, each combinatorial formula of degron signal and ubiquitin signal is applied in a specific way to a specific substrate.

    That’s why we have all those E2, and especially E3, complex molecule.

    Substrate recognition is the key to all, and it strictly conditions degron signal recognition and ubiquitin signal application.

    Again, the engineering resources linked to tis system are amazing, and of course the real purpose of all this must be to make an extremely fine-tuned regulation of cellular processes possible.

  226. DATCG, Upright Biped, Dionisio:

    It’s also interesting to remark that protein degradation (which, again, is not the only task of ubiquitination) has at least wto completely different purposes:

    a) Degrading proteins which are not really functional, or are not really functional any more (so calle Protein Quality control). If I understand well, this is mainly accomplished by lysosome autophagy, especially for damaged big structures, like mitochondria.

    b) Controlling the concentration of key regulatory or effector proteins, whose levels must be stable but readily adjustable according to conditions. This is of course a much finer task, accomplished by intervening on highly dynamic physical systems.

    The two things are completely different in concept and, of course, implementation, and it’s really surprising that the same ubiquitin system presides to both! 🙂

  227. DATCG,

    Please help me with this:

    @219 you wrote:

    Dio,
    Here’s the DOI for #156
    DOI: 10.1186/1687-4153-2012-8 · Source: PubMed

    Referring to your comment @156, where you wrote:

    FYI, has anyone seen Abel’s paper from 2015?
    off-topic: PDF Document…
    Functional Sequence Complexity (FSC) Measured in Fits(Functional bits)

    The link you provided @156 points to this:
    https://www.researchgate.net/publication/275892308_Functional_Sequence_Complexity_FSC_measured_in_fits

    Which includes the full text PDF of the given paper.

    However, the DOI you just provided @219 points to this:

    https://bsb-eurasipjournals.springeropen.com/articles/10.1186/1687-4153-2012-8

    Am I missing something?

    Thanks

  228. “Evolution of the Ubiquitin system? … ”

    The problem of evolution of anything always boils down to the distinction between specificity and non-specificity. That is why the problem of evolution is generic and we don’t need to name or identify any particular biological component, system or function, but only two things:

    A) the number of particles comprising biological components
    B) the level of specificity that retains a functional role of a component in a system.

    Here is detailed explanation:

    https://biospecificity.wordpress.com/

  229. gpuccio @228:

    b) Controlling the concentration of key regulatory or effector proteins, whose levels must be stable but readily adjustable according to conditions. This is of course a much finer task, accomplished by intervening on highly dynamic physical systems.

    Does the above item ‘b’ relate to the issues addressed @226?

  230. forexhr @230:

    That’s an interesting contribution to the discussion. Thanks.

  231. #229 Dio,

    my mistake, looked again, there is no DOI for that paper by Abel. Might be it’s only published on his Emergence Project site.

  232. #228 Gpuccio,

    Precisely, “extremely fine-tuned” in the 2nd mode for Ubiquitin. It’s quite precarious process and explains why so many diseases crop up if not in sync.

    On item a) degradation, I read a little bit about one scenario farther up processing ladder if aggregation forms. In eyes causing cataracts if not corrected.

    But then there’s this news reported in 2015 on communications issues with Calpain from Tufts.

    Mechanism involved in causing cataracts in mice identified by researchers

    from Sciencedaily; Science News 2015

    Summary

    A communications breakdown between two biochemical pathways is involved in causing cataracts in mice, scientists have discovered. The newfound relationship between the ubiquitin and calpain pathways may lead to pharmaceuticals and dietary approaches that can prolong the function of the relevant pathways and delay the onset of cataracts in people.

    Cataracts is one of the Most Common Eye Diseases

    Cataracts are caused in part by the accumulation of abnormal proteins. Normally, obsolete or damaged proteins are removed by the ubiquitin and lysosomal pathways. The authors noticed when the ubiquitin pathway falters calcium flows into the cells of the lens, causing a third pathway to activate. It is this third pathway that causes cataract-related damage in the eye.

    “We discovered that the ubiquitin pathway and the calpain pathway communicate with one another. When their conversation goes awry, cells start a vicious cycle in which proteins are improperly degraded,” said senior and corresponding author Allen Taylor, Ph.D., the director of Laboratory for Nutrition and Vision at the USDA HNRCA and a professor at the Friedman School of Nutrition Science and Policy at Tufts University. “This leads to alterations in proteins and the beginning of the clouding of the lens that signals the onset of cataract.”

  233. Gpuccio,

    re: your points…

    “That’s why we have all those E2, and especially E3, complex molecule.”

    It escalates for E3 as a need for Conditional Processing too? As well as specificity and context of the proteins.

    Is that overstating it?

    “Substrate recognition is the key to all, and it strictly conditions degron signal recognition and ubiquitin signal application.”

    And if there are any changes, disease. My question would be what are threshold levels for limiting mutations to these working areas. And if this is well understood yet. We know abnormal proteins accumulate if a, b, c or d happen. But what are Fault-Tolerance levels.

    “Again, the engineering resources linked to tis system are amazing, and of course the real purpose of all this must be to make an extremely fine-tuned regulation of cellular processes possible.”

    Fine-tuned with error correction from step 1 to 1001 😉

    Protein Quality Control Systems appear by blind process?

    It is an amazing labyrinth of pathways, locks, keys, signals, cleavings, stackings and communications networks zooming around in the cells.

  234. forexhr:

    I think we probably agree on the main ideas.

    Of course, I have some definite approaches to measuring functional information (specificity), and I rely a lot on sequence conservation throughout long evolutionary times.

  235. DATCG at #234:

    Calpain seems another additional calcium-regulated network for protein degradation. How many of them are there? 🙂

    From Uniprot:

    “Calcium-regulated non-lysosomal thiol-protease which catalyzes limited proteolysis of substrates involved in cytoskeletal remodeling and signal transduction. Proteolytically cleaves MYOC at ‘Arg-226’ (PubMed:17650508). Proteolytically cleaves CPEB3 following neuronal stimulation which abolishes CPEB3 translational repressor activity, leading to translation of CPEB3 target mRNAs (By similarity).”

  236. @237 Gpuccio,

    How many and Why? 🙂

    If it’s not due to random mutations and natural selection. If RM&NS there is no rhyme or reason.

    But there must be practical reasons if by Design.

    Calpain assist ubiquitin. Without it ubiquitination will not occur.

    Non-lysosomal, hmmmm. We know the Proteosome cannot handle all proteins for degradation.

    Tissue Specificity maybe or limits and controls, but there are others as you pointed out as well.

    So we have multiple interfaces for specific task in and outside of the Ubiquitin framework that also coordinate with it for degradation and other task.

    Each in it’s own realm of duties, rules and constraints.

  237. DATCG @233,

    OK, no problem.
    Thanks.

  238. #237 Gpuccio, Dionisio,

    Another regulated network in coordination with mono and poly-Ubiquitin System. p97 or VCP.

    Don’t think it was covered yet. Like you said Gpuccio, there must be many more. But I found this interesting as they’re using it to inhibit the Proteasome, RP11 and slow down cancerous tumors.

    After watching Dr. Deshaies Video 2 & 3 again, I became curious of his commercial work.

    Turns out he’s a Scientific Founder at Cleave Biosciences
    working with different aspects of ubiquitin pathways and coordinated systems like VCP-p97.

    They have research ongoing into P97 – a celluar AAA ATPase, with a “described role as master regulator in the (UPS) Ubiquitin Proteasome System.”

    Here…
    Cleave BioSciences Pipeline – P97

    They have a very short Summary…

    P97 is an important cellular AAA ATPase also known as VCP (Valosin Containing Protein) that has a well described role as a master regulator in the Ubiquitin Proteasome System. P97/VCP extracts misfolded proteins from membranes, such as the endoplasmic reticulum, and chaperones them to the proteasome for degradation.1 Emerging evidence suggests that cancer cells become over-dependent on protein homeostasis systems and therefore inhibitors of p97 are expected to have meaningful anti-cancer activity.2 Genetic knockdown of p97 leads to cell death in a number of solid tumor cell lines and compounds that inhibit p97 have been discovered which lead to cancer cell death in various pre-clinical models.3, 4

    .

  239. Continuing below…

  240. Delete #241 please.

    Continued…

    Searched for more on VCP-p97 as regulator in Ubiqutin process.

    Found this paper…
    The VCP/p97 system at a glance: connecting cellular function to disease pathogenesis

    2014. Published by The Company of Biologists Ltd | Journal of Cell Science (2014) 127, 1–7 doi:10.1242/jcs.093831
    Hemmo Meyer1, and Conrad C. Weihl2

    Most research connects VCP/p97 to ubiquitin-dependent processes (see Box 1), as it directly and indirectly binds to ubiquitylated substrates and facilitates steps downstream of ubiquitylation (Ye, 2006; Jentsch and Rumpf, 2007; Meyer et al., 2012).

    A common theme is that VCP/p97 extracts ubiquitylated proteins from membranes or cellular structures, or segregates them from binding proteins. Importantly, the degree of the requirement for VCP/p97 varies and might depend on substrate localization, structure or solubility (Beskow et al., 2009; Gallagher et al., 2014).

    In many cases, VCP/p97 facilitates the degradation of polyubiquitylated substrates in the proteasome. However, VCP/p97 also targets proteins with monoubiquitin or non-degradative ubiquitin chains, and recent examples of the role of VCP/p97 in segregating transcription factors from chromatin and disassembling RNA-protein complexes (see below) cement this notion (Stolz et al., 2011; Meyer et al., 2012; Ndoja et al., 2014).

    p97 functions also as interaction hub

    The second aspect of VCP/p97 function is its role as an interaction hub, and different sets of at least 30 cofactors have been shown to be responsible for modulating VCP/p97-mediated processes (Schuberth and Buchberger, 2008; Yeung et al., 2008; Meyer et al., 2012). These cofactors contain specific interaction domains or motifs that bind to VCP/p97 either at its N-terminal domain or C-terminal tail (see poster).
    Some of these cofactors serve as ubiquitin adaptors or recruit VCP/p97 to intracellular membranes.

    Deubiquitylating Enzymes

    In addition, VCP/p97 directly or indirectly binds to ubiquitin ligases and deubiquitylating enzymes (DUBs), including a large number of cullin-RING ligases (Alexandru et al., 2008; Sowa et al., 2009).

    VCP/p97-associated DUBs and ligases edit the ubiquitin chains on the substrate protein to either improve its targeting to the proteasome or recycle the substrate, thus determining its fate (Jentsch and Rumpf, 2007; Meyer et al., 2012).

    p92 VCP ensures homeostasis in facilitating
    proteasomal degradation of damaged or misfolded proteins

    Its main role in ensuring protein homeostasis is well established, as it facilitates the proteasomal degradation of large cohorts of damaged or misfolded proteins in different compartments including:

    – ER (termed ER-associated degradation or ERAD),
    – outer mitochondrial membrane and the nucleus,
    – co-translational degradation at the ribosome (see below).

    Governs crucial signaling pathways

    Besides its quality control function, VCP/p97 also governs crucial signaling pathways (Meyer et al., 2012; Yamanaka et al., 2012). Important examples are the degradation of IkBa (also known as NFKBIA), which leads to NFkB activation (Li et al., 2014), or degradation of HIF1a, which downregulates the hypoxic response (Alexandru et al., 2008).

    Not sure if this involves ubiquitin or not. But it does NFkB, so most likely.

    VCP-p97 non-degradative process of TFs

    In other cases, the function of VCP/p97 is non-degradative; for instance, the extraction of transcription factor precursors from membranes for their subsequent activation, such as has been reported for Spt23, which regulates the expression of the fatty acid desaturase Ole1 in yeast, or Nrf1(TF), which is involved in the homeostatic response (Jentsch and Rumpf, 2007; Radhakrishnan
    et al., 2014).

    Chromatin

    An emerging aspect is the role of VCP/p97 in cell cycle
    progression and chromatin-associated functions that ensure
    genomic stability (see poster).

    VCP/p97 dissociates proteins from chromatin, either for their degradation or for recycling, to modulate the dynamics of chromatin regulators, and the list of substrates that are regulated in this way is growing (Meyer et al., 2012; Vaz et al., 2013).

    DNA double-stranded breaks

    In response to double-strand breaks, VCP/p97 removes L3MBTL1 and unidentified K48-linked ubiquitin conjugates from damaged sites to orchestrate DNA repair and facilitates CDC25A degradation to enforce the G2/M checkpoint (Acs et al., 2011; Meerang et al., 2011; Riemer et al., 2014).

    Whereas these are all functions that are mediated by its heterodimeric adaptor Ufd1–Npl4, VCP/p97 also cooperates with DVC1 (also known as SPRTN) to extract ubiquitylated DNA polymerase g and rescue stalled replication forks (Davis et al., 2012; Mosbech et al., 2012) in human cells.

  241. continued…

    Figure 1 for above comment Includes VCP-p97 diagram functions and interactions with ubiquitin system.

    Unfortunately not an SVG format. Does not scale well.

    VCP-p97 – Ubiqiutin Systems and other Functions

  242. DATCG:

    You have definitely found a very important actor in the scene we have been debating! 🙂

    VCP-p97 seems to be as elusive as its many names (TERA, Transitional endoplasmic reticulum ATPase, VCP, Valosin-containing protein, CDC48, and so on).

    The same name of its protein family is astounding:

    AAA+: extended family of ATPases associated with various cellular activities.

    And various it is!

    From Uniprot:

    Necessary for the fragmentation of Golgi stacks during mitosis and for their reassembly after mitosis. Involved in the formation of the transitional endoplasmic reticulum (tER). The transfer of membranes from the endoplasmic reticulum to the Golgi apparatus occurs via 50-70 nm transition vesicles which derive from part-rough, part-smooth transitional elements of the endoplasmic reticulum (tER). Vesicle budding from the tER is an ATP-dependent process. The ternary complex containing UFD1, VCP and NPLOC4 binds ubiquitinated proteins and is necessary for the export of misfolded proteins from the ER to the cytoplasm, where they are degraded by the proteasome. The NPLOC4-UFD1-VCP complex regulates spindle disassembly at the end of mitosis and is necessary for the formation of a closed nuclear envelope. Regulates E3 ubiquitin-protein ligase activity of RNF19A. Component of the VCP/p97-AMFR/gp78 complex that participates in the final step of the sterol-mediated ubiquitination and endoplasmic reticulum-associated degradation (ERAD) of HMGCR. Involved in endoplasmic reticulum stress-induced pre-emptive quality control, a mechanism that selectively attenuates the translocation of newly synthesized proteins into the endoplasmic reticulum and reroutes them to the cytosol for proteasomal degradation (PubMed:26565908). Also involved in DNA damage response: recruited to double-strand breaks (DSBs) sites in a RNF8- and RNF168-dependent manner and promotes the recruitment of TP53BP1 at DNA damage sites (PubMed:22020440, PubMed:22120668). Recruited to stalled replication forks by SPRTN: may act by mediating extraction of DNA polymerase eta (POLH) to prevent excessive translesion DNA synthesis and limit the incidence of mutations induced by DNA damage (PubMed:23042607, PubMed:23042605). Required for cytoplasmic retrotranslocation of stressed/damaged mitochondrial outer-membrane proteins and their subsequent proteasomal degradation (PubMed:16186510, PubMed:21118995). Essential for the maturation of ubiquitin-containing autophagosomes and the clearance of ubiquitinated protein by autophagy (PubMed:20104022, PubMed:27753622). Acts as a negative regulator of type I interferon production by interacting with DDX58/RIG-I: interaction takes place when DDX58/RIG-I is ubiquitinated via ‘Lys-63’-linked ubiquitin on its CARD domains, leading to recruit RNF125 and promote ubiquitination and degradation of DDX58/RIG-I (PubMed:26471729). May play a role in the ubiquitin-dependent sorting of membrane proteins to lysosomes where they undergo degradation (PubMed:21822278). May more particularly play a role in caveolins sorting in cells (PubMed:21822278, PubMed:23335559).

    By the way, it is an 806 AAs long protein (in humans), extremely conserved in all eukaryotes (almost as much as ubiquitin).

    78% identities and 89% positives in fungi, 1313 bits, 1.63 baa.

    Complex structure, complex interactions, and definitely a prima donna in the ubiquitin drama.

    The amazing thing is that, as usual, so many things are known about it (I am just starting to dig), and yet so little is really understood.

    Maybe just a look at Wikipedia for a brief summary:

    Function[edit]
    p97/CDC48 performs diverse functions through modulating the stability and thus the activity of its substrates. The general function of p97/CDC48 is to segregate proteins from large protein assembly or immobile cellular structures such as membranes or chromatin, allowing the released protein molecules to be degraded by the proteasome. The functions of p97/CDC48 can be grouped into the following three major categories.

    Protein quality control[edit]
    The best characterized function of p97 is to mediate a network of protein quality control processes in order to maintain protein homeostasis.[49] These include endoplasmic reticulum-associated protein degradation (ERAD) and mitochondria-associated degradation.[14][50] In these processes, ATP hydrolysis by p97/CDC48 is required to extract aberrant proteins from the membranes of the ER or mitochondria. p97/CDC48 is also required to release defective translation products stalled on ribosome in a process termed ribosome-associated degradation.[51][52][53] It appears that only after extraction from the membranes or large protein assembly like ribosome, can polypeptides be degraded by the proteasome. In addition to this ‘segregase’ function, p97/CDC48 might have an additional role in shuttling the released polypeptides to the proteasome. This chaperoning function seems to be particularly important for degradation of certain aggregation-prone misfolded proteins in nucleus.[54] Several lines of evidence also implicate p97 in autophagy, a process that turns over cellular proteins (including misfolded ones) by engulfing them into double-membrane-surrounded vesicles named autophagosome, but the precise role of p97 in this process is unclear.[55]

    Chromatin-associated functions[edit]
    p97 also functions broadly in eukaryotic nucleus by releasing protein molecules from chromatins in a manner analogous to that in ERAD.[56] The identified p97 substrates include transcriptional repressor ?2 and RNA polymerase (Pol) II complex and CMG DNA helicase in budding yeast, and the DNA replicating licensing factor CDT1, DNA repairing proteins DDB2 and XPC, mitosis regulator Aurora B, and certain DNA polymerases in mammalian cells. These substrates link p97 function to gene transcription, DNA replication and repair, and cell cycle progression.

    Membrane fusion and trafficking[edit]
    Biochemical and genetic studies have also implicated p97 in fusion of vesicles that lead to the formation of Golgi apparatus at the end of mitosis.[57] This process requires the ubiquitin binding adaptor p47 and a p97-associated deubiquitinase VCIP135, and thus connecting membrane fusion to the ubiquitin pathways. However, the precise role of p97 in Golgi formation is unclear due to lack of information on relevant substrate(s). Recent studies also suggest that p97 may regulate vesicle trafficking from plasma membrane to the lysosome, a process termed endocytosis.[55]

    Now, I would say that what is called here the “general function”:

    “The general function of p97/CDC48 is to segregate proteins from large protein assembly or immobile cellular structures such as membranes or chromatin, allowing the released protein molecules to be degraded by the proteasome.”

    is as intriguing and undetailed as it can be!

    However, the “structure” part (always in Wikipedia) is even better:

    Structure[edit]
    According to the crystal structures of full-length wild-type p97,[17][18] six p97 subunits assemble into a barrel-like structure, in which the N-D1 and D2 domains form two concentric, stacked rings (Figure 2).

    The N-D1 ring is larger (162 Å in diameter) than the D2 ring (113 Å) due to the laterally attached N-domains. The D1 and D2 domains are highly homologous in both sequence and structure, but they serve distinct functions. For example, the hexameric assembly of p97 only requires the D1 but not the D2 domain.[19] Unlike many bacterial AAA+ proteins, assembly of p97 hexamer does not depend on the presence of nucleotide. The p97 hexameric assembly can undergo dramatic conformational changes during nucleotide hydrolysis cycle,[20][21][22][23][24] and it is generally believed that these conformational changes generate mechanical force, which is applied to substrate molecules to influence their stability and function. However, how precisely p97 generates force is unclear.

    Emphasis mine.

  243. Gpuccio,

    Thanks for your detailed response, specifically on conservation in eukaryotes and humans, including the Blast stats!

    On the conformational changes, it’s quite amazing what it goes through and this is yet again conditional.

    By the way, it is an 806 AAs long protein (in humans), extremely conserved in all eukaryotes (almost as much as ubiquitin).

    78% identities and 89% positives in fungi, 1313 bits, 1.63 baa.

    Conserved together as they would have to be from the beginning for any of this to make sense, correct?

    There is no(or very little) room for mutations here. Disease is the result if mutations impact these different systems working together on crucial time-dependent delivery.

    Thus all the Quality Control systems and constraints in place to clear out mutations and damaged goods.

  244. Hey guys, Dionisio, Gpuccio, UB, etc.,

    What more is there to add? Anything we’re missing or have not covered? Or to highlight?

    I’m tempted to post more papers but did not want to do so Gpuccio if you think the subject matter for this post has been fully expanded upon.

    I think for me, there’s the overall picture, big image of the systems control aspects, semiosis, then Conserved Functions over time in eukaryotes you’ve highlighted.

    These systems are so large, complex, integrated it’s hard to set back and look upon them as well understood units in a larger frame of reference.

    Even with all the infographics, step by step processes, and videos, still hard to comprehend it all in formalized actions and conditions.

    There’s one issue I do not understand in video 3 by Dr. Deshaies presentation on inhibiting cancerous tumors by blocking the Proteasome.

    Looking at his commercial site, including some prescriptions approved we can see even targeted solutions still result in possible serious repercussions to people as side affects.

    It seems the methods utilized today, though much better are a bit like using a hammer on a screw.

    I thought from a Design perspective, it would be more upstream in detection systems or cutting off supply to the tumor by a better method. Or, farther upstream, detecting mutations that allow tumors to form in the first place and replacing those mutations – maybe – not saying it’s easy. Just thinking through the process. That would mean fully understanding the detailed circumstances that allowed the mutation upstream.

    If it could be done, eliminating need for post-treatment of tumors after they’ve started. Which is late in the process. Though I would not rule out better Post-treatment methodology.

    Just some thoughts.

  245. @217 updated list (added 3 references posted after #217):

    1. Abel, David L., and Jack T. Trevors. “Three Subsets of Sequence Complexity and Their Relevance to Biopolymeric Information.” Theoretical Biology and Medical Modelling 2, no. 1 (August 11, 2005): 29. https://doi.org/10.1186/1742-4682-2-29.

    2. Durston, Kirk K, David KY Chiu, Andrew KC Wong, and Gary CL Li. “Statistical Discovery of Site Inter-Dependencies in Sub-Molecular Hierarchical Protein Structuring.” EURASIP Journal on Bioinformatics and Systems Biology 2012, no. 1 (December 2012). https://doi.org/10.1186/1687-4153-2012-8.

    3. Liu, Ke, Lei Lyu, David Chin, Junyuan Gao, Xiurong Sun, Fu Shang, Andrea Caceres, et al. “Altered Ubiquitin Causes Perturbed Calcium Homeostasis, Hyperactivation of Calpain, Dysregulated Differentiation, and Cataract.” Proceedings of the National Academy of Sciences 112, no. 4 (January 27, 2015): 1071–76. https://doi.org/10.1073/pnas.1404059112.

    4. Meyer, H., and C. C. Weihl. “The VCP/P97 System at a Glance: Connecting Cellular Function to Disease Pathogenesis.” Journal of Cell Science 127, no. 18 (September 15, 2014): 3877–83. https://doi.org/10.1242/jcs.093831.

    5. Pla, A, M Pascual, J Renau-Piqueras, and C Guerri. “TLR4 Mediates the Impairment of Ubiquitin-Proteasome and Autophagy-Lysosome Pathways Induced by Ethanol Treatment in Brain.” Cell Death & Disease 5, no. 2 (February 2014): e1066–e1066. https://doi.org/10.1038/cddis.2014.46.

    6. Ravid, Tommer, and Mark Hochstrasser. “Diversity of Degradation Signals in the Ubiquitin–proteasome System.” Nature Reviews Molecular Cell Biology 9, no. 9 (September 2008): 679–89. https://doi.org/10.1038/nrm2468.

    7. Rogers, J. M., V. Oleinikovas, S. L. Shammas, C. T. Wong, D. De Sancho, C. M. Baker, and J. Clarke. “Interplay between Partner and Ligand Facilitates the Folding and Binding of an Intrinsically Disordered Protein.” Proceedings of the National Academy of Sciences 111, no. 43 (October 28, 2014): 15420–25. https://doi.org/10.1073/pnas.1409122111.

    8. Ruiz i Altaba, Ariel, Vân Nguyên, and Verónica Palma. “The Emergent Design of the Neural Tube: Prepattern, SHH Morphogen and GLI Code.” Current Opinion in Genetics & Development 13, no. 5 (October 2003): 513–21. https://doi.org/10.1016/j.gde.2003.08.005.

    9. Savage, Kienan I., and D. Paul Harkin. “BRCA1, a ‘Complex’ Protein Involved in the Maintenance of Genomic Stability.” The FEBS Journal 282, no. 4 (February 2015): 630–46. https://doi.org/10.1111/febs.13150.

    10. Srikanthan, S., W. Li, R. L. Silverstein, and T. M. McIntyre. “Exosome Poly-Ubiquitin Inhibits Platelet Activation, Downregulates CD36 and Inhibits pro-Atherothombotic Cellular Functions.” Journal of Thrombosis and Haemostasis 12, no. 11 (November 2014): 1906–17. https://doi.org/10.1111/jth.12712.

    11. Uversky, Vladimir N. “Functional Roles of Transiently and Intrinsically Disordered Regions within Proteins.” FEBS Journal 282, no. 7 (April 2015): 1182–89. https://doi.org/10.1111/febs.13202.

    12. Wang, Yi-Ting, and Guang-Chao Chen. “The Role of Ubiquitin System in Autophagy.” In Autophagy in Current Trends in Cellular Physiology and Pathology, edited by Nikolai V. Gorbunov and Marion Schneider. InTech, 2016. https://doi.org/10.5772/64728.

  246. DATCG:

    Well, I think we have certainly covered a lot of important and interesting issues. I certainly agree with your thoughts.

    Please, feel free to post something new if you think it is worthwhile, or not to post if you prefer so. I will do more or less the same.

    I think this whole subject is really good evidence for design, for a lot of different and intertwining reasons that we have tried to highlight in our “private party” here.

    But our kind interlocutors probably think that all this is trivial or irrelevant, otherwise they would certainly have joined the discussion to show us out serious errors! 🙂

  247. DATCG:

    The problem of a functional, or “pathogenetic” therapy of tumours is complex, and in general rather frustrating.

    For a lot of time the therapy of tumors and leukemias has been highly empirical, and based essentially on drugs which are toxic to all cells.

    Understanding the biological features of neoplastic cells has always been a great aim, but unfortunately our increase in understanding has not always provided really useful therapeutic strategies.

    However, things are probably changing, and maybe with time we can get better results.

    About going upstream: I don’t know, “detecting mutations that allow tumors to form in the first place and replacing those mutations” seems still rather far away. Of course detecting tumors when they are still at the beginning would be great, but the problem is that they are really a lot of different things, from a biological point of view, even when they have similar clinical manifestations. And a lot of random events are probably implied in the initial phases of the disease. Here again complexity makes it difficult for us to really understand, and unfortunately it is here the complexity of possible random devastations of extremely complex functions.

    But understanding is always the foundation for all. After understanding, some power of intervention must come, sooner or later.

  248. Dionisio:

    Thank you for your updates! 🙂

  249. DATCG:

    Again about VCP/p97/CDC48 (February 2017):

    A Cdc48 “Retrochaperone” Function Is Required for the Solubility of Retrotranslocated, Integral Membrane Endoplasmic Reticulum-associated Degradation (ERAD-M) Substrates

    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5336148/

    Retrochaperone?

    Endoplasmic reticulum (ER)2-associated degradation (ERAD) refers to a group of quality control pathways that degrade damaged or misfolded ER-localized proteins (1, 2). ERAD occurs through the ubiquitin-proteasome pathway, by which ubiquitin is attached to ERAD substrates to cause proteasomal degradation (3,–5).

    ERAD pathways present the cell with a spatial challenge. The 26S proteasome resides in the cytosol and connected compartments, along with the E1 ubiquitin-activating enzyme and most E2s. Accordingly, a unifying feature of all ERAD pathways is the requirement for movement of substrates from the ER membrane or lumen to the cytosol for degradation. This transport component of ERAD is broadly referred to as dislocation or retrotranslocation, and it has been known to occur since the earliest studies of ERAD

    In this work we sought to discover the retrochaperones that allow multispanning membrane proteins to remain soluble after retrotranslocation during ERAD.

    We then used unbiased proteomics to identify and confirm Cdc48 as the principal retrochaperone allowing multispanning membrane proteins to remain soluble in the cytosol.

    We explored the features of Cdc48-client interaction. The tripartite Cdc48 complex binds polyubiquitin chains. Proteolytic removal of the polyubiquitin from Hmg2-GFP abolished its Cdc48 association and rendered the retrotranslocated Hmg2-GFP insoluble. Similarly, addition of excess polyubiquitin chains to the assay supernatant resulted in complete loss of Cdc48 binding to retrotranslocated Hmg2-GFP, and again it caused drastic loss of Hmg2-GFP solubility. Thus, it was clear that polyubiquitin-mediated association of the Cdc48 complex is critical for the maintained solubility of the eight-spanning Hmg2-GFP, indicating that Cdc48 is a bone fide retrochaperone in addition to being a ubiquitin-dependent “dislocase.

    The polyubiquitin binding of Cdc48 was a critical component of Cdc48 retrochaperone function.

    Cdc48/p97 are AAA hexameric ATPases. They are thought to use ATP hydrolysis to generate the considerable conformational force needed for the many versions of ubiquitin-dependent extraction and dislocation for which they are well known (19). Especially in the case of ERAD-M retrotranslocation, prodigious energy would be required for substrate removal from the membrane. However, it is less clear if the retrochaperoning role of Cdc48 is also ATP-dependent. It will be important to evaluate the role of ATP in this novel holdase function, and a number of straightforward experiments are now possible with the assays and techniques developed herein.

    Taken together, these studies show that the Cdc48 complex has a critical and general function as an “ERAD holdase” or retrochaperone. It has been clear for a number of years that Cdc48/p97 accompanies ERAD substrates on their way to the proteasome, but this work demonstrates that the solubility of the ubiquitinated substrates is only possible due to the chaperoning functions of the complex. There are both functional and pathological consequences of this critical new action. It will be intriguing to understand the mechanics and structural aspects of Cdc48 chaperoning and to eventually understand the breadth of this function in cellular processes, including both degradation and possible refolding of damaged proteins that engage the AAA-ATPases in the course of proteostasis.

    So, a new function for our protein, as though the “old” functions were not enough!

    Retrochaperone. 🙂

    And, again, a critical role of ubiquitin chains.

  250. DATCG @246:

    What more is there to add? Anything we’re missing or have not covered? Or to highlight?

    gpuccio has covered (along with your contributions) a substantial area of important subtopics within the main theme of this thread. But note that the number of biology-related research papers seem to increase quite rapidly revealing interesting things that had not been considered until now. I would refrain from thinking the discussion has been exhausted.
    As we saw, many details remain elusive at best.
    As outstanding questions get answered, new ones are raised. This gives the impression of a never-ending story. The complexity of the functionally specified informational organization keeps deepening with no end in sight yet.

  251. The mitotic checkpoint system ensures the fidelity of chromosome segregation in mitosis by preventing premature initiation of anaphase until correct bipolar attachment of chromosomes to the mitotic spindle is reached. It promotes the assembly of a mitotic checkpoint complex (MCC), composed of BubR1, Bub3, Cdc20, and Mad2, which inhibits the activity of the anaphase-promoting complex/cyclosome (APC/C) ubiquitin ligase. When the checkpoint is satisfied, anaphase is initiated by the disassembly of MCC. Previous studies indicated that the dissociation of APC/C-bound MCC requires ubiquitylation and suggested that the target of ubiquitylation is the Cdc20 component of MCC. However, it remained unknown how ubiquitylation causes the release of MCC from APC/C and its disassembly and whether ubiquitylation of additional proteins is involved in this process. We find that ubiquitylation causes the dissociation of BubR1 from Cdc20 in MCC and suggest that this may lead to the release of MCC components from APC/C. BubR1 in MCC is ubiquitylated by APC/C, although to a lesser degree than Cdc20. The extent of BubR1 ubiquitylation was markedly increased in recombinant MCC that contained a lysine-less mutant of Cdc20. Mutation of lysine residues to arginines in the N-terminal region of BubR1 partially inhibited its ubiquitylation and slowed down the release of MCC from APC/C, provided that Cdc20 ubiquitylation was also blocked. It is suggested that ubiquitylation of both Cdc20 and BubR1 may be involved in their dissociation from each other and in the release of MCC components from APC/C.

    Role of ubiquitylation of components of mitotic checkpoint complex in their dissociation from anaphase-promoting complex/cyclosome.
    Sitry-Shevah D, Kaisari S, Teichner A, Miniowitz-Shemtov S, Hershko A
    Proc Natl Acad Sci U S A. 2018 Feb 20;115(8):1777-1782.
    doi: 10.1073/pnas.1720312115.

  252. Gpuccio @248,

    Well OK then, more to follow 🙂 Just wanted to make sure I was staying in the right theme of things.

    Sounds great! And agree the neo-darwinist would certainly show up if they had a rebuttal.

  253. #249 Gpuccio,

    Yep agree with all you said. I think as I get an opportunity to peek inside how it’s done which is quite remarkable the explanation by Dr. Deshaies actually to do that it was very eye opening and at same time humbling at how far we have to go.

    About going upstream: I don’t know, “detecting mutations that allow tumors to form in the first place and replacing those mutations” seems still rather far away. Of course detecting tumors when they are still at the beginning would be great, but the problem is that they are really a lot of different things, from a biological point of view, even when they have similar clinical manifestations.

    I humbly have no idea where to start, just a vague understanding at all. I’m guessing openly here, much out of ignorance that if we’re looking at a designed system, then we may find a pattern of weak-links(?) so to speak? That may eventually be understood as hot spots for deleterious mutations.

    And a lot of random events are probably implied in the initial phases of the disease. Here again complexity makes it difficult for us to really understand, and unfortunately it is here the complexity of possible random devastations of extremely complex functions.

    Ohhh yes… agree at the difficulty. Maybe I’m reaching to far when I say upstream, but for some reason think it’s not out of the realm of possibility to discover in the future.

    Maybe a bit naive too. But I keep thinking if designed, then based upon environmental input – see how the branching of deleterious mutations form patterns of failure. That end in tumors. Then tracing it back upstream to the regulatory functions or master regulators even and other coordinated interdependencies.

    It’s a mouthful of networking semantic diagnosis and reverse engineering! 🙂

  254. #251 Gpuccio,

    Congrats on finding a rarely used term, RetroChaperone! 🙂

    Haha, I checked and wiki still does not have it. Only used mainly in this paper and a few others. Maybe chaperone is good enough, but retro sounds cool to identify along with retrotranslocation to the cytosol. They do speak about regulation of retro-translocation of chaperones but interestly not updated with Cdc48 yet! Wiki is falling behind 😉

    And wow… yeah Cdc48 chaperoning misfolded ER local proteins to the cytosol. Maintaining solubility by binding to ubiquitinated ERAD M-substrates – retrotranslocated.

    What can possibly go wrong? 😉

    To degrade or not to degrade, this is the question of the protein life cycle.

    I’ll review this new informatoin you provided. And I have other papers I put on hold as I’m working thru them.

    Oh, during review of other papers, maybe my use of “upstream” is inappropriate? I’ve an old habit of thinking in terms of Top-down structured programming.

    I’ll search for an example.

  255. a bit of humor to add to the mix from some college students I’m guessing.

    The sad case of a misfolded protein and UPR(Unfolded Protein Response) on the “Ugly Protein Network”

    https://www.youtube.com/watch?v=XYGlzNnHoTw

  256. #252, 253,

    Agreed 🙂 And thanks for interesting abstract on chromosome segregation in mitosis and MCC Mitotitic Checkpoint Complex. And role of Ubiquitylation.

  257. This comment is off-topic a bit. My pet peeve of Nomenclature and chaotic naming conventions of functions.

    While searching on Choromsome Segregation and ubiquitination of open access papers I came across a Chapter by Mitsuhiro Yanagida.

    Besides the main chapter, on basics of Chromosome Segregation which mentions ubiquitin interplay and roles
    Yanagida mentions Nomenclature.

    He points out the problem of Nomenclature at Chapter 2.4, pg 25. You may not be interested in this at all, but it’s 2nd person I’ve found frustrated a bit, detailing why it’s important for easy identification of functions. Another reason I personally think this is important is from a Design perspective.

    The chapter automatically opens a PDF btw for download from Springer.com…

    Basics of Chromosome Segregation – Mitsuhiro Yanagida – 2009

    I like the points he is making about recognize Functions across organisms!

    The nomenclature used for genes involved in chromosome segregation is a serious problem in communicating results obtained in different organisms. Many genes are initially identified through the use of mutants, antibodies, or amino acid sequences of purified proteins and their molecular functions are not known.

    Thus, many of the gene names do not give functional clues and are difficult to remember. Although similar proteins exist in other organisms, researchers tend to use their own organism’s nomenclature, as it is often unclear whether these genes are functionally equivalent to similar genes in other organisms.

    Indeed, genes with analogous sequences but distinct functions are not uncommon. It is therefore very difficult for researchers in other fields and for newcomers to the field to understand the functions of a particular gene by reading the literature.

    Thank you as a newcomer! 🙂 But hmmm, just as a systems molecular biologist it sure seems cumbersome, chaotic and unproductive as well.

    A number of protein complexes essential for chromosome segregation, however, have been given common names across organisms. The presence of multiple subunits that all share sequence similarity in different organisms is convincing evidence of the functional similarity of these complexes, such as condensin, cohesin, anaphase-promoting complex (APC/C), and mitotic checkpoint complex (MCC).

    The use of a common nomenclature for these complexes promotes integrated studies.

    For example, condensin is a hetero-pentameric complex required for mitotic chromosome architecture. It consists of two subunits belonging to the structural maintenance of chromosome (SMC) ATPase protein family, and three
    non-SMC components (reviewed in Nasmyth and Haering 2005, Belmont 2006, Hirano 2006). Frog condensin contains XCAP-C (SMC4) and XCAP-E (SMC2), two heterodimeric coiled-coil SMCs and three non-SMC proteins: XCAP-H, -G, and -D2. In S. cerevisiae, the dimeric Smc2 and Smc4 associate with three nonSMC subunits, Ycg1, Ycs4, and Brn1. Similarly, two SMC proteins of S. pombe, Cut3 and Cut14, form a heterodimer and bind to three non-SMC subunits, Cnd1, Cnd2, and Cnd3 (Nasmyth and Haering 2005, Belmont 2006, Hirano 2006).

    The sequences of each of these sets of five subunits are similar from fungi to human, indicating that they are functionally conserved.

    Although different names remain for individual subunits, they are less important than those of complexes.

    Complexes required for chromosome segregation are often multifunctional. Condensin (see above) is also required for interphase activities, such as DNA-damage repair (Heale et al., 2006).

    Cohesin, the multiprotein complex that holds sister chromatids together following DNA replication, is also
    required for DNA-damage repair (Strom et al., 2007, Unal et al., 2007, Ball¨ and Yokomori 2008) and developmental transcriptional regulation (Dorsett et al., 2005, Dorsett 2007, Gullerova and Proudfoot 2008, Wendt et al., 2008).

    The name, usually based on the initially discovered function, might only partially represent the functions mediated by the complex and could be misleading.

    Therefore, biologists and geneticists should use caution when naming a complex according to its originally discovered function.

    The anaphase-promoting complex/cyclosome (APC/C) has an instructive history with regard to the naming. The APC/C was discovered as a complex and called a cyclosome (Sudakin et al., 1995), as it is essential for the degradation
    of mitotic cyclin in vitro. This same complex was also called the APC, as it was defined as an anaphase-promoting complex (King et al., 1995). The APC/C, which contains 15 subunits (Passmore et al., 2005), is the E3 ubiquitin ligase that poly-ubiquitylates mitotic cyclin and securin for degradation in a destruction-box (DB)-dependent manner (reviewed in Sullivan and Morgan 2007).

    APC/C activation is inhibited by the spindle assembly checkpoint (also called the spindle checkpoint or mitotic checkpoint; see Chapter 11). Poly-ubiquitylated
    cyclin and securin are rapidly degraded by the 26S proteasome,
    leading to the activation of separase, the cleavage of cohesin, the separation of the sister
    chromatids, and the onset of anaphase (Morgan 2006, Fig. 2.1).

    Because the abbreviation APC also refers to the frequently cited tumor suppressor protein adenomatous polyposis coli, it is currently recommended that the abbreviation
    APC/C be used to avoid confusion. This distinction has become particularly necessary as the tumor suppressor APC interacts with the plus ends of the microtubules and is implicated in the spindle checkpoint (Draviam et al., 2006).

    While the APC/C regulates the exit of mitosis in dividing cells (Sullivan and Morgan 2007), it is also abundant in non-dividing cells such as neurons and muscles (van Roessel et al., 2004, Zarnescu and Moses 2004). The APC/C seems
    to have a postmitotic role at Drosophila neuromuscular synapses: in neurons, the APC/C controls synaptic size, and in muscles, it regulates synaptic transmission
    (van Roessel et al., 2004). The roles of the APC/C in non-dividing differentiated cells are elusive, but clearly different from its role in mitotic progression and exit.

    Thus, a new name, particularly one based on a single function, could cause misconceptions concerning the roles of these complexes.

    Thank you Mitsuhiro Yanagida! He deserves an award for common sense 🙂 Not much about Ubiquitin but thank you for elucidating the problems of naming conventions across disparate areas.

  258. @215 update the list – add the following item from gpuccio @251::

    Neal, Sonya, Raymond Mak, Eric J. Bennett, and Randolph Hampton. “A Cdc48 ‘Retrochaperone’ Function Is Required for the Solubility of Retrotranslocated, Integral Membrane Endoplasmic Reticulum-Associated Degradation (ERAD-M) Substrates.” Journal of Biological Chemistry 292, no. 8 (February 24, 2017): 3112–28. https://doi.org/10.1074/jbc.M116.770610.

  259. Dionisio, Gpuccio,

    Question, do either of you have sources for images of Proteins that you like to refer to? Or for any active process and genetic material? If so, please share. Would like to build up different resources for viewing.

    Came across a resource trying to find images of Ubiquitin Proteins. This is of WWP1(WW domain containing E3 ubiquitin protein ligase 1) This includes a HECT domain.

    Atlas of Genetics and Cytogenetics in Oncology and Haematology – WWP1 containing E3 ubiquitin ligase 1 – Alias AIP5

    The images of above link are of WWP1 expression in 22Rv1 prostate cancer cell line.

    Descriptions, notations and info…

    A: WWP1 protein

    B: Exogenous WWP1 expression in the 22Rv1 prostate cancer cell line was detected under a confocal microscopy. The endosomes are indicated by GFP-Rab5.

    C: Protein structure of WWP1
    Description: 922 amino acids; approximatively 110 kDa protein; The C2 domain at N-terminus is responsible for calcium-dependent phospholipid binding. The four WW domains in the middle are responsible for protein-protein interaction with PY motifs. The HECT domain at the C-terminus is responsible for the ubiquitin transfer. The Cystein 890 is the catalytic center. The underlined WWP1 substrates do not have a PY motif (PPXY). A smaller WWP1 protein isoform was detected in two prostate cancer cell lines PC-3 and LAPC-4 (Chen C, 2007).
    Protein structure: The HECT domain of WWP1 (see Figure 2C.)(Verdecia MA., 2003).

    Expression: The WWP1 protein is lowly expressed in normal prostate and breast but is frequently upregulated in prostate and breast cancers due to the gene amplification.

    Localisation: Predominately on membrane structures in cytoplasm and occasionally in nucleus (see Figure 2B.).

    Function: WWP1 is an E3 ubiquitin ligase.

    WWP1 negatively regulates the transforming growth factor-beta (TGF-b) signaling by targeting its molecular components, including TGF-beta receptor 1 (TbR1) (Komuro A, 2004), Smad2 (Seo SR, 2004), and Smad4 (Moren A., 2005) for ubiquitin mediated degradation.

    In addition, WWP1 has been reported to target the epithelial Na+ channel (ENaC) (Malbert-Colas L, 2003), Notch (Shaye DD, 2005), Runx2 (Jones DC, 2006; Shen R, 2006), KLF2 (Zhang X, 2004), and KLF5 (Chen C, 2005) for ubiquitin-mediated proteolysis.

    Recently, WWP1 has been demonstrated to inhibit p53 activity through exporting p53 from the nucleus after ubiquitination (Laine A,.2007). Overall, WWP1 may play a pro-survival role in several tumor types including breast (Chen C, 2007) and prostate (Chen C, 2007). WWP1 has also shown to promote virus budding (Martin-Serrano J, 2005; Heidecker G, 2007).

    Homology: WWP1 belongs to the C2-WW-HECT E3 family which contains 8 other members (Chen C, 2007). The WWP1 gene is highly-conserved among species (from human to c. elegant).

    Mutations

    Somatic: The WWP1 gene is rarely mutated in human prostate cancer (Chen C, 2007). Two sequence alterations were detected in prostate cancer xenografts. One was 2393A–>T (Glu798Val) in CWR91 and the other was 721A–>T (Thr241Ser) in LuCaP35. Additionally, some mutations in the HECT domain decrease the E3 ligase activity (Verdecia MA., 2003).

  260. Morning Dio 🙂 Have a good day. I’m out for now.

  261. DATCG @246:

    What more is there to add? Anything we’re missing or have not covered? Or to highlight?

    gpuccio @251:

    So, a new function for our protein, as though the “old” functions were not enough!
    Retrochaperone. 🙂
    And, again, a critical role of ubiquitin chains.

  262. DATCG @259,
    That’s interesting.

    DATCG @261,
    That’s interesting.

    DATCG @262,
    Thanks.

  263. Dionisio @253:

    You too are not bad at picking pèapers which “just came out of the printing press”! 🙂

    I specially liked this phrase:

    “When the checkpoint is satisfied, anaphase is initiated by the disassembly of MCC.”

    (Emphasis mine)

    After all, design is the tool to satisfy a desire. Maybe ubiqutin chians have a role in expressing satisfaction, too! 🙂

  264. DATCG:

    “And agree the neo-darwinist would certainly show up if they had a rebuttal.”

    Maybe they are simply shy! 🙂

  265. DATCG at #255:

    “It’s a mouthful of networking semantic diagnosis and reverse engineering!”

    Yes, but unfortunately sometimes it’s easier to buid something again than to repair it.

    The problem with neoplastic cells is that, once the initial transformation takes place, a lot of further mutations or functional impairments is very likely to follow.

    That’s also the reason for resistance to therapy in relapsed neploasias.

  266. DATCG at #256:

    It’s great to be ahead of Wikipedia! 🙂 🙂

  267. gpuccio @265:

    After all, design is the tool to satisfy a desire. Maybe ubiquitin chains have a role in expressing satisfaction, too!

    [emphasis added]

    That reminds me of some loud musicians that couldn’t get no satisfaction, even though they kept trying (or at least that’s what they claimed) since the mid 1960s. 🙂

    Maybe they didn’t know much about ubiquitin back then? 🙂

  268. DATCG @254:
    “And agree the neo-darwinist would certainly show up if they had a rebuttal.”

    gpuccio @266:
    “Maybe they are simply shy!”

    No, they aren’t shy at all! 🙂

    The most probable reason behind their conspicuous absence here is that most of what is discussed can be easily explained through RV+NS, hence this discussion is trivial and not worth their time. 🙂

  269. Off topic.

    CHARLEMAGNE DISTINGUISHED LECTURE SERIES
    with Prof. Denis Noble Ph.D.
    Title: From Pacing the Heart to the Pace of Evolution
    Abstract
    Multi-mechanism interpretations of cardiac pacemaker function reveal the extent to which many physiological functions are buffered against genomic change. Contrary to Schrodinger’s claim in What is Life? (1944) which led to the Central Dogma of Molecular Biology (Crick 1970), biological functions at higher levels harness stochasticity at lower levels. This harnessing of stochasticity is a prerequisite for the processes by which the pace of evolution can be accelerated through guided control of mutation rates and of buffering by regulatory networks in organisms.
    Schrodinger E. 1944 What is life? Cambridge, UK: Cambridge University Press.
    Crick FHC. 1970 Central dogma of molecular biology. Nature 227, 561 – 563. (doi:10.1038/227561a0)
    Noble D. Dance to the Tune of Life. Biological Relativity. Cambridge University Press 2016
    Noble D. Evolution viewed from physics, physiology and medicine. Interface Focus 2017, 7, 20160159.
    Noble R & Noble D. Was the Watchmaker Blind? Or was she One-eyed? Biology, 2017, 6, 47.

    Check this out at your convenience.

    If you don’t want to watch the whole video, just skip to around the mark 30:00 and listen to the last ten minutes.

    The visual part of the presentation is not very clear, which seems like a defect of the way the presentation was recorded. Maybe there is a better video of the same lecture?

    https://www.youtube.com/embed/XbS2dwn04fQ

    https://www.aices.rwth-aachen.de/en/

  270. @271 addendum

    isn’t professor Denis Noble one of the pioneers of systems biology and founder of the 3rd way (the raft some evolutionists are using to jump out of the sinking neo-Darwinian ship)?

  271. The Ubiquitin Ligase (E3) Psh1p Is Required for Proper Segregation of both Centromeric and Two-Micron Plasmids in Saccharomyces cerevisiae
    Meredith B. Metzger, Jessica L. Scales, Mitchell F. Dunklebarger and Allan M. Weissman
    G3: Genes, Genomes, Genetics November 1, 2017 vol. 7 no. 11 3731-3743; https://doi.org/10.1534/g3.117.300227

    http://www.g3journal.org/conte.....1.full.pdf

  272. Dionisio at #273:

    Interesting.

    This strange E3 ligase, Psh1p, 406 AAs long, is practically taxonomycally restricted to Saccharomycetes. This is really amazing.

    Indeed, it shares practically no homology (except for a few low hits limited to the RING domain) with any organism outside of fungi, and even in fungi the homology is rather low (100 – 200 bits) outside of Saccharomycetes.

    Its function remains elusive, even after reading the interesting paper you linked. Its only known target seems to be CSE4p, a strange Histone H3 like protein, 229 AAs long. From Uniprot:

    Histone H3-like variant which exclusively replaces conventional H3 in the nucleosome core of centromeric chromatin at the inner plate of the kinetochore. Required for recruitment and assembly of kinetochore proteins, mitotic progression and chromosome segregation. May serve as an epigenetic mark that propagates centromere identity through replication and cell division. Required for functional chromatin architecture at the yeast 2-micron circle partitioning locus and promotes equal plasmid segregation.

    This strange variant seems, too, essentially restricted to Saccharomycetes, except for the partial homology (about 130 bits) to histone H3 in the C terminal part.

    So, this complex biological system linked to yeast plasmids seems to be a remarkable example of taxonomically restricted complexity.

    Involving, of course, ubiquitin! 🙂

  273. DATCG at #259:

    I agree with you that protein nomenclature is often misleading. Proteins that are clear homologues in many organisms receive often a lot of different names. You can find that multitude of names in their Uniprot record, usually.

    For example, our much discussed p97 is reported in Uniprot, for humans, as:

    TERA_HUMAN: Transitional endoplasmic reticulum ATPase

    But also:

    TER ATPase

    VCP: Valosin-containing protein

    15S Mg(2+)-ATPase p97 subunit

    and we know that, in yeast, it is called:

    CDC48: Cell division control protein 48

    In papers, you can often find those different names, and it can be difficult sometimes to understand that some papers are referring to the same protein!

    And, in this case, we are talking of a very conserved protein, and there can be no doubt that human TERA and yeast CDC48 are homologues, because they share 1178 bits and 68% identities and 83% positives.

  274. DATCG, Dionisio:

    Again our friend TERA/VCP/p97/CDC48, in some new role! 🙂

    The following paper is of January 2018:

    Cdc48 regulates a deubiquitylase cascade critical for mitochondrial fusion

    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5798933/

    (Public access)

    Abstract:

    Cdc48/p97, a ubiquitin-selective chaperone, orchestrates the function of E3 ligases and deubiquitylases (DUBs). Here, we identify a new function of Cdc48 in ubiquitin-dependent regulation of mitochondrial dynamics. The DUBs Ubp12 and Ubp2 exert opposing effects on mitochondrial fusion and cleave different ubiquitin chains on the mitofusin Fzo1. We demonstrate that Cdc48 integrates the activities of these two DUBs, which are themselves ubiquitylated. First, Cdc48 promotes proteolysis of Ubp12, stabilizing pro-fusion ubiquitylation on Fzo1. Second, loss of Ubp12 stabilizes Ubp2 and thereby facilitates removal of ubiquitin chains on Fzo1 inhibiting fusion. Thus, Cdc48 synergistically regulates the ubiquitylation status of Fzo1, allowing to control the balance between activation or repression of mitochondrial fusion. In conclusion, we unravel a new cascade of ubiquitylation events, comprising Cdc48 and two DUBs, fine-tuning the fusogenic activity of Fzo1.

    Mitochondrial fusion?

    Yes, because we learn that:

    Mitochondria are little compartments within a cell that produce the energy needed for most biological processes. Each cell possesses several mitochondria, which can fuse together and then break again into smaller units. This fusion process is essential for cellular health.

    Mitochondria are dynamic organelles constantly undergoing fusion and fission events, modulated by a variety of post-translational modifiers including ubiquitin

    The ubiquitin-specific chaperone Cdc48/p97 is required to maintain mitochondrial morphology (Esaki and Ogura, 2012). However, the underlying molecular mechanism of how Cdc48 regulates mitochondrial dynamics is not understood.

    Here, we identify a role of Cdc48 in mitochondrial fusion, as part of a novel enzymatic cascade consisting of Cdc48, Ubp12 and Ubp2. Cdc48 negatively regulates Ubp12, which negatively regulates Ubp2, explaining why these two DUBs exert opposite effects on their targets and on ubiquitin homeostasis.

    If someone reading this thread is starting to believe that I am probably making up things, I am certainly not offended! 🙂

  275. gpuccio @276:

    If someone reading this thread is starting to believe that I am probably making up things, I am certainly not offended!

    Yes, it seems like “fake news” indeed. 🙂

  276. The endoplasmic reticulum (ER) serves as a warehouse for factors that augment and control the biogenesis of nascent proteins entering the secretory pathway. In turn, this compartment also harbors the machinery that responds to the presence of misfolded proteins by targeting them for proteolysis via a process known as ER-associated degradation (ERAD). During ERAD, substrates are selected, modified with ubiquitin, removed from the ER, and then degraded by the cytoplasmic 26S proteasome. While integral membrane proteins can directly access the ubiquitination machinery that resides in the cytoplasm or on the cytoplasmic face of the ER membrane, soluble ERAD substrates within the lumen must be retrotranslocated from this compartment. In either case, nearly all ERAD substrates are tagged with a polyubiquitin chain, a modification that represents a commitment step to degrade aberrant proteins. However, increasing evidence indicates that the polyubiquitin chain on ERAD substrates can be further modified, serves to recruit ERAD-requiring factors, and may regulate the ERAD machinery. Amino acid side chains other than lysine on ERAD substrates can also be modified with ubiquitin, and post-translational modifications that affect substrate ubiquitination have been observed. Here, we summarize these data and provide an overview of questions driving this field of research.

    The evolving role of ubiquitin modification in endoplasmic reticulum-associated degradation
    G. Michael Preston, Jeffrey L. Brodsky
    Biochemical Journal
    Feb 03, 2017,
    474
    (4)
    445-469;
    DOI: 10.1042/BCJ20160582
    http://biochemj.org/lookup/doi/10.1042/BCJ20160582

  277. Mitochondrial integrity relies on homotypic fusion between adjacent outer membranes, which is mediated by large GTPases called mitofusins. The regulation of this process remains nonetheless elusive. Here, we report a crosstalk between the ubiquitin protease Ubp2 and the ubiquitin ligases Mdm30 and Rsp5 that modulates mitochondrial fusion. Ubp2 is an antagonist of Rsp5, which promotes synthesis of the fatty acids desaturase Ole1. We show that Ubp2 also counteracts Mdm30-mediated turnover of the yeast mitofusin Fzo1 and that Mdm30 targets Ubp2 for degradation thereby inducing Rsp5-mediated desaturation of fatty acids. Exogenous desaturated fatty acids inhibit Ubp2 degradation resulting in higher levels of Fzo1 and maintenance of efficient mitochondrial fusion. Our results demonstrate that the Mdm30-Ubp2-Rsp5 crosstalk regulates mitochondrial fusion by coordinating an intricate balance between Fzo1 turnover and the status of fatty acids saturation. This pathway may link outer membrane fusion to lipids homeostasis.

    Cavellini, Laetitia, Julie Meurisse, Justin Findinier, Zoi Erpapazoglou, Naïma Belgareh-Touzé, Allan M. Weissman, and Mickael M. Cohen. “An Ubiquitin-Dependent Balance between Mitofusin Turnover and Fatty Acids Desaturation Regulates Mitochondrial Fusion.” Nature Communications 8 (June 13, 2017): 15832. https://doi.org/10.1038/ncomms15832.

  278. A conserved AAA+ ATPase, called Cdc48 in yeast and p97 or VCP in metazoans, plays an essential role in many cellular processes by segregating polyubiquitinated proteins from complexes or membranes. For example, in endoplasmic reticulum (ER)-associated protein degradation (ERAD), Cdc48/p97 pulls polyubiquitinated, misfolded proteins out of the ER and transfers them to the proteasome. Cdc48/p97 consists of an N-terminal domain and two ATPase domains (D1 and D2). Six Cdc48 monomers form a double-ring structure surrounding a central pore. Cdc48/p97 cooperates with a number of different cofactors, which bind either to the N-terminal domain or to the C-terminal tail. The mechanism of Cdc48/p97 action is poorly understood, despite its critical role in many cellular systems. Recent in vitro experiments using yeast Cdc48 and its heterodimeric cofactor Ufd1/Npl4 (UN) have resulted in novel mechanistic insight. After interaction of the substrate-attached polyubiquitin chain with UN, Cdc48 uses ATP hydrolysis in the D2 domain to move the polypeptide through its central pore, thereby unfolding the substrate. ATP hydrolysis in the D1 domain is involved in substrate release from the Cdc48 complex, which requires the cooperation of the ATPase with a deubiquitinase (DUB). Surprisingly, the DUB does not completely remove all ubiquitin molecules; the remaining oligoubiquitin chain is also translocated through the pore. Cdc48 action bears similarities to the translocation mechanisms employed by bacterial AAA ATPases and the eukaryotic 19S subunit of the proteasome, but differs significantly from that of a related type II ATPase, the NEM-sensitive fusion protein (NSF). Many questions about Cdc48/p97 remain unanswered, including how it handles well-folded substrate proteins, how it passes substrates to the proteasome, and how various cofactors modify substrates and regulate its function.

    Bodnar, Nicholas, and Tom Rapoport. “Toward an Understanding of the Cdc48/P97 ATPase.” F1000Research 6 (August 3, 2017): 1318. https://doi.org/10.12688/f1000research.11683.1.

  279. For over a century, the abnormal movement or number of centrosomes has been linked with errors of chromosomes distribution in mitosis. While not essential for the formation of the mitotic spindle, the presence and location of centrosomes has a major influence on the manner in which microtubules interact with the kinetochores of replicated sister chromatids and the accuracy with which they migrate to resulting daughter cells. A complex network has evolved to ensure that cells contain the proper number of centrosomes and that their location is optimal for effective attachment of emanating spindle fibers with the kinetochores. The components of this network are regulated through a series of post-translational modifications, including ubiquitin and ubiquitin-like modifiers, which coordinate the timing and strength of signaling events key to the centrosome cycle. In this review, we examine the role of the ubiquitin system in the events relating to centriole duplication and centrosome separation, and discuss how the disruption of these functions impacts chromosome segregation.

    Zhang, Ying, and Paul J. Galardy. “Ubiquitin, the Centrosome, and Chromosome Segregation.” Chromosome Research 24, no. 1 (January 2016): 77–91. https://doi.org/10.1007/s10577-015-9511-7.

    https://www.researchgate.net/profile/Paul_Galardy/publication/287971598_Ubiquitin_the_centrosome_and_chromosome_segregation/links/5759653208ae9a9c954ed1f7/Ubiquitin-the-centrosome-and-chromosome-segregation.pdf

  280. Post-translational modification of proteins by ubiquitylation is increasingly recognised as a highly complex code that contributes to the regulation of diverse cellular processes. In humans, a family of almost 100 deubiquitylase enzymes (DUBs) are assigned to six subfamilies and many of these DUBs can remove ubiquitin from proteins to reverse signals. Roles for individual DUBs have been delineated within specific cellular processes, including many that are dysregulated in diseases, particularly cancer. As potentially druggable enzymes, disease-associated DUBs are of increasing interest as pharmaceutical targets. The biology, structure and regulation of DUBs have been extensively reviewed elsewhere, so here we focus specifically on roles of DUBs in regulating cell cycle processes in mammalian cells. Over a quarter of all DUBs, representing four different families, have been shown to play roles either in the unidirectional progression of the cell cycle through specific checkpoints, or in the DNA damage response and repair pathways. We catalogue these roles and discuss specific examples. Centrosomes are the major microtubule nucleating centres within a cell and play a key role in forming the bipolar mitotic spindle required to accurately divide genetic material between daughter cells during cell division. To enable this mitotic role, centrosomes undergo a complex replication cycle that is intimately linked to the cell division cycle. Here, we also catalogue and discuss DUBs that have been linked to centrosome replication or function, including centrosome clustering, a mitotic survival strategy unique to cancer cells with supernumerary centrosomes.

    Darling, Sarah & Fielding, Andrew & Sabat-Po?piech, Dorota & Prior, Ian & Coulson, Judy. (2017). Regulation of the cell cycle and centrosome biology by deubiquitylases. Biochemical Society Transactions. 45. BST20170087. 10.1042/BST20170087.

    http://www.biochemsoctrans.org.....l-text.pdf

  281. DATCG and gpuccio,

    Please, be alert for repeated references. I may have messed up some required steps in the Zotero rules, causing some papers to get posted twice by mistake. Just raise a red flag if you notice such a case. Thanks.

  282. Ubiquitin-specific protease 15 (USP15) is a widely expressed deubiquitylase that has been implicated in diverse cellular processes in cancer. Here we identify topoisomerase II (TOP2A) as a novel protein that is regulated by USP15. TOP2A accumulates during G2 and functions to decatenate intertwined sister chromatids at prophase, ensuring the replicated genome can be accurately divided into daughter cells at anaphase. We show that USP15 is required for TOP2A accumulation, and that USP15 depletion leads to the formation of anaphase chromosome bridges. These bridges fail to decatenate, and at mitotic exit form micronuclei that are indicative of genome instability. We also describe the cell cycle-dependent behaviour for two major isoforms of USP15, which differ by a short serine-rich insertion that is retained in isoform-1 but not in isoform-2. Although USP15 is predominantly cytoplasmic in interphase, we show that both isoforms move into the nucleus at prophase, but that isoform-1 is phosphorylated on its unique S229 residue at mitotic entry. The micronuclei phenotype we observe on USP15 depletion can be rescued by either USP15 isoform and requires USP15 catalytic activity. Importantly, however, an S229D phospho-mimetic mutant of USP15 isoform-1 cannot rescue either the micronuclei phenotype, or accumulation of TOP2A. Thus, S229 phosphorylation selectively abrogates this role of USP15 in maintaining genome integrity in an isoform-specific manner. Finally, we show that USP15 isoform-1 is preferentially upregulated in a panel of non-small cell lung cancer cell lines, and propose that isoform imbalance may contribute to genome instability in cancer. Our data provide the first example of isoform-specific deubiquitylase phospho-regulation and reveal a novel role for USP15 in guarding genome integrity.

    Fielding, Andrew & Concannon, Matthew & Darling, Sarah & V. Rusilowicz-Jones, Emma & Sacco, Joseph & Prior, Ian & J. Clague, Michael & Urbé, Sylvie & Coulson, Judy. (2018). The deubiquitylase USP15 regulates topoisomerase II alpha to maintain genome integrity. Oncogene. 10.1038/s41388-017-0092-0.

    https://www.researchgate.net/publication/323127826_The_deubiquitylase_USP15_regulates_topoisomerase_II_alpha_to_maintain_genome_integrity/fulltext/5a81cb2aa6fdcc6f3ead658d/323127826_The_deubiquitylase_USP15_regulates_topoisomerase_II_alpha_to_maintain_genome_integrity.pdf

  283. Deregulation of centriole duplication has been implicated in cancer and primary microcephaly. Accordingly, it is important to understand how key centriole duplication factors are regulated. E3 ubiquitin ligases have been implicated in controlling the levels of several duplication factors, including PLK4, STIL and SAS-6, but the precise mechanisms ensuring centriole homeostasis remain to be fully understood. Here, we have combined proteomics approaches with the use of MLN4924, a generic inhibitor of SCF E3 ubiquitin ligases, to monitor changes in the cellular abundance of centriole duplication factors. We identified human STIL as a novel substrate of SCF-?TrCP. The binding of ?TrCP depends on a DSG motif within STIL, and serine 395 within this motif is phosphorylatedin vivoSCF-?TrCP-mediated degradation of STIL occurs throughout interphase and mutations in the DSG motif causes massive centrosome amplification, attesting to the physiological importance of the pathway. We also uncover a connection between this new pathway and CDK2, whose role in centriole biogenesis remains poorly understood. We show that CDK2 activity protects STIL against SCF-?TrCP-mediated degradation, indicating that CDK2 and SCF-?TrCP cooperate via STIL to control centriole biogenesis.

    Arquint, Christian & Cubizolles, Fabien & Morand, Agathe & Schmidt, Alexander & Nigg, Erich. (2018). The SKP1-Cullin-F-box E3 ligase ?TrCP and CDK2 cooperate to control STIL abundance and centriole number. Open Biology. 8. 170253. 10.1098/rsob.170253.

    https://www.researchgate.net/profile/Alexander_Schmidt3/publication/323170017_The_SKP1-Cullin-F-box_E3_ligase_bTrCP_and_CDK2_cooperate_to_control_STIL_abundance_and_centriole_number/links/5aa1389da6fdcc22e2d10921/The-SKP1-Cullin-F-box-E3-ligase-bTrCP-and-CDK2-cooperate-to-control-STIL-abundance-and-centriole-number.pdf

  284. #267

    “Yes, but unfortunately sometimes it’s easier to buid something again than to repair it.”

    Oh, like what you’re pointing out. So, “naturally” speaking or by Design, we have multiple routes to organized redistribution and/or total destruction of proteins.

    The Proteaosome itself is not total destruction of all cellular matter, correct? It’s not a garbage disposal per say as an apt analogy? The proteins, misfolded, etc., go in and are broken down to component parts that can then be recycled for new parts, correct?

    I’m bypassing or leaving out the full spectrum. But there’s apoptosis and other methods as well.

    To add, we are expected to believe that a system decision like this – to prevent proteolysis, or to allow, then recycle is by a blind, unguided RM & NS “process.”

    “The problem with neoplastic cells is that, once the initial transformation takes place, a lot of further mutations or functional impairments is very likely to follow.”

    Agree!

    “That’s also the reason for resistance to therapy in relapsed neploasias.”

    Agree again, so my question is, what is correctly terminology for molecular biology? I used “upstream” for me meaning to a) detect, b) correct the problem prior to neoplasia. Is that to difficult? Is the current process that corrects missing critical points of mutation? And can it be… hmmm, helped to recognize them?

    I may be assuming to much to take on here from an overall systems perspective.

  285. Dionisio, you are on a roll 🙂

    LOL @RollingStone reference.

  286. Playing devil’s advocate since we have no participation by opponents to Design in favor of a blind, unguided “process,” I’m putting my Hunter-cap on. You may want to Google it. 😉

    (ps if this is considered to off-topic we can discuss another time)

    The challenge..

    “The hslV protein has been hypothesized to resemble the likely ancestor of the 20S proteasome.HslV is generally not essential in bacteria, and not all bacteria possess it, while some protists possess both the 20S and the hslV systems.”

    So, is hslV a possible ancestor to 20S Proteasome? Could it be? Might it be?

    These scientist may have found a likely candidate which might be related to an ancestral gene, which could be a breakthrough in understanding the possible evolution of the Proteasome by natural sequence of events by a gradual process of random mutations and natural selection.

    Evolution of Proteasome Regulators in Eukaryotes

    The 20S (alpha) and (beta) subunits share structural similarity and likely originated from an ancestral gene that duplicated before the divergence of archaea and eukaryotes (Gille et al. 2003). In contrast to the 20S proteasome, the evolutionary history of PAs remains fragmentary and scattered. Here, we present a comprehensive view of the evolution of the three types of activators and of PI31 from archaeal to eukaryotic lineages, using the classification of eukaryotes recently revised by Adl et al. (2012). We examined genomic data available for a total of 17 clades, spreading over 3.5 billion years of evolution and covering archaea and most of the eukaryote supergroups, that is, Opisthokonta (including Metazoans, Choanoflagellida, Ichthyosporea, and Fungi), Amoebozoans, Excavates (including Metamonads [Diplomonadida and Parabasalia] and Discoba [Heterolobosea and Englenozoa/Kinetoplastids]), Archaeplastida (Choloroplastida and Rhodophyceae), SAR (Stramenopiles, Alveolates, and Rhizaria), and two unclassified clades, Cryptophyta and Haptophyta, previously classified as Chromalveolates with the SAR group. We show that the full current repertoire of proteasome regulators was already present in the last eukaryotic common ancestor (LECA) and has subsequently evolved through independent duplication/loss events in specific lineages.

  287. #267 Gpuccio, follow-up to 287,

    I may be asking wrong questions and mistaken on pathways to tumor cells. If so, it explains my confusion of a Proteasome “lockout” solution to stop cancer cells from growing and elimination of them. I do recognize it is a solution, but was thinking there might be more efficient methods with less side effects in treatment of multiple myeloma by Kyprolis. As an example, but no means trying to target any specific medication.

    Certainly it works in a certain percentage of patients.

    But it can lead to other consequences in patients. So it’s knocking out one problem, but creating another.

    I don’t know of better solutions, but thinking if we were to look at the different pathways to the cancerous cells, what logical points along the way would we find the breakdown(deleterious mutations) and then see if there’s an alternative solution to a blocking attempt in the proteasome. Maybe a recognition of mutation prior to the signal for degradation is a way. Not easy, but maybe in breakdown of systems immunity, there’s a missing conditional check of mutations by error correction.

    Is it even feasible to think of adding such a new “check” for error correction. And then what would be downside of doing so.

    If there’s a SNP, point mutation, or… well, in searching came across this a Stop Codon mutation and a way to correct it in research done in Yeast.

    It’s a fairly good review of why this is so difficult as well.

    http://sitn.hms.harvard.edu/flash/2011/issue97/

    These are broad and difficult questions I know or researchers would already have these answers.

  288. follow up to #290…

    Recognizing different solutions may or may not be advantageous dependent upon different mechanisms within cells and error correction features.

    I was thinking one reason to use Error Correction is it already exist as a functional step.

    It would be like adding another Conditional Check? Maybe.

    But recognize researchers do not know all the steps to simply add a new check feature(or override) at this time. Nor do researchers know all the rules especially in human cells. But the Harvard article gives me hope.

  289. Gpuccio,
    I am reminded now of your Open Access paper you referenced at #89, Ubiquitin Enzymes in the Regulation of Immune Responses

    and Figure 3… 😉

    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5490640/figure/F0003/

    So, yeah, whew… trying to impact these steps, rules interactions is mind boggling.

    So we have, trying to think through this multiple checks and balances on disease fighting systems, heavily regulated by Ubiquitin Systems and DUBS, etc.

    Maybe I’ve conflated the two in my rush to think of different solutions. Though I’m guessing SNPs can cause problems for E1, E2, and E3 steps breaking down, then the cascade of steps following.

    OK, apologies for going to far off topic.

  290. Dionisio at #278 and 279:

    These two processes of ERAD (ER-associated degradation, with associated retrotranslocation) and Mitochondrial fusion are really surprising.

    They are both critically dependant on an unexpected dynamin plasticity of membranes in inner organelles. I am not suprised at all, instead, that the related mechanisms and controls remain “poorly understood” or “elusive”.

    But ubiquitin certainly a major role in both.

    Now, that’s really, really weird! That the same regulation system, which is in itself as complex and multi-faceted as we have seen in this thread, can at the same time be the key regulator in such different processes (together with all the others we have discussed) is IMO mind-blogging.

    I really cannot imagine any way to put such a system to work with any bottom-up strategy, even if designed. You really need a strict top-down engineering to get that kind of results. And even then, you need an unbelievable attention to details and connections between systems, and a perfect control of the symbolic system you are using.

    The cross-talk between inner compartments in the cell is really a fascinating issue: too often we think of the cell as some rather homogeneous environment, or we just acknowledge the separation between nucleus and cytoplasm in eukaryotes. But the cytoplasm is anything but homogeneous. Organelles are separated by membranes, and membranes, as we have seen, are dynamic tools which are almost magically shaped and reshaped by complex molecular systems.

    And, even where mebranes are not present, a lot of functional sub-sections can be dynamically assessed, continuosly created and destroyed, shaping a functional landscape of cytoplasmic events which we still have to start to understand: second messengers, signaling pathways, and so on. No part of cytoplasm is the same as any other, each landscape is unique and functional.

    See, for example, here:

    Membrane-bound organelles versus membrane-less compartments and their control of anabolic pathways in Drosophila

    https://www.sciencedirect.com/science/article/pii/S0012160617300131

    (Public access)

    Abstract

    Classically, we think of cell compartmentalization as being achieved by membrane-bound organelles. It has nevertheless emerged that membrane-less assemblies also largely contribute to this compartmentalization. Here, we compare the characteristics of both types of compartmentalization in term of maintenance of functional identities. Furthermore, membrane less-compartments are critical for sustaining developmental and cell biological events as they control major metabolic pathways. We describe two examples related to this issue in Drosophila, the role of P-bodies in the translational control of gurken in the Drosophila oocyte, and the formation of Sec bodies upon amino-acid starvation in Drosophila cells.

  291. Dionisio, Gpuccio,

    When I began looking at ERAD, and translocation, or retrotranslocation, I was like, wow, wow, wow… when you posted on retroChaperones.

    Great papers Dionisio, now when is there time to review all of these ubiquitin-related networks and functions?

    🙂

  292. #293 Cpuccio, interesting…

    It is nevertheless emerging that cell compartmentalization is also achieved by steady-state membrane-less assemblies in the nucleus, such as nucleoli, Cajal bodies and nuclear speckles, and in the cytoplasm, such as RNA based C. elegans P-granules, P-bodies, ribosomes, as well as others that do not contain RNA, like centrosome, proteasome and aggresome (Rajan et al., 2001).

    In this case, what is “steady-state” referring too?

    Also, here is “intrinsically disordered domains”

    They are flexible and have the propensity to adopt a large range of conformations. These proteins often display intrinsically disordered domains that have low complexity sequences (Huntley and Golding, 2002). Low complexity sequences are regions of poor amino-acid diversity, such as repeats of certain amino-acids (Q, N, S, G, Y, R) in prion-like domains (Alberti et al., 2009), repeats of alternating charges, such as RG, and other domains without regular sequences.

    Then ubiquitination…

    Last, it is also clear that stress specific post-translational modifications promote the formation of membrane-less compartments in vivo and phase separation in vitro (Han et al., 2012; Kato et al., 2012). This is case for SUMOylation and phosphorylation (Banani et al., 2016), ubiquitination for proteasome storage granules (Peters et al., 2013), poly-ADP ribosylation (Leung et al., 2011) and mono-ADP-ribosylation (Aguilera-Gomez et al., 2016). Conversely, arginine methylation by PRMT1 has been shown to be inhibitory (Jun et al., 2017; Nott et al., 2015), and phosphorylation by DYRK3 leads to stress granule dissolution (Wippich et al., 2013).

    Reaction to stress, creation of stress induced solutions, reversible(!) after stress period is finished.

    Amazing stuff!

    And this all done in “compartment-less” area through what I assume is unique solution? Not sure if this is replicated in prokaryotes? Or, unique to eukaryotes?

    I really cannot imagine any way to put such a system to work with any bottom-up strategy, even if designed. You really need a strict top-down engineering to get that kind of results. And even then, you need an unbelievable attention to details and connections between systems, and a perfect control of the symbolic system you are using.

    Great thing is, if it’s designed, molecular engineers, communications and network engineers, coders, etc., can reverse engineer it 🙂 Which is why Design Theory is a better heuristic going forward!

    Darwin is dead, neo-Darwinism is too as an overall solution guide, and only ancillary mutations it appears, mostly deleterious or weak form of survival mechanism.

    oh wow…

    In Drosophila cells, the stress of amino-acid starvation also inhibits protein transport through the secretory pathway (Zacharogianni et al., 2011) and leads to the remodeling of the ERES components into a novel membrane-less stress assembly, the Sec body (Zacharogianni et al., 2014). During the period of stress, Sec bodies store and protect most of the COPII components and Sec16 from degradation. They are round and display FRAP properties compatible with having liquid droplet properties. Importantly, they are pro-survival and rapidly disassemble upon stress relief. When stress is relieved, Sec bodies rapidly dissolve releasing their functional components that resume protein transport (Zacharogianni et al., 2014).

    Yep a system spontaneously generated with stress reaction mediation and then dispersion and back to normal.

    Sure… from abiogenesis to coordinated systems networking.

  293. off-topic,
    lncRNA treatments, although I assume, somewhere ubiquitin is in the pudding 😉

    Through this approach, the team identified 570 lncRNA molecules that were expressed differently in healthy and cancerous tissues. Further, they were able to uncover 633 previously unknown biomarkers that could act as predictive tools for 14 cancer types.

    The team then used this knowledge to try to treat mice that had been grafted with human lung cancer tissue. They injected each mouse with an agent that blocked the activity of the relevant lnRNA (locked nucleic acid antisense oligonucleotides) twice a week and examined the effects to the tumours. They found that within 15 days, their treatment had led to a tumour size reduction of almost 50%.

    Epigenetics, what once was thought to be Junk turns out to be crucial for optimized health and a key part of solving health issues.

    http://www.frontlinegenomics.c.....ng-cancer/

  294. DATCG,

    FYI – the prolific Italian composer GP got another ‘song’ in the ‘hit parade’ top 5 in the first 3 weeks since its release!

    This is interesting because the ‘pure science’ genre doesn’t seem very popular in this world. Who are those anonymous readers?

    🙂

    Popular Posts (Last 30 Days)

    News-watch: yet another incident of mass violence in FL, USA (1,980)

    My conclusion (so far) on the suggested infinite past,… (1,692)

    Stephen Hawking continues to talk widely celebrated nonsense (1,409)

    Becky’s Lesson, a Viginette (1,321)

    The Ubiquitin System: Functional Complexity and Semiosis… (1,282)

  295. DATCG @294:

    “…is there time to review all of these ubiquitin-related networks and functions?”

    Good question.

    Have you heard of the “Big Data Problem in Biology”?

  296. #297 Curious Bio-technophiles maybe? 😉

    Woot, wooot… Which reminds me, I was going to post something the other day on ER and celluar structures – organelles and your postings reminded me, I like pictures 😉 or videos.

    And I’m guessing some readers and lurkers do as well. There are many fine examples on youtube, but this is a good start and people can then see all the other choices should they like to learn more refined knowledge of each structure. This video shows Eukaryote and Prokaryote cells. Including a special guest performance by the irreducibly complex flagella 😉

    https://www.youtube.com/watch?v=URUJD5NEXC8

    Protein Synthesis…
    https://www.youtube.com/watch?v=kmrUzDYAmEI

    and maybe more later.

  297. #298-188… I think someone mentioned it before 😉

    Might be a good job to get into, high demand 😉 and good pay for sure!

  298. #297 Dionisio,

    and congrats to the systems ID review to the Mastro Gpuccio for another Top 5 composition 🙂

  299. Ubiquitin Chain formation, simplified overview shown by Fun with Tinker toys, multiple Chain positions…

    https://www.youtube.com/watch?v=miZYmuDKO2s

    Lecture in below video on Ubiquitin and Autophagy, can go to minute 4.20 mark for a quick look.

    Lecturer states about 90% of proteins are controlled by one of these two systems. Main message for good health? Don’t stop exercising!

    Resting for to long activates the proteolytic systems of Ubiquitin and autophagy! Muscles become weaker…. as Contractile Proteins are removed.

    https://www.youtube.com/watch?v=tliw477USx0

  300. Dionisio, DATCG:

    “FYI – the prolific Italian composer GP got another ‘song’ in the ‘hit parade’ top 5 in the first 3 weeks since its release!”

    Well, it would have been impossible without your constant support! 🙂

    “This is interesting because the ‘pure science’ genre doesn’t seem very popular in this world. Who are those anonymous readers?”

    Maybe we are in some kind of niche market…

  301. DATCG at #302:

    The thing I like most in the Tinker toys video is how she tries to be as precise as possible in wrapping the string around the wooden nucleosome, so that it is more or less 1.67 turns! (OK, more or less…) 🙂

  302. #304, hahahaha… you caught that did you? 😉

    here’s a image representation of our little friend Cdc48, ubiquitin, retrotranslocation complex, proteasome, ERAD-C & ERAD-L

    no tinker toys here 😉 She would have to bring a bigger box!

    http://www.cell.com/cms/attach.....46/gr3.jpg

    and interestingly, another representation from August 2014 of threading and Cdc48 regulation to the proteasome. Notice all the Question Marks at end of each explanation. Not sure if that’s an error or just a valid – we don’t know for sure…

    http://www.mdpi.com/cells/cell.....824-ag.png

    and the associated paper…
    Regulation of Endoplasmic Reticulum-Associated Protein Degradation (ERAD) by Ubiquitin

    http://www.mdpi.com/2073-4409/3/3/824/htm

  303. #303,

    “Maybe we are in some kind of niche market…”

    not by accident, only by Design 😉

    BTW, many readers might be searching for these papers we have all listed and come across this site as well. I’ve noticed several times UD gets listed fairly high, even on 1st page organic search sometimes on past references. Of course the searches are usually highly specific, long-tail SEO type searches.

  304. Special Issue “Protein Ubiquitination” 14 papers… published 2014.

    http://www.mdpi.com/journal/ce.....uitination

    One of interest is:

    Versatile Roles of K63-Linked Ubiquitin Chains in Trafficking
    Zoi Erpapazoglou 1,2, Olivier Walker 3 and Rosine Haguenauer-Tsapis 1,*

    http://www.mdpi.com/2073-4409/3/4/1027

    Abstract

    Modification by Lys63-linked ubiquitin (UbK63) chains is the second most abundant form of ubiquitylation.

    In addition to their role in DNA repair or kinase activation, UbK63 chains interfere with multiple steps of intracellular trafficking. UbK63 chains decorate many plasma membrane proteins, providing a signal that is often, but not always, required for their internalization. In yeast, plants, worms and mammals, this same modification appears to be critical for efficient sorting to multivesicular bodies and subsequent lysosomal degradation. UbK63 chains are also one of the modifications involved in various forms of autophagy (mitophagy, xenophagy, or aggrephagy). Here, in the context of trafficking, we report recent structural studies investigating UbK63 chains assembly by various E2/E3 pairs, disassembly by deubiquitylases, and specifically recognition as sorting signals by receptors carrying Ub-binding domains, often acting in tandem. In addition, we address emerging and unanticipated roles of UbK63 chains in various recycling pathways that function by activating nucleators required for actin polymerization, as well as in the transient recruitment of signaling molecules at the plasma or ER membrane. In this review, we describe recent advances that converge to elucidate the mechanisms underlying the wealth of trafficking functions of UbK63 chains.

    Keywords of the publication:

    – ubiquitin
    – ubiquitin chain
    – Sumo
    – 26S proteasome
    – protein stability
    – protein localization
    – E3 ligases
    – cellular regulation
    – signal transduction
    – development


    Example of Trafficking Steps involving UbK63 chains…

    http://www.mdpi.com/cells/cell.....7-g002.png

    “It is now clear that an expanding list of mammalian membrane proteins are modified by UbK63 chains at the plasma membrane (Table S1).”

    .

  305. Gpuccio, a bit off topic again, but thought it’s related as well in so many ways as we keep being amazed by all the intricate interactions and interdependency of so many systems working, coordinating together with Ubiquitin and multiple functions of different genes and proteins.

    When I came across this, made me think of you and TFs. And this shows TF’s constrain evolution.

    Transcription Factors, Pleiotropy and Constraints on Evolution

    hattip: Jeffery Tompkins PhD – ICR.org

    My overall thoughts are there are Constraints, conserved regions and hot spots meant for rapid evolution according to environmental queues, or stress, but very limited in novel forms. Like finch beaks. Sure, they get large or small based upon seasons, rain, droughts, but overall body plan, the bird is still a bird.

    And I simply cannot imagine the Ubiquitin System allowing for much more change.

    What level of constraint is the Ubiquitin System on evolution? And how would that begin to be measured? Cross-posted this here at UD post:

    Best guesses fail with plant evolution

  306. DATCG @308:

    “[…] there are Constraints, conserved regions and hot spots meant for rapid evolution according to environmental queues, or stress, but very limited in novel forms.”

    environmental queues?

    huh?

    did you mean “cues”?

  307. gpuccio @293:

    That the same regulation system, which is in itself as complex and multi-faceted as we have seen in this thread, can at the same time be the key regulator in such different processes (together with all the others we have discussed) is IMO mind-boggling.

    I really cannot imagine any way to put such a system to work with any bottom-up strategy, even if designed. You really need a strict top-down engineering to get that kind of results. And even then, you need an unbelievable attention to details and connections between systems, and a perfect control of the symbolic system you are using.

    The cross-talk between inner compartments in the cell is really a fascinating issue: too often we think of the cell as some rather homogeneous environment, or we just acknowledge the separation between nucleus and cytoplasm in eukaryotes. But the cytoplasm is anything but homogeneous. Organelles are separated by membranes, and membranes, as we have seen, are dynamic tools which are almost magically shaped and reshaped by complex molecular systems.

    And, even where mebranes are not present, a lot of functional sub-sections can be dynamically assessed, continuosly created and destroyed, shaping a functional landscape of cytoplasmic events which we still have to start to understand: second messengers, signaling pathways, and so on. No part of cytoplasm is the same as any other, each landscape is unique and functional.

    Agree. Beyond fascinating.

  308. In the dividing eukaryotic cell the spindle assembly checkpoint (SAC) ensures each daughter cell inherits an identical set of chromosomes.

    The SAC coordinates the correct attachment of sister chromatid kinetochores to the mitotic spindle with activation of the anaphase-promoting complex/cyclosome (APC/C), the E3 ubiquitin ligase that initiates chromosome separation.

    In response to unattached kinetochores, the SAC generates the mitotic checkpoint complex (MCC), a multimeric assembly that inhibits the APC/C, delaying chromosome segregation.

    Conformational variability of the complex allows for UbcH10 association, and we show from a structure of APC/CMCC in complex with UbcH10 how the Cdc20 subunit intrinsic to the MCC (Cdc20MCC) is ubiquitinated, a process that results in APC/C reactivation when the SAC is silenced.

    Molecular basis of APC/C regulation by the spindle assembly checkpoint
    Claudio Alfieri,#1 Leifu Chang,#1 Ziguo Zhang,1 Jing Yang,1 Sarah Maslen,1 Mark Skehel,1 and David Barford1
    Nature. 2016 Aug 25; 536(7617): 431–436.
    doi: 10.1038/nature19083

    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5019344/pdf/emss-69174.pdf

  309. Correct segregation of the mitotic chromosomes into daughter cells is a highly regulated process critical to safeguard genome stability. During M phase the spindle assembly checkpoint (SAC) ensures that all kinetochores are correctly attached before its inactivation allows progression into anaphase. Upon SAC inactivation, the anaphase promoting complex/cyclosome (APC/C) E3 ligase ubiquitinates and targets cyclin B and securin for proteasomal degradation. Here, we describe the identification of Ribonucleic Acid Export protein 1 (RAE1), a protein previously shown to be involved in SAC regulation and bipolar spindle formation, as a novel substrate of the deubiquitinating enzyme (DUB) Ubiquitin Specific Protease 11 (USP11). Lentiviral knock-down of USP11 or RAE1 in U2OS cells drastically reduces cell proliferation and increases multipolar spindle formation. We show that USP11 is associated with the mitotic spindle, does not regulate SAC inactivation, but controls ubiquitination of RAE1 at the mitotic spindle, hereby functionally modulating its interaction with Nuclear Mitotic Apparatus protein (NuMA).

    USP11 deubiquitinates RAE1 and plays a key role in bipolar spindle formation
    Anna Stockum, Ambrosius P. Snijders, Goedele N. Maertens
    PLoS One. 2018; 13(1): e0190513.
    doi: 10.1371/journal.pone.0190513

    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5749825/pdf/pone.0190513.pdf

  310. The anaphase promoting complex or cyclosome (APC/C) is a large multi-subunit E3 ubiquitin ligase that orchestrates cell cycle progression by mediating the degradation of important cell cycle regulators. During the two decades since its discovery, much has been learnt concerning its role in recognizing and ubiquitinating specific proteins in a cell-cycle-dependent manner, the mechanisms governing substrate specificity, the catalytic process of assembling polyubiquitin chains on its target proteins, and its regulation by phosphorylation and the spindle assembly checkpoint. The past few years have witnessed significant progress in understanding the quantitative mechanisms underlying these varied APC/C functions. This review integrates the overall functions and properties of the APC/C with mechanistic insights gained from recent cryo-electron microscopy (cryo-EM) studies of reconstituted human APC/C complexes.

    Visualizing the complex functions and mechanisms of the anaphase promoting complex/cyclosome (APC/C)
    Claudio Alfieri,† Suyang Zhang,† and David Barford
    Open Biol. 2017 Nov; 7(11): 170204.
    doi: 10.1098/rsob.170204
    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5717348/pdf/rsob-7-170204.pdf

  311. The anaphase-promoting complex (APC/C) is a multimeric RING E3 ubiquitin ligase that controls chromosome segregation and mitotic exit. Its regulation by coactivator subunits, phosphorylation, the mitotic checkpoint complex, and interphase inhibitor Emi1 ensures the correct order and timing of distinct cell cycle transitions. Here, we used cryo-electron microscopy to determine atomic structures of APC/C-coactivator complexes with either Emi1 or a UbcH10-ubiquitin conjugate. These structures define the architecture of all APC/C subunits, the position of the catalytic module, and explain how Emi1 mediates inhibition of the two E2s UbcH10 and Ube2S. Definition of Cdh1 interactions with the APC/C indicates how they are antagonized by Cdh1 phosphorylation. The structure of the APC/C with UbcH10-ubiquitin reveals insights into the initiating ubiquitination reaction. Our results provide a quantitative framework for the design of experiments to further investigate APC/C functions in vivo.

    Atomic structure of the APC/C and its mechanism of protein ubiquitination.
    Chang L#1, Zhang Z#1, Yang J1, McLaughlin SH1, Barford D1.
    Nature. 2015 Jun 25;522(7557):450-454.
    doi: 10.1038/nature14471. Epub 2015 Jun 15.
    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4608048/pdf/emss-65381.pdf

  312. DATCG at #308:

    Very interesting paper about TFs and their evolutionary rate!

    I think the really interesting data is in Fig. 2A, where it is shown that the evolutionary pattern of TFs, as referred to the whole molecule, is strongly related to the number of known TF -TF interactions.

    The analysis here is done with 1552 TFs, and it is a linear regression, but I suppose that a p value of 5e-36 can never be questioned by anybody! 🙂

    That is the true, strong point: TFs which have a high number of interactions with other TFs are highly contrained (IOWs, their whole sequence is strongly conserved).

    A few comments:

    a) Just as a clarification for possible readers, the parameter they are using to measure sequence conservation is the dN/dS ratio, which is nothing else than the Ka/Ks ratio (against, nomenclature!) that I have often used in my discussions, IOWs the ration between non synonimous mutations (per non synonimous site) and synonimous mutations (per synonimous site). The lower this value, the higher the sequence conservation. The reference to synonimous mutations makes the measure relatively independent from evolutionary times (at least for evolutionary times which are not too long).

    b) I am not too sure that the number of TF – TF interactions can be interpreted only as a measure of pleiotropy, IOWs of multiple function. As the working of TFs for one single function is often combinatorial, with many TFs joining in very big protein complexes to achieve the fine tuning of the function itself, I would say that the number of known TF – TF interactions is also a measure of the complexity of the individual functions regulated by those TFs, and not only of the number of functions to which each TF contributes.

    c) The important point is: TFs are highly functional molecules, and their whole molecule contributes to their function, not only the DBD, or even the known protein interaction domains. As we have seen, the sequences with “conditional folding” are probably the most important in the final regulatory functions.

  313. DATCG at #307:

    Great review of the known roles of K63 ubiquitin chains, “the second most abundant form of ubiquitylation”!

    This kind of ubiquitination is specially interesting because it is usually proteasome independent (K48 and K11 being the proteasome linked ubiquitinations).

    And look at the number of intriguing and complex functions implemented by K63 ubiquitination: modifications of plasma membrane proteins and cargoes, internalization of receptors, sorting to multivesicular bodies, other forms of cell trafficking, signaling pathways, selective autophagy, mitophagy, xenophagy.

    These are just the main titles of the various sections in the paper, where each of these complex subjects is well summarized according to our present understanding.

    And all these functions must be added to the multitude of specific functions that the ubiquitin system implements by K48 ubiquitination and proteosamal degradation, as we have discussed in detail previously! 🙂

  314. Ubiquitin-like proteins (Ubl’s) are conjugated to target proteins or lipids to regulate their activity, stability, subcellular localization, or macromolecular interactions. Similar to ubiquitin, conjugation is achieved through a cascade of activities that are catalyzed by E1 activating enzymes, E2 conjugating enzymes, and E3 ligases. In this review, we will summarize structural and mechanistic details of enzymes and protein cofactors that participate in Ubl conjugation cascades. Precisely, we will focus on conjugation machinery in the SUMO, NEDD8, ATG8, ATG12, URM1, UFM1, FAT10, and ISG15 pathways while referring to the ubiquitin pathway to highlight common or contrasting themes. We will also review various strategies used to trap intermediates during Ubl activation and conjugation.

    Ubiquitin-like Protein Conjugation: Structures, Chemistry, and Mechanism
    Laurent Cappadocia† and Christopher D. Lima
    Chem Rev. 2018 Feb 14; 118(3): 889–918.
    doi: 10.1021/acs.chemrev.6b00737
    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5815371/pdf/cr6b00737.pdf

  315. […] protein turnover by the ubiquitin-proteasome system provides a vital mechanism for the regulation of centrosome protein levels.

    PC/CFZR-1 Controls SAS-5 Levels To Regulate Centrosome Duplication in Caenorhabditis elegans
    Jeffrey C. Medley, Lauren E. DeMeyer, Megan M. Kabara, and Mi Hye Song
    G3 (Bethesda). 2017 Dec; 7(12): 3937–3946.
    doi: 10.1534/g3.117.300260
    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5714490/pdf/3937.pdf

  316. The homeostasis of MCPH1 in association with the ubiquitin-proteasome system ensures mitotic entry independent of cell cycle checkpoint.

    The E3 ubiquitin ligase APC/CCdh1 degrades MCPH1 after MCPH1-?TrCP2-Cdc25A-mediated mitotic entry to ensure neurogenesis.
    Liu X1, Zong W1, Li T1,2, Wang Y3, Xu X4,5, Zhou ZW6, Wang ZQ
    EMBO J. 2017 Dec 15;36(24):3666-3681.
    doi: 10.15252/embj.201694443.

  317. Dionisio at #317:

    Yes, ubiquitin like proteins certainly add a lot to the complexity of the system. And the paper you linked is a very good and very recent review of what is known about them.

    SUMO is one of the most important in the group.

    SUMO1 is a 101 AAs long protein in humans. Strangley, it does not exhibit a great sequence homology with ubiqutin (13 identities, 33 positives, 29.3 bits, a weakly significant e-value of 9e-07).

    However, its sequence is highly conserved in eukaryotes. Not so much as ubiquitin, but highly conserved just the same.

    The human protein shows 47 identities and 66 positives with fungi (102 bits, e value 2e-27). But those values of homology rapidly increase in metazoa.

    The protein is one of those which undergo important engineering in vertebrates, passing from 138 to 178 bits of homology, a 0.396 baa jump.

    The protein in cartilaginous fish shows 84% identities and 92% positives with the human form. This is very strong conservation.

    So, the obvious point is: SUMO is an ubiquitin-related protein, but it is different: different in sequence, different in functions and functional networks. It has its specific E1-E2-E3 systems.

    And this “different” protein is already present in single celled eukaryotes, and is well conserved throughout the whole eukaryotic history. Much more conserved than it is similar to ubiquitin itself.

    So, what does that mean? It means that this is certainly a variant of the ubiquitin concept, but it appears from the beginning and is different form the beginning, and it maintains its difference, because its difference is functional, is specific, and is therefore conserved.

  318. gpuccio @320:

    “[…] this is certainly a variant of the ubiquitin concept, but it appears from the beginning and is different form the beginning, and it maintains its difference, because its difference is functional, is specific, and is therefore conserved.”

    Interesting. Thanks.

  319. Human gut Bacteroides species produce different types of toxins that antagonize closely related members of the gut microbiota. Some are toxic effectors delivered by type VI secretion systems, and others are non-contact-dependent secreted antimicrobial proteins. Many strains of Bacteroides fragilis secrete antimicrobial molecules, but only one of these toxins has been described to date (Bacteroidales secreted antimicrobial protein 1 [BSAP-1]). In this study, we describe a novel secreted protein produced by B. fragilis strain 638R that mediated intraspecies antagonism. Using transposon mutagenesis and deletion mutation, we identified a gene encoding a eukaryotic-like ubiquitin protein (BfUbb) necessary for toxin activity against a subset of B. fragilis strains. The addition of ubb into a heterologous background strain conferred toxic activity on that strain. We found this gene to be one of the most highly expressed in the B. fragilis genome. The mature protein is 84% similar to human ubiquitin but has an N-terminal signal peptidase I (SpI) signal sequence and is secreted extracellularly. We found that the mature 76-amino-acid synthetic protein has very potent activity, confirming that BfUbb mediates the activity. Analyses of human gut metagenomic data sets revealed that ubb is present in 12% of the metagenomes that have evidence of B. fragilis. As 638R produces both BSAP-1 and BfUbb, we performed a comprehensive analysis of the toxin activity of BSAP-1 and BfUbb against a set of 40 B. fragilis strains, revealing that 75% of B. fragilis strains are targeted by one or the other of these two secreted proteins of strain 638R.

    Gut Symbiont Bacteroides fragilis Secretes a Eukaryotic-Like Ubiquitin Protein That Mediates Intraspecies Antagonism
    Maria Chatzidaki-Livanis, Michael J. Coyne, Kevin G. Roelofs,* Rahul R. Gentyala,* Jarreth M. Caldwell,* and Laurie E. Comstock
    mBio. 2017 Nov-Dec; 8(6): e01902-17.
    doi: 10.1128/mBio.01902-17
    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5705921/pdf/mBio.01902-17.pdf

  320. Posttranslational modification with ubiquitin chains controls cell fate in all eukaryotes. Depending on the connectivity between subunits, different ubiquitin chain types trigger distinct outputs, as seen with K48- and K63-linked conjugates that drive protein degradation or complex assembly, respectively. Recent biochemical analyses also suggested roles for mixed or branched ubiquitin chains, yet without a method to monitor endogenous conjugates, the physiological significance of heterotypic polymers remained poorly understood. Here, we engineered a bispecific antibody to detect K11/K48-linked chains and identified mitotic regulators, misfolded nascent polypeptides, and pathological Huntingtin variants as their endogenous substrates. We show that K11/K48-linked chains are synthesized and processed by essential ubiquitin ligases and effectors that are mutated across neurodegenerative diseases; accordingly, these conjugates promote rapid proteasomal clearance of aggregation-prone proteins. By revealing key roles of K11/K48-linked chains in cell-cycle and quality control, we establish heterotypic ubiquitin conjugates as important carriers of biological information.

    Assembly and Function of Heterotypic Ubiquitin Chains in Cell-Cycle and Protein Quality Control.
    Yau RG1, Doerner K2, Castellanos ER3, Haakonsen DL1, Werner A2, Wang N4, Yang XW4, Martinez-Martin N5, Matsumoto ML6, Dixit VM7, Rape M
    Cell. 2017 Nov 2;171(4):918-933.e20.
    doi: 10.1016/j.cell.2017.09.040

  321. Ubiquitylation is a tightly regulated process that is essential for appropriate cell survival and function, and the ubiquitin pathway has shown promise as a therapeutic target for several forms of cancer. In this issue of the JCI, Kedves and colleagues report the identification of a subset of gynecological cancers with repressed expression of the polyubiquitin gene UBB, which renders these cancer cells sensitive to further decreases in ubiquitin production by inhibition of the polyubiquitin gene UBC. Moreover, inducible depletion of UBC in mice harboring tumors with low UBB levels dramatically decreased tumor burden and prolonged survival. Together, the results of this study indicate that there is a synthetic lethal relationship between UBB and UBC that has potential to be exploited as a therapeutic strategy to fight these devastating cancers.

    Ubiquitin levels: the next target against gynecological cancers?
    Haakonsen DL, Rape M
    J Clin Invest. 2017 Dec 1;127(12):4228-4230.
    doi: 10.1172/JCI98262
    https://www.jci.org/articles/view/98262/pdf

  322. Protein ubiquitylation is an important post-translational modification, regulating aspects of virtually every biochemical pathway in eukaryotic cells. Hundreds of enzymes participate in the conjugation and deconjugation of ubiquitin, as well as the recognition, signaling functions, and degradation of ubiquitylated proteins. Regulation of ubiquitylation is most commonly at the level of recognition of substrates by E3 ubiquitin ligases. Characterization of the network of E3-substrate relationships is a major goal and challenge in the field, as this expected to yield fundamental biological insights and opportunities for drug development. There has been remarkable success in identifying substrates for some E3 ligases, in many instances using the standard protein-protein interaction techniques (e.g., two-hybrid screens and co-immunoprecipitations paired with mass spectrometry). However, some E3s have remained refractory to characterization, while others have simply not yet been studied due to the sheer number and diversity of E3s. This review will discuss the range of tools and techniques that can be used for substrate profiling of E3 ligases.

    O’Connor, Hazel F., and Jon M. Huibregtse. “Enzyme–substrate Relationships in the Ubiquitin System: Approaches for Identifying Substrates of Ubiquitin Ligases.” Cellular and Molecular Life Sciences 74, no. 18 (September 2017): 3363–75. https://doi.org/10.1007/s00018-017-2529-6.

  323. Covalent, reversible, post-translational modification of cellular proteins with the small modifier, ubiquitin (Ub), regulates virtually every known cellular process in eukaryotes. The process is carried out by a trio of enzymes: a Ub-activating (E1) enzyme, a Ub-conjugating (E2) enzyme, and a Ub ligase (E3) enzyme. RING-in-Between-RING (RBR) E3s constitute one of three classes of E3 ligases and are defined by a RING-HECT-hybrid mechanism that utilizes a E2-binding RING domain and a second domain (called RING2) that contains an active site Cys required for the formation of an obligatory E3~Ub intermediate. Albeit a small class, RBR E3s in humans regulate diverse cellular process. This review focuses on non-Parkin members such as HOIP/HOIL-1L (the only E3s known to generate linear Ub chains), HHARI and TRIAD1, both of which have been recently demonstrated to work together with Cullin RING E3 ligases. We provide a brief historical background and highlight, summarize, and discuss recent developments in the young field of RBR E3s. Insights reviewed here include new understandings of the RBR Ub-transfer mechanism, specifically the role of RING1 and various Ub-binding sites, brief structural comparisons among members, and different modes of auto-inhibition and activation.

    Dove, Katja K., and Rachel E. Klevit. “RING-Between-RING E3 Ligases: Emerging Themes amid the Variations.” Journal of Molecular Biology 429, no. 22 (November 2017): 3363–75. https://doi.org/10.1016/j.jmb.2017.08.008.

  324. Since its discovery as a post-translational signal for protein degradation, our understanding of ubiquitin (Ub) has vastly evolved. Today, we recognize that the role of Ub signaling is expansive and encompasses diverse processes including cell division, the DNA damage response, cellular immune signaling, and even organismal development. With such a wide range of functions comes a wide range of regulatory mechanisms that control the activity of the ubiquitylation machinery. Ub attachment to substrates occurs through the sequential action of three classes of enzymes, E1s, E2s, and E3s. In humans, there are 2 E1s, ? 35 E2s, and hundreds of E3s that work to attach Ub to thousands of cellular substrates. Regulation of ubiquitylation can occur at each stage of the stepwise Ub transfer process, and substrates can also impact their own modification. Recent studies have revealed elegant mechanisms that have evolved to control the activity of the enzymes involved. In this minireview, we highlight recent discoveries that define some of the various mechanisms by which the activities of E3-Ub ligases are regulated.

    Vittal, Vinayak, Mikaela D. Stewart, Peter S. Brzovic, and Rachel E. Klevit. “Regulating the Regulators: Recent Revelations in the Control of E3 Ubiquitin Ligases.” Journal of Biological Chemistry 290, no. 35 (August 28, 2015): 21244–51. https://doi.org/10.1074/jbc.R115.675165.

    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4571856/pdf/zbc21244.pdf

  325. The eukaryotic 26S proteasome is a large multisubunit complex that degrades the majority of proteins in the cell under normal conditions. The 26S proteasome can be divided into two subcomplexes: the 19S regulatory particle and the 20S core particle. Most substrates are first covalently modified by ubiquitin, which then directs them to the proteasome. The function of the regulatory particle is to recognize, unfold, deubiquitylate, and translocate substrates into the core particle, which contains the proteolytic sites of the proteasome. Given the abundance and subunit complexity of the proteasome, the assembly of this ~2.5MDa complex must be carefully orchestrated to ensure its correct formation. In recent years, significant progress has been made in the understanding of proteasome assembly, structure, and function. Technical advances in cryo-electron microscopy have resulted in a series of atomic cryo-electron microscopy structures of both human and yeast 26S proteasomes. These structures have illuminated new intricacies and dynamics of the proteasome. In this review, we focus on the mechanisms of proteasome assembly, particularly in light of recent structural information.

    Budenholzer, Lauren, Chin Leng Cheng, Yanjie Li, and Mark Hochstrasser. “Proteasome Structure and Assembly.” Journal of Molecular Biology 429, no. 22 (November 2017): 3500–3524. https://doi.org/10.1016/j.jmb.2017.05.027.

  326. Cellular protein homeostasis is maintained by two major degradation pathways, namely the ubiquitin-proteasome system (UPS) and autophagy. Until recently, the UPS and autophagy were considered to be largely independent systems targeting proteins for degradation in the proteasome and lysosome, respectively. However, the identification of crucial roles of molecular players such as ubiquitin and p62 in both of these pathways as well as the observation that blocking the UPS affects autophagy flux and vice versa has generated interest in studying crosstalk between these pathways. Here, we critically review the current understanding of how the UPS and autophagy execute coordinated protein degradation at the molecular level, and shed light on our recent findings indicating an important role of an autophagy-associated transmembrane protein EI24 as a bridging molecule between the UPS and autophagy that functions by regulating the degradation of several E3 ligases with Really Interesting New Gene (RING)-domains.

    Nam, Taewook, Jong Hyun Han, Sushil Devkota, and Han-Woong Lee. “Emerging Paradigm of Crosstalk between Autophagy and the Ubiquitin-Proteasome System.” Molecules and Cells 40, no. 12 (December 31, 2017): 897–905. https://doi.org/10.14348/molcells.2017.0226.
    http://www.molcells.org/journa......2017.0226

  327. is this off-topic? not sure…

    We present an atomic model of a substrate-bound inner mitochondrial membrane AAA+ quality control protease in yeast, YME1. Our ~3.4-angstrom cryo-electron microscopy structure reveals how the adenosine triphosphatases (ATPases) form a closed spiral staircase encircling an unfolded substrate, directing it toward the flat, symmetric protease ring. Three coexisting nucleotide states allosterically induce distinct positioning of tyrosines in the central channel, resulting in substrate engagement and translocation to the negatively charged proteolytic chamber. This tight coordination by a network of conserved residues defines a sequential, around-the-ring adenosine triphosphate hydrolysis cycle that results in stepwise substrate translocation. A hingelike linker accommodates the large-scale nucleotide-driven motions of the ATPase spiral relative to the planar proteolytic base. The translocation mechanism is likely conserved for other AAA+ ATPases.

    Puchades, Cristina, Anthony J. Rampello, Mia Shin, Christopher J. Giuliano, R. Luke Wiseman, Steven E. Glynn, and Gabriel C. Lander. “Structure of the Mitochondrial Inner Membrane AAA+ Protease YME1 Gives Insight into Substrate Processing.” Science 358, no. 6363 (November 3, 2017): eaao0464. https://doi.org/10.1126/science.aao0464.

    The hexameric AAA ATPase Vps4 drives membrane fission by remodeling and disassembling ESCRT-III filaments. Building upon our earlier 4.3 Å resolution cryo-EM structure (Monroe et al., 2017), we now report a 3.2 Å structure of Vps4 bound to an ESCRT-III peptide substrate. The new structure reveals that the peptide approximates a ?-strand conformation whose helical symmetry matches that of the five Vps4 subunits it contacts directly. Adjacent Vps4 subunits make equivalent interactions with successive substrate dipeptides through two distinct classes of side chain binding pockets formed primarily by Vps4 pore loop 1. These pockets accommodate a wide range of residues, while main chain hydrogen bonds may help dictate substrate-binding orientation. The structure supports a ‘conveyor belt’ model of translocation in which ATP binding allows a Vps4 subunit to join the growing end of the helix and engage the substrate, while hydrolysis and release promotes helix disassembly and substrate release at the lagging end.

    Han, Han, Nicole Monroe, Wesley I Sundquist, Peter S Shen, and Christopher P Hill. “The AAA ATPase Vps4 Binds ESCRT-III Substrates through a Repeating Array of Dipeptide-Binding Pockets.” ELife 6 (November 22, 2017). https://doi.org/10.7554/eLife.31324.
    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5716660/pdf/elife-31324.pdf

  328. @327:

    Since its discovery as a post-translational signal for protein degradation, our understanding of ubiquitin (Ub) has vastly evolved.

    Perhaps this is a case where the meaning of the term “evolution” is unanimously accepted? 🙂

    Evolution of our understanding of ubiquitin?

  329. maybe the functional complexity of cellular* membranes could be a future topic for an OP?

    (*) including organelle membrane too

    https://www.ncbi.nlm.nih.gov/Structure/pdb/6BMF

    https://www.ncbi.nlm.nih.gov/Structure/pdb/6AP1

  330. Dionisio,

    “… queue” is a word I abuse quite frequently. Think it is a leftover from many visits to London and parts of England and Scotland where I had to stand in a queue 🙂

    For some reason my brain mistranslates cue to queue. Not the first time. Maybe I’ll just use a different word entirely.

    Environmental Factors 😉 EFs. or Input.

  331. Layman, Awo A. K., and Paula M. Oliver. “Ubiquitin Ligases and Deubiquitinating Enzymes in CD4 + T Cell Effector Fate Choice and Function.” The Journal of Immunology 196, no. 10 (May 15, 2016): 3975–82. https://doi.org/10.4049/jimmunol.1502660.

    Skieterska, Kamila, Pieter Rondou, and Kathleen Van Craenenbroeck. “Regulation of G Protein-Coupled Receptors by Ubiquitination.” International Journal of Molecular Sciences 18, no. 12 (April 27, 2017): 923. https://doi.org/10.3390/ijms18050923.

    Ohtake, Fumiaki, Hikaru Tsuchiya, Yasushi Saeki, and Keiji Tanaka. “K63 Ubiquitylation Triggers Proteasomal Degradation by Seeding Branched Ubiquitin Chains.” Proceedings of the National Academy of Sciences 115, no. 7 (February 13, 2018): E1401–8. https://doi.org/10.1073/pnas.1716673115.

    Grice, Guinevere L., and James A. Nathan. “The Recognition of Ubiquitinated Proteins by the Proteasome.” Cellular and Molecular Life Sciences 73, no. 18 (September 2016): 3497–3506. https://doi.org/10.1007/s00018-016-2255-5.

  332. Poot, Stefanie A.H. de, Geng Tian, and Daniel Finley. “Meddling with Fate: The Proteasomal Deubiquitinating Enzymes.” Journal of Molecular Biology 429, no. 22 (November 2017): 3525–45. https://doi.org/10.1016/j.jmb.2017.09.015.

    Boutouja, Fahd, Rebecca Brinkmeier, Thomas Mastalski, Fouzi El Magraoui, and Harald Platta. “Regulation of the Tumor-Suppressor BECLIN 1 by Distinct Ubiquitination Cascades.” International Journal of Molecular Sciences 18, no. 12 (November 27, 2017): 2541. https://doi.org/10.3390/ijms18122541.

  333. Dionisio @ 327/331

    Ha! 🙂

    As knowledge increases of Functional Sequence Complex – inter-Dependent Organized Systems(FSC-iDOS), I think we find “evolve” is an over-hyped term in “evolutionary” biology.

    Gpuccio always points out Variation? Random Variation.

    There is deleterious mutations and then significantly controlled, programmatic, conditional logic of Allowed Variation. Based off environmental qu…. uh stimuli 😉

    Or internal factors with prescribed actionable network systems response.

    All the papers you posted and Gpuccio commented on shows overwhelming evidence of Design and planning. At so many different levels of expertise, not only coding, but of engineering which as you’ve said, we’ve not seen nothing yet!

    Like “junk” DNA, functions abound in places Darwinist once said had no function…

    Appendix Might Save Your Life – 2012 SciAm

    ou may have heard the appendix is vestigial, a relict of our past like the hind leg bones of a whale. Parker heard that too, he just disagrees. Parker thinks the appendix serves as a nature reserve for beneficial bacteria in our guts. When we get a severe gut infection such as cholera (which happened often during much of our history and happens often in many regions even today), the beneficial bacteria in our gut are depleted. The appendix allows them to be restored. In essence, Parker sees the appendix as a sanctuary for our tiny mutualist friends, a place where there is always room at the inn4. If he is right, the appendix nurtures beneficial bacteria even as our conscious brains and cultures tell us to kill, kill, kill them with wipes and pills.

    “Evolve” “Junk” “Vestigial”

    You keep using that word, I do not think it means what you think it means

    .

  334. Ran a quick search on technology and Ubiquitin to determine how far science and scientist have “evolved” in use of technology to decode the Ubiquitin Code.

    Tracing down linear ubiquitination

    New technology enables detailed analysis of target proteins
    Date: March 20, 2017
    Source: Goethe University Frankfurt
    Summary: Researchers have developed a novel technology to decipher the secret ubiquitin code.

    Scientists often refer to it as the secret ubiquitin code, which still needs to be fully deciphered.

    Recently, scientists discovered that ubiquitin molecules are not only assembled in a non-linear manner, but also build linear chains, in which the head of one ubiquitin is linked to the tail of another ubiquitin molecule. So far, only two highly specific enzymes are known capable of synthesizing and degrading such linear ubiquitin chains, and both are being extensively studied at the Institute of Biochemistry II at the Goethe University Frankfurt. However, target proteins of linear ubiquitination, as well as their specific cellular functions, have largely remained elusive. The novel technology developed by the team around Koraljka Husnjak from the Goethe University Frankfurt now enables the systematic analysis of linear ubiquitination targets.

    “The slow progress in this research area was mainly due to the lack of suitable methods for proteomic analysis of proteins modified with linear ubiquitin chains,” explains Koraljka Husnjak whose native country is Croatia. Her team solved the problem by internally modifying the ubiquitin molecule in such a way that it maintains its cellular functions whilst at the same time enabling the enrichment and further analysis of linear ubiquitin targets by mass spectrometry.

    Only a year ago this technique began. Amazing. So many, many more papers to come in future using this technique of identification.

    With this technology at hand, it is now possible to identify target proteins modified by linear ubiquitin, and to detect the exact position within the protein where the linear chain is attached. Scientists praise this highly sensitive approach as an important breakthrough that will strongly improve our understanding of the functions of linear ubiquitination and its role in diseases.

    Dr. Husnjak already provided the proof of this concept and identified several novel proteins modified by linear ubiquitin chains. Amongst them are essential components of one of the major pro-inflammatory pathways within cells. “Linear ubiquitin chains relay signals that play an important role in the regulation of immune responses, in pathogen defence and immunological disorders. Until now we know very little about how small slips in this system contribute to severe diseases, and how we can manipulate it for therapeutic purposes” comments Husnjak the potential of the new technology.

    Errors in the ubiquitin system have been linked to numerous diseases including cancer and neurodegenerative disorders such as Parkinson’s disease, but also to the development and progression of infections and inflammatory diseases.

    Great work by Dr. Husnjak and his team(s) at Goethe University Frankfurt.

    Katarzyna Kliza, Christoph Taumer, Irene Pinzuti, Mirita Franz-Wachtel, Simone Kunzelmann, Benjamin Stieglitz, Boris Macek & Koraljka Husnjak
    Nature Methods volume 14, pages 504–512 (2017)
    doi:10.1038/nmeth.4228

    Scientific paper behind pay wall…

    Nature – Internally tagged ubiquitin: a tool to identify linear polyubiquitin-modified proteins by mass spectrometry

    Decipher:
    1 – decode
    1a – decipher a secret message
    3a – to make out the meaning of despite indistinctness or obscurity
    3b – to interpret the meaning of

    Code:
    3a – a system of signals or symbols for communication
    3b – a system of symbols (such as letters or numbers) used to represent assigned and often secret meanings
    4 – genetic code
    5 – instructions for a computer (as within a piece of software

  335. Dionisio at #131:

    “Evolution of our understanding of ubiquitin?”

    Well, our understanding of ubiquitin has certainly “evolved” from the beginning of this thread! 🙂

    I suppose it was not a completely unguided process, however. It took some specific work and attempts at understanding by a small group of rather insubordinate people (including me), a lot of not really “natural” selection of papers from the literature, and some effort to express relevant thoughts in the 300+ comments by 4+ commenters in the discussion.

    I would say that environmental pressure (comments from the other side) had no relevant role in shaping that evolution (indeed, no role at all!).

    RV certainly was present, mostly in the form of typos, even if our small structure has a very efficient proof checking system (you know what I mean! 😉 )

    That said, the results are not bad. And I can see a lot of convergent evolution all around! 🙂

  336. Dionisio:

    “Maybe the functional complexity of cellular* membranes could be a future topic for an OP?
    (*) including organelle membrane too”

    It’s certainly a possibility.

  337. DATCG at #336:

    “Or internal factors with prescribed actionable network systems response.”

    The subject of intelligent and functional algorithmic responses to environment is fascinating. We have certainly many examples of that.

    One is well known, and I have written about it in a previous OP:

    https://uncommondescent.com/intelligent-design/antibody-affinity-maturation-as-an-engineering-process-and-other-things/

    Antibody affinity maturation is indeed a wonderful example of algorithmic process which creates important functional information based on the acquisition of information from the environment (the contact with the antigen) and from a highly complex computational process (the maturation process), essentially of the bottom-up type.

    It is interesting that many times it gas been pointed to as an example of “darwinian evolution”, which is good evidence of how confused are sometimes our kind interlocutors.

    Of course, such a refined computational process is outstanding evidence of design: designed objects can indeed comput new information about the pre-defined function and using new information inputs and their pre-programmed computing information: that’s what computers, or neural networks, do all the time.

    I think that another system designed to provide that kind of functionality is probably the plasmid system in prokaryotes.

    However, computational systems, even computers, always have the same fundamental limit in themselves: they can only compute what they have been directly or indirectly programmed to compute, and nothing else.

    That’s why the generation of really new complex functional information always requires a conscious designer.

  338. DATCG at #336:

    “All the papers you posted and Gpuccio commented on shows overwhelming evidence of Design and planning. At so many different levels of expertise, not only coding, but of engineering which as you’ve said, we’ve not seen nothing yet!”

    Of course. We have been witnessing here, in this interesting thread, a perfect example of design of the highest kind, a kind that vastly outperforms anything we can yet try to conceive.

    Just to sum up, we have seen tons of examples of:

    a) Huge functional complexity, and of the highest type, the regulatory type.

    b) The ubiquitous presence of refined semiosis, everywhere.

    c) Hundreds, maybe thousands, of individual systems exhibiting, each of them, irreducible complexity.

    But our kind interlocutors seem not to be interested in all that. OK, but they miss a lot of fun! 🙂

  339. gpuccio @340:

    Where are DNA_Jock, sparc, and other politely dissenting interlocutors that were so active in your interesting 2015 OP and discussion thread that you pointed at?

    It would definitely add some “spice” to the discussion to have a couple of serious opponents actively participating, but where have they all gone? 🙂

    Has anybody heard of professor Arthur Hunt lately?
    I’m willing to get off this thread if that’s the condition for professor Larry Moran to come back. At least that would reassure him that nobody will ask dishonest questions with “tricky” words like “exactly” subliminally embedded in the questions. GP as the owner and moderator of this thread will ensure that all “tricky” words are written in bold font so nobody misses their presence in the text. 🙂

  340. gpuccio @341:

    Just to sum up, we have seen tons of examples of:
    a) Huge functional complexity, and of the highest type, the regulatory type.
    b) The ubiquitous presence of refined semiosis, everywhere.
    c) Hundreds, maybe thousands, of individual systems exhibiting, each of them, irreducible complexity.

    Excellent summary! Thanks.

  341. gpuccio @338:

    Very refreshing sense of humor pointing to what’s going on here. Thanks.

    I have learned (and still learning) much from this OP + discussion thread. Much more than I expected at the start, even though my expectations were high. Thanks.

  342. gpuccio @339:

    That’s encouraging. I look forward to reading it someday. Thanks.

    BTW, are your OPs part of your intensive preparation for a potentially future OP on “procedures”?

    As UB stated before, if you ever decide to write a book with all your OP + follow up comments, you’ll make many happy campers around here and out there! 🙂

  343. gpuccio @338:

    To illustrate the refreshingly funny assessment of this discussion and its effect on our knowledge, let’s add that around 120 papers have been referenced in this thread so far.

  344. Dionisio:

    “BTW, are your OPs part of your intensive preparation for a potentially future OP on “procedures”?”

    I suppose they are. I must say that the “preparation” is much more “intensive” than I could imagine! 🙂

  345. Dionisio:

    “To illustrate the refreshingly funny assessment of this discussion and its effect on our knowledge, let’s add that around 120 papers have been referenced in this thread so far.”

    Yes. Not bad.

    And, I would say, almost all rather pertinent. And many of them extremely recent.

  346. Dionisio:

    “It would definitely add some “spice” to the discussion to have a couple of serious opponents actively participating, but where have they all gone?”

    I would like to know. Some of them were pretty good!

    “Has anybody heard of professor Arthur Hunt lately?”

    Apparently not.

    “I’m willing to get off this thread if that’s the condition for professor Larry Moran to come back.”

    I don’t believe that it would work! 🙂

    I don’t think that I have ever discussed directly with Larry Moran, even if I have commented about some of his statements a couple of times.

    “GP as the owner and moderator of this thread will ensure that all “tricky” words are written in bold font so nobody misses their presence in the text.”

    Well, I have never “moderated” anything in my life, I would not like to begin with you! 🙂

    I confide in your self-discipline to ensure that all bolds are assigned in a politically correct way.

  347. DATCG at #337:

    Decipher:
    1 – decode
    1a – decipher a secret message
    3a – to make out the meaning of despite indistinctness or obscurity
    3b – to interpret the meaning of

    Code:
    3a – a system of signals or symbols for communication
    3b – a system of symbols (such as letters or numbers) used to represent assigned and often secret meanings
    4 – genetic code
    5 – instructions for a computer (as within a piece of software

    Wonderful clarification of terms which are often badly used.

    It’s refreshing to see how the subjective experience of meaning is central even in simple definitions. And how the symbolic nature of codes is crystal clear in language.

    Codes and design are connected just from the beginning by their definitions themselves: both words have no sense if we don’t refer in some way to the subjective experience of understanding meanings!

  348. Chapard, C., P. Meraldi, T. Gleich, D. Bachmann, D. Hohl, and M. Huber. “TRAIP Is a Regulator of the Spindle Assembly Checkpoint.” Journal of Cell Science 127, no. 24 (December 15, 2014): 5149–56. https://doi.org/10.1242/jcs.152579.

    Hoffmann, Saskia, Stine Smedegaard, Kyosuke Nakamura, Gulnahar B. Mortuza, Markus Räschle, Alain Ibañez de Opakua, Yasuyoshi Oka, et al. “TRAIP Is a PCNA-Binding Ubiquitin Ligase That Protects Genome Stability after Replication Stress.” The Journal of Cell Biology 212, no. 1 (January 4, 2016): 63–75. https://doi.org/10.1083/jcb.201506071.

    Ma, Xingjie, Junjie Zhao, Fan Yang, Haitao Liu, and Weibo Qi. “Ubiquitin Conjugating Enzyme E2 L3 Promoted Tumor Growth of NSCLC through Accelerating P27kip1 Ubiquitination and Degradation.” Oncotarget 8, no. 48 (October 13, 2017). https://doi.org/10.18632/oncotarget.20449.

    Min, M., T. E. T. Mevissen, M. De Luca, D. Komander, and C. Lindon. “Efficient APC/C Substrate Degradation in Cells Undergoing Mitotic Exit Depends on K11 Ubiquitin Linkages.” Molecular Biology of the Cell 26, no. 24 (December 1, 2015): 4325–32. https://doi.org/10.1091/mbc.E15-02-0102.

    Nath, Somsubhra, Taraswi Banerjee, Debrup Sen, Tania Das, and Susanta Roychoudhury. “Spindle Assembly Checkpoint Protein Cdc20 Transcriptionally Activates Expression of Ubiquitin Carrier Protein UbcH10.” Journal of Biological Chemistry 286, no. 18 (May 6, 2011): 15666–77. https://doi.org/10.1074/jbc.M110.160671.

    Iimura, Akira, Fuhito Yamazaki, Toshiyasu Suzuki, Tatsuya Endo, Eisuke Nishida, and Morioh Kusakabe. “The E3 Ubiquitin Ligase Hace1 Is Required for Early Embryonic Development in Xenopus Laevis.” BMC Developmental Biology 16, no. 1 (December 2016). https://doi.org/10.1186/s12861-016-0132-y.

    Kai, Masatake, Naoto Ueno, and Noriyuki Kinoshita. “Phosphorylation-Dependent Ubiquitination of Paraxial Protocadherin (PAPC) Controls Gastrulation Cell Movements.” Edited by Jung Weon Lee. PLOS ONE 10, no. 1 (January 12, 2015): e0115111. https://doi.org/10.1371/journal.pone.0115111.

  349. #340 Gpuccio,

    After going back to 2015 post, I must have landed in a Wagner Hyper-astronomical subspace. Ha!

    grrr… lost my original comment Wagner’s hypercube somewhere 😉 and no amount of steps to find it!

    That was some fun reading! 🙂 I went back into dimensional subspace where everything was connected by a single step 😉

    I see the light now, one step here, another there and voila from pink kittens to pink unicorns!

    What was so funny is using Wagner as a defense laid open the failures of neo-Darwinism. Yet the claim was self-organization and hypothetical dimensions solve the very problem Darwinist claimed did not exist. Interesting!

    Forgotten about that OP by you Gpuccio! Interesting look back! And even that I participated in it. Must have dropped off before the talk of astronomical library poofed into existence.

    And today, here we are with more evidence that your positions are rock solid. And where is the blind, unguided hyper-astronomical solution?

    We know Darwinism is dead, that Extended Synthesis is now at play in attempts to hold on to materialist doctrine. And neo-darwinism is dying on the vine as well, as some form of subset to whatever is next, hyper-astronomical libraries I guess.

    That is desperation for sure. But then, maybe from the beginning so was Darwin’s attempt to write off Design by a series of gradual steps.

  350. Dionisio, Gpuccio,

    “Procedures” and future OPs?

    What procedures? I’ve missed something after visiting hyper-astronomical dimensions.

  351. DATCG:

    Some time ago, I expressed my desire to write an OP about the problem of the “missing procedures”.

    What I meant was (and is) that with all that we know about the genome, and I would add also about epigenetics, we see a wonderful display of intelligent coordination and differentiation of programs and of regulations, but in the end we don’t know where and how the real procedures that control all that are written.

    In software, we have to write down the effectors, but we also need to write down the procedures that control the effectors.

    In biological software, we know much about the proteins (the effectors), but we understand too little about what controls the ordered and differentiated manifestations of proteins (and other components) in all the various engineered outcomes that we see in cells: above all, the different types of cells and cellular states in multicellular beings.

    Of course, today we know much more than yesterday. This same thread is evidence of that. And there is the huge fiels of epiganetics, which has added a lot of understanding.

    And we know a lot also of cell differentiation. A lot more than we knew, certainly.

    That’s the reason why I have renounced for the moment to deal with the problem of the missing procedures: I wanted to understand better what we know and what we don’t know. And, as I said to Dionisio, that has been, and is, a very “intensive preparation”.

    Because what we know is really amazing. And what we don’t know is esponentially more than what we know.

    However, I continue to believe that the procedures, those which are really essential, those which explain how things really work, are still missing. We know a lot of details, but we never understand what really controls the details.

    The fact remains that in that elusive genome, with its 20000+ protein coding genes, and its non coding DNA, plus all the possible information in the cytoplasm or other epigenetic markings, there must be the information sufficient to guide all the various cell differentiations which lead to tissues and organs. And there must be a satisying answer to the problem of morphogenesis.

    And there must be an answer to the architecure of systems like the immune system or, even worse, the nervous system of humans, for those who are not satisfied with the blind belief that a minor bundle of nucleotide variations in a few genes are enough to guide and determine the structure of 10^11 interconnected neurons, with all the advanced functions that, undeniably, the human brain seems to own.

    Those are the missing procedures. I don’t think that we have any good idea of where and how that information is written. But I hope that, as we gather billions and billions of new details (the famous “Big data” problem), sooner or later some major breakthrough will take place.

  352. Gpuccio,

    Thanks, it’s a bit overwhelming, amazing and adventurous 🙂

    I guess what we do know or can infer is it’s a Rules Based system, Modular and Conditional with massive amounts of parallel processing(billions of cells) and semiotic. The nervous system… whew…. back to being overwhelmed but in a good way 🙂

    But as you point out…

    Because what we know is really amazing. And what we don’t know is exponentially more than what we know.

    However, I continue to believe that the procedures, those which are really essential, those which explain how things really work, are still missing.

    We know a lot of details, but we never understand what really controls the details.

    Yes, many more details have come to light in just those few years since your 2015 post. And yet, so much more details to learn, remains! This is the great architecture of all systems in the world combined.

    The Sensors and signals processing fascinates me as that is part of control, procedural and rules based. The control systems are massive. Once a sensor detects an aberration, it’s not enough to react. It must react, signal, monitor, then decide to keep reacting or stop the reaction – in case of immune systems defense. Or in any other case as we discussed on the balance of protein synthesis and degradation and recycling, a delicate balance.

    I’ve been reading when I can about Sensors discovered in these or other processes. We know how important some systems are conserved. And we know why some are intentionally allowed to vary.

    We need a Big Table of Rules and Conditions for Big Data.

    Problem, Detection, Rules(If Then, Boolean, etc.), Signal, Proteins, Actions, Do-Until … Next Step

    Maybe that’s just to simplified, but somehow all of these actions need to be codified, procedurally assigned and pattern checked.

    Before I saw you and Dionisio discuss Procedures, an idea came that maybe one method of realization and visualization would be to track a specific change, damage, or protein life from start to finish across all systems interaction. From birth to death or recycling(degradation) in a cell?

    And then codify the specific actions along the way, Step 1, 2, 3, 4… Detection, Alerts, Rules to Alerts, Actions to Take, etc.

    Maybe Protein Synthesis, damage, repair, splicing, modification and eventual targeting for recycling. So far, all these excellent post by you from many different OPs has been looking at Modular Systems and Components. And those are overwhelming when we look at the multiplication of interactions like Ubiquitin. But if we dial it down to one specific point of functional interaction, then follow maybe it narrows down the thought process of uncovering procedures and rules?

    Is that fair to say? Along with a variety of interactions of course that grow and influence them. Again, I may have missed some other post in the past.

    Maybe it could be limited by intentional direction of steps. Leave out some conditional steps, maybe even whole steps at first and fill in those steps later. To keep it from being an overwhelming amount of information at once.

    Even mark areas “unknown” or TBD(to be discovered).

    Whatever we don’t know – will be fun to discover! Really appreciate all the efforts you provide and explanations. Thanks again. Looking back on your 2015 post was good to review.

  353. And… now I jump to the Kinetochore 🙂

    Which Dionisio first posted in Comment #22, then subsequent comments with differing roles of ubiquitin.

    We live in amazing times where we enjoy all of this rich information available to us these days.

    Here’s original video from 2012 starting 6 min mark of the Kinetochore broadcasting signals for eventual separation of chromosomes…

    https://www.youtube.com/watch?v=WFCvkkDSfIU

    Hope you guys had a great weekend.

  354. here’s another “simple” animation for Damaged DNA and Repair from 2011…

    DNA Damage Response to double-stranded DNA break — Homologous Recombination

    And accompanying information summary of work flow. Check out all the ubiqutin tags and ubiquitin chains.

    The DNA damage response (DDR) of the cell includes:
    (i) sensing,
    (ii) signalling
    (iii) repair of such damage

    Double strand breaks (DSBs) are the most toxical DNA damage for the cell. They can be induced by ionizing radiation, laser beam, bleomycin, Topoisomerase II enzyme, endonucleases or also can be produced during the repair of single-stranded breaks (SSB) DNA.

    It has been reported that approximately nine DSB per cell and day are produced in physiological conditions.

    The Homologus Recombination (HR) pathway is in charge of DSBs repair, in a error-free fashion, during S or G2 phases of the cell cycle, by using sister chromatid as template3.

    A proficient sensing and signaling of DSBs is very important for the maintenance of the genome and chromosomal stability.

    Recent research works, stressed out the key role of post-traslational modification of DDR proteins such as: phosphorilation, acetylation, methylation, ubiquitination and sumoylation in regulating the DNA damage signalling and response.

    The HR DDR signalling is believed to act in the following order:

    first, the DSB lesion is recognized by MRN complex (MRE11-RAD50-NBS1),

    that recruits the ATM (mutated in Ataxia Telangiectasia) kinase into the damage site.

    ATM phosphorilates the serine 139 of the ?H2AX at the damage site and also in large number of nucleosomes around the DSB.

    ATM also phosphorilates MDC1 (mediator of DNA damage checkpoint protein 1).

    At this point the ?H2AX-ATM-MDC1 connection generates a possitive feedback loop, that contributes to amplify DSB signal along the whole nucleous.

    Following this, RNF8 (an ubiquitin ligase), modifies ?H2AX.

    Then RNF168, an E3 ubiquitin ligase, detects the RNF8 signal in histones and amplify it by creating poly-ubiquitin chains.

    At this time, the RAP80-Abraxas complex is recruited into the damage site, followed by the BRCA1-BRCC36 complex.

    BARD1 it is believed to win the competition against Ctip protein for coupling with BRCA1-BRCC36 complex.

    An important role is attributed to RAP80 ubiquitination by RNF8, and BRCA1 SUMOylation by PIAS1-UBC9 complex.

    At this time, Ctip couples to MRN complex, displace most of the DDR proteins at DSB site, and catalyzes the DSBs ends resection by using its exonuclease activity.

    Immediately RPA binds to ssDNA. Then BRCA2 displaces RPA and enhance the RAD51 binding to the ssDNA filaments.

    This is the main signal for recruitment of recombination machinery to complete the DSB repair.

    Well, that’s a load… for sure!

    .

  355. DATCG at #355:

    Very good thoughts.

    I really don’t know how the problem of Big Data and of overwhelming details can be treated to get a glimpse of the controlling procedures.

    The problem in the end is to find where the information is.

    Let’s take for example the fundamental problem of the transition from single celled organisms to multicellular. The problem could be summed up as follows: if I start with soem single celled organism, let’s say yeast, and I want to get a multicellular organism, let’s say c. elegans, what kind of information do I need to add? Is it all genomic? And what is it exactly?

    Now, of course we know that yeast has “only” 6000 genes, while c. elegans has almost 20000. But the mere number of genes is not a good parameter, considering for example that some other fungus, like Phanerochaete chrysosporium, has almost 12000, and that drosophila has 13600, while humans themselves are at about 20000. The nubler of genes is not a good answer.

    Non coding DNA is probably better, but of course it is still difficult to understand the role of great part of it.

    Epigenetic states can keep a lot of information, but are they dependent on genomic information? Or can they store supplementary information which is tgransmitted directly, and does not rely on genomic stoiring?

    In the end, we know that the information, whatever its form, must be in some way connected to the physical organism that reproduces, because there is no doubt that it is transmitted: otherwise, we could not explain how the miracles of function and differentiation take place each time, with remarkable precision.

    I think we should probably consider a living being as a whole system, that includes genetic and epigenetic information which, at any moment, is in a dynamic state of interaction. The whole system probably bears the necessary information for life and reproduction and cell differentiation.

    But that system could store infromation in many different ways, some of which we certainly have to discover yet, some of which could be very different from what we usually think of.

    For example, I believe that we must go beyond the mere biochemical states, and look more deeply to what biophysics can tell us. We already know that for DNA biophysical states are complex and still poorly understood, and that they certainly have a major role in transcription regulation. The same can be said for proteins, especially considering what we have found about conditional folding, intrinsical disorder, and so on.

    The same can be said for cellular states, including organelles and compartments, both membrane-linked and membraneless.

    In a sense, the whole cell is a multi-faceted information system, that we still need to decode.

    Extremely interesting, from this point of view, is the goldfish-carp experiment referenced in the Denis Noble video linked by Dionisio at #271, starting at mark 31:46, and whose original paper can be found here:

    Cytoplasmic Impact on Cross-Genus Cloned Fish Derived from Transgenic Common Carp (Cyprinus carpio) Nuclei and Goldfish (Carassius auratus) Enucleated Eggs

    https://academic.oup.com/biolreprod/article/72/3/510/2666963

    And here is a follow-up, with interesting (and unexpected) news about mitochondria:

    The carp–goldfish nucleocytoplasmic hybrid has mitochondria from the carp as the nuclear donor species

    https://www.sciencedirect.com/science/article/pii/S0378111913016855?via%3Dihub

    Here is a more general review:

    The egg and the nucleus: a battle for supremacy

    http://dev.biologists.org/cont.....ong#sec-10

    And here, too:

    Interspecies Somatic Cell Nuclear Transfer: Advancements and Problems

    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3787369/

  356. gpuccio @358:

    “In the end, we know that the information, whatever its form, must be in some way connected to the physical organism that reproduces, because there is no doubt that it is transmitted: otherwise, we could not explain how the miracles of function and differentiation take place each time, with remarkable precision.”

    Oops! You used a politically incorrect word in scientific discussions: “miracles”

    You better watch out! Next time, please refrain from using words like that. 🙂

  357. DATCG at #356:

    Great video!

    The Kinetochore is certainly another incredible structure.

    From Wikipedia:

    A 2010 study uses a complex method (termed multiclassifier combinatorial proteomics or MCCP) to analyze the proteomic composition of vertebrate chromosomes, including kinetochores.[32] Although this study does not include a biochemical enrichment for kinetochores, obtained data include all the centromeric subcomplexes, with peptides from all 125 known centromeric proteins. According to this study, there are still about one hundred unknown kinetochore proteins, doubling the known structure during mitosis, which confirms the kinetochore as one of the most complex cellular substructures. Consistently, a comprehensive literature survey indicated that there had been at least 196 human proteins already experimentally shown to be localized at kinetochores

  358. Byrne, Robert, Thomas Mund, and Julien D. F. Licchesi. “Activity-Based Probes for HECT E3 Ubiquitin Ligases.” ChemBioChem 18, no. 14 (July 18, 2017): 1415–27. https://doi.org/10.1002/cbic.201700006.

    Flack, Joshua E., Juliusz Mieszczanek, Nikola Novcic, and Mariann Bienz. “Wnt-Dependent Inactivation of the Groucho/TLE Co-Repressor by the HECT E3 Ubiquitin Ligase Hyd/UBR5.” Molecular Cell 67, no. 2 (July 2017): 181–193.e5. https://doi.org/10.1016/j.molcel.2017.06.009.

    Gabrielsen, Mads, Lori Buetow, Mark A. Nakasone, Syed Feroj Ahmed, Gary J. Sibbet, Brian O. Smith, Wei Zhang, Sachdev S. Sidhu, and Danny T. Huang. “A General Strategy for Discovery of Inhibitors and Activators of RING and U-Box E3 Ligases with Ubiquitin Variants.” Molecular Cell 68, no. 2 (October 2017): 456–470.e10. https://doi.org/10.1016/j.molcel.2017.09.027.

    Gorelik, Maryna, and Sachdev S. Sidhu. “Specific Targeting of the Deubiquitinase and E3 Ligase Families with Engineered Ubiquitin Variants: Gorelik and Sidhu.” Bioengineering & Translational Medicine 2, no. 1 (March 2017): 31–42. https://doi.org/10.1002/btm2.10044.

    Iimura, Akira, Fuhito Yamazaki, Toshiyasu Suzuki, Tatsuya Endo, Eisuke Nishida, and Morioh Kusakabe. “The E3 Ubiquitin Ligase Hace1 Is Required for Early Embryonic Development in Xenopus Laevis.” BMC Developmental Biology 16, no. 1 (December 2016). https://doi.org/10.1186/s12861-016-0132-y.

    Kai, Masatake, Naoto Ueno, and Noriyuki Kinoshita. “Phosphorylation-Dependent Ubiquitination of Paraxial Protocadherin (PAPC) Controls Gastrulation Cell Movements.” Edited by Jung Weon Lee. PLOS ONE 10, no. 1 (January 12, 2015): e0115111. https://doi.org/10.1371/journal.pone.0115111.

    Lorenz, Sonja. “Structural Mechanisms of HECT-Type Ubiquitin Ligases.” Biological Chemistry 399, no. 2 (January 26, 2018). https://doi.org/10.1515/hsz-2017-0184.
    Mlodzik, Marek. “Ubiquitin Connects with Planar Cell Polarity.” Cell 137, no. 2 (April 2009): 209–11. https://doi.org/10.1016/j.cell.2009.04.002.

    Mund, Thomas, Michael Graeb, Juliusz Mieszczanek, Melissa Gammons, Hugh R. B. Pelham, and Mariann Bienz. “Disinhibition of the HECT E3 Ubiquitin Ligase WWP2 by Polymerized Dishevelled.” Open Biology 5, no. 12 (December 2015): 150185. https://doi.org/10.1098/rsob.150185.

    Narimatsu, Masahiro, Rohit Bose, Melanie Pye, Liang Zhang, Bryan Miller, Peter Ching, Rui Sakuma, et al. “Regulation of Planar Cell Polarity by Smurf Ubiquitin Ligases.” Cell 137, no. 2 (April 2009): 295–307. https://doi.org/10.1016/j.cell.2009.02.025.

    Ramakrishnan, Aravinda-Bharathi, Abhishek Sinha, Vinson B. Fan, and Ken M. Cadigan. “The Wnt Transcriptional Switch: TLE Removal or Inactivation?” BioEssays 40, no. 2 (February 2018): 1700162. https://doi.org/10.1002/bies.201700162.

    Xie, Zhongdong, Han Liang, Jinmeng Wang, Xiaowen Xu, Yan Zhu, Aizhen Guo, Xian Shen, Fuao Cao, and Wenjun Chang. “Significance of the E3 Ubiquitin Protein UBR5 as an Oncogene and a Prognostic Biomarker in Colorectal Cancer.” Oncotarget 8, no. 64 (December 8, 2017). https://doi.org/10.18632/oncotarget.22531.

    Zhang, Wei, Maria A. Sartori, Taras Makhnevych, Kelly E. Federowicz, Xiaohui Dong, Li Liu, Satra Nim, et al. “Generation and Validation of Intracellular Ubiquitin Variant Inhibitors for USP7 and USP10.” Journal of Molecular Biology 429, no. 22 (November 2017): 3546–60. https://doi.org/10.1016/j.jmb.2017.05.025.

  359. Dionisio:

    “Oops! You used a politically incorrect word in scientific discussions: “miracles””

    It’s an old problem. In my whole life, I have never been able to be politically correct! 🙂

  360. DATCG at #357:

    DNA repair: another key issue!

    Just a curiosity: many of the proteins involved have a similar evolutionary history, more or less as can be seen in Fig. 5 in the OP for BRCA1, with a late development of the human sequence, and some important late jump in mammals.

    They are:

    RNF8 (485 AAs)
    RNF168 (571 AAs)
    BRCA1 (1863 AAs)
    BRCA2 (3418 Aas)
    MDC1 (2089 AAs)
    RAP80 (719 AAs)
    BARD1 (777 AAs)
    Ctip (897 AAs)

    Other proteins in the process, instead, are older (highly conserved in eukaryotes), and a couple mof them are mainly engineered (in human form) at the vertebrate transition.

    The presence of so many late-engineered proteins is maybe surprising, because DNA repair seems to be an old problem. But it seems that it requires new or more specific solutions, especially in mammals. It would be interesting to understand why.

  361. Gpuccio #358, re: Dionisio’s #271,

    Great video Find Dionisio!

    I love it when we see the other side admit the real issues are wide open and unsolved by Darwinian mechanisms!

    Excellent, it’s another addition of information I had not reviewed yet. I quickly went to minute 30 Dionisio remarked on and very interesting to hear Denis Noble’s remarks.

    Gold Fish, Carp -> nucleus replacment = -> Something in the Middle 🙂 🙂 🙂

    Thanks for a clear and sober look at state of genomics, epigenome and many other issues related to cellular organisms and evolution.

    Going back to a previous OP you recommended earlier, where Darwinist supporters kept trying to say it was easy to see evolution “did it” mantra. I see they would glance over these difficult issues in favor of story telling.

    Leaving out vast amounts of details and frankly, millions of gradual steps if they were to be honest about Random mutations and Natural Selection via gradual process.

    And what was interesting is the appeal to Wagner as if his wild imaginations about hyper-astronomical library eliminates all the steps and makes it easier to evolve.

    So, now they’ve eliminated Darwin?

    Haha… they really are in a Catch-22 these days.

    At least Noble of Third Way admits Darwin’s dead and one of his earlier videos shreds Richard Dawkins.

    Though they refuse to admit Design to the table, Third Way is admitting failure of neo-darwinism to save Darwin.

  362. From the video Dionisio posted @271, Denis Noble,

    @31.40 mark…

    Denis Noble @31.44

    “I include this slide because it beautifully illustrates that vastly more must be transmitted to the next generation than just the DNA of the nucleus…”

    As he’s showing the slide of the Goldfish, the Carp and the resulting combination leading to something aligned in the middle to a Gold-Carp 😉

    He then goes on to quote cytoplasmic factors in the egg cell from a paper by a Chinese Scientist.

    BTW, anyone else notice how Asian nations are not held down by Darwinism? They think outside the “Black Box?”

    Darwin I think half the time remains due to England and Western influence of society, not due to scientific rigor.

    It’s as if the old guard cannot let go of a 19th century failed paradigm.

  363. DATCG:

    “And what was interesting is the appeal to Wagner as if his wild imaginations about hyper-astronomical library eliminates all the steps and makes it easier to evolve.”

    Frankly, I could never find anything credible in what Wagner says.

    “Though they refuse to admit Design to the table, Third Way is admitting failure of neo-darwinism to save Darwin.”

    Yes, in a sense they are.

    But I must admit that it is more difficult for me to understand people who understand and don’t admit. In a sense, I have more sympathy for Dawkins…

    “BTW, anyone else notice how Asian nations are not held down by Darwinism? They think outside the “Black Box?””

    Yes, I noticed. They are doing a lot of good work. I think they are probably more competitive, and interested to the results. And truth is often needed to get results! 🙂

    “Darwin I think half the time remains due to England and Western influence of society, not due to scientific rigor.”

    It’s the lingering power of a static Academia, still conditioned by old philosophies and by ever young political feuds.

  364. DATCG, Dionisio:

    This is new and interesting:

    Ubiquitin Modulates Liquid-Liquid Phase Separation of UBQLN2 via Disruption of Multivalent Interactions.

    http://www.cell.com/molecular-.....0102-3.pdf

    Abstract:

    Under stress, certain eukaryotic proteins and RNA assemble to form membraneless organelles known as stress granules. The most well-studied stress granule components are RNA-binding proteins that undergo liquid-liquid phase separation (LLPS) into protein-rich droplets mediated by intrinsically disordered low-complexity domains (LCDs). Here we show that stress granules include proteasomal shuttle factor UBQLN2, an LCD-containing protein structurally and functionally distinct from RNA-binding proteins. In vitro, UBQLN2 exhibits LLPS at physiological conditions. Deletion studies correlate oligomerization with UBQLN2’s ability to phase-separate and form stress-induced cytoplasmic puncta in cells. Using nuclear magnetic resonance (NMR) spectroscopy, we mapped weak, multivalent interactions that promote UBQLN2 oligomerization and LLPS. Ubiquitin or polyubiquitin binding, obligatory for UBQLN2’s biological functions, eliminates UBQLN2 LLPS, thus serving as a switch between droplet and disperse phases. We postulate that UBQLN2 LLPS enables its recruitment to stress granules, where its interactions with ubiquitinated substrates reverse LLPS to enable shuttling of clients out of stress granules.

    There seems to be almost averything here: ubiquitin chains, membraneless organelles, intrinsically disordered domains.

    UBQLN2 (Ubiquilin 2) is a strange object. It is a 624 AAs protein, and here is the Uniprot function section:

    Plays an important role in the regulation of different protein degradation mechanisms and pathways including ubiquitin-proteasome system (UPS), autophagy and the endoplasmic reticulum-associated protein degradation (ERAD) pathway. Mediates the proteasomal targeting of misfolded or accumulated proteins for degradation by binding (via UBA domain) to their polyubiquitin chains and by interacting (via ubiquitin-like domain) with the subunits of the proteasome (PubMed:10983987). Plays a role in the ERAD pathway via its interaction with ER-localized proteins FAF2/UBXD8 and HERPUD1 and may form a link between the polyubiquitinated ERAD substrates and the proteasome (PubMed:24215460, PubMed:18307982). Involved in the regulation of macroautophagy and autophagosome formation; required for maturation of autophagy-related protein LC3 from the cytosolic form LC3-I to the membrane-bound form LC3-II and may assist in the maturation of autophagosomes to autolysosomes by mediating autophagosome-lysosome fusion (PubMed:19148225, PubMed:20529957). Negatively regulates the endocytosis of GPCR receptors: AVPR2 and ADRB2, by specifically reducing the rate at which receptor-arrestin complexes concentrate in clathrin-coated pits (CCPs) (PubMed:18199683).

    It has an ubiquitin-like domain, but technically it does not seem to be an E3 ligase, nor an ubiquitin binding protein. But it certainly regulates many ubiquitin related pathways.

    Things get stranger and stranger! 🙂

  365. DATCG, Dionisio:

    In plants:

    E3 ubiquitin ligases: key regulators of hormone signaling in plants

    http://www.mcponline.org/conte.....6.full.pdf

    Abstract
    Ubiquitin-mediated control of protein stability is central to most aspects of plant hormone signaling. Attachment of ubiquitin to target proteins occurs via an enzymatic cascade with the final step being catalyzed by a family of enzymes known as E3 ubiquitin ligases, which have been classified based on their protein domains and structures. While E3 ubiquitin ligases are conserved among eukaryotes, in plants they are well-known to fulfill unique roles as central regulators of phytohormone signaling, including hormone perception and regulation of hormone biosynthesis. This review will highlight up-to-date findings that have refined well-known E3 ligase-substrate interactions and defined novel E3 ligase substrates that mediate numerous hormone signaling pathways. Additionally, examples of how particular E3 ligases may mediate hormone crosstalk will be discussed as an emerging theme. Looking forward, promising experimental approaches and methods that will provide deeper mechanistic insight into the roles of E3 ubiquitin ligases in plants will be considered.

    Now, I suppose that regulating hormone crosstalk in plants is something again completely different from all the functions we have already listed.

    It is really amazing how the ubiquitin system seems capable of regulating practically anything! 🙂

  366. DATCG, Dionisio:

    This is really interesting. It introduces us to a new code, the redox code, and a nex cross-talk with the ubiquitin code.

    Ube2V2 Is a Rosetta Stone Bridging Redox and Ubiquitin Codes, Coordinating DNA Damage Responses.

    https://pubs.acs.org/doi/pdf/10.1021/acscentsci.7b00556

    Abstract:

    Posttranslational modifications (PTMs) are the lingua franca of cellular communication. Most PTMs are enzyme-orchestrated. However, the reemergence of electrophilic drugs has ushered mining of unconventional/non-enzyme-catalyzed electrophile-signaling pathways. Despite the latest impetus toward harnessing kinetically and functionally privileged cysteines for electrophilic drug design, identifying these sensors remains challenging. Herein, we designed “G-REX”-a technique that allows controlled release of reactive electrophiles in vivo. Mitigating toxicity/off-target effects associated with uncontrolled bolus exposure, G-REX tagged first-responding innate cysteines that bind electrophiles under true kcat/Km conditions. G-REX identified two allosteric ubiquitin-conjugating proteins-Ube2V1/Ube2V2-sharing a novel privileged-sensor-cysteine. This non-enzyme-catalyzed-PTM triggered responses specific to each protein. Thus, G-REX is an unbiased method to identify novel functional cysteines. Contrasting conventional active-site/off-active-site cysteine-modifications that regulate target activity, modification of Ube2V2 allosterically hyperactivated its enzymatically active binding-partner Ube2N, promoting K63-linked client ubiquitination and stimulating H2AX-dependent DNA damage response. This work establishes Ube2V2 as a Rosetta-stone bridging redox and ubiquitin codes to guard genome integrity.

    (Public access. Emphasis mine)

    Some interesting passages:

    Through a phenomenal research effort we now understand much about complex post-translational regulation in cell signaling. Approximately 10% of the genome is involved in phosphorylation and ubiquitination

    Against the backdrop of these exquisite enzyme-regulated signaling subsystems, the cell has also harnessed reactive smallmolecule signaling mediators to fine-tune responses. In this paradigm, reactive oxygen or electrophilic species (ROS/RES) directly modify a specific signal-sensing protein, preempting decision-making.

    These data indicate that redox signaling HNEylation of one regulatory protein (at a site with no “expected” reactivity) can affect ubiquitin signaling via a third-party enzyme containing a catalytic cysteine (Ube2N).

  367. DATCG, Dionisio:

    This is about ROS (Reactive oxygen species) and RES (Reactive electrophile species). RES seem to be specially important in signaling.

    ROS-mediated lipid peroxidation and RES-activated signaling.

    https://www.annualreviews.org/doi/abs/10.1146/annurev-arplant-050312-120132?rfr_dat=cr_pub%3Dpubmed&url_ver=Z39.88-2003&rfr_id=ori%3Arid%3Acrossref.org&journalCode=arplant

    Abstract;

    Nonenzymatic lipid oxidation is usually viewed as deleterious. But if this is the case, then why does it occur so frequently in cells? Here we review the mechanisms of membrane peroxidation and examine the genesis of reactive electrophile species (RES). Recent evidence suggests that during stress, both lipid peroxidation and RES generation can benefit cells. New results from genetic approaches support a model in which entire membranes can act as supramolecular sinks for singlet oxygen, the predominant reactive oxygen species (ROS) in plastids. RES reprogram gene expression through a class II TGA transcription factor module as well as other, unknown signaling pathways. We propose a framework to explain how RES signaling promotes cell “REScue” by stimulating the expression of genes encoding detoxification functions, cell cycle regulators, and chaperones. The majority of the known biological activities of oxygenated lipids (oxylipins) in plants are mediated either by jasmonate perception or through RES signaling networks.

    4-Hydroxynonenal (HNE) is the RES product quoted in the paper linked at my previous comment.

  368. DATCG, Dionisio:

    The degron scenario is not complete, yet. Here comes the role of N terminal arginylation:

    N-terminal arginylation generates a bimodal degron that modulates autophagic proteolysis.

    http://www.pnas.org/content/ea.....10115.long

    Abstract
    The conjugation of amino acids to the protein N termini is universally observed in eukaryotes and prokaryotes, yet its functions remain poorly understood. In eukaryotes, the amino acid l-arginine (l-Arg) is conjugated to N-terminal Asp (Nt-Asp), Glu, Gln, Asn, and Cys, directly or associated with posttranslational modifications. Following Nt-arginylation, the Nt-Arg is recognized by UBR boxes of N-recognins such as UBR1, UBR2, UBR4/p600, and UBR5/EDD, leading to substrate ubiquitination and proteasomal degradation via the N-end rule pathway. It has been a mystery, however, why studies for the past five decades identified only a handful of Nt-arginylated substrates in mammals, although five of 20 principal amino acids are eligible for arginylation. Here, we show that the Nt-Arg functions as a bimodal degron that directs substrates to either the ubiquitin (Ub)-proteasome system (UPS) or macroautophagy depending on physiological states. In normal conditions, the arginylated forms of proteolytic cleavage products, D101-CDC6 and D1156-BRCA1, are targeted to UBR box-containing N-recognins and degraded by the proteasome. However, when proteostasis by the UPS is perturbed, their Nt-Arg redirects these otherwise cellular wastes to macroautophagy through its binding to the ZZ domain of the autophagic adaptor p62/STQSM/Sequestosome-1. Upon binding to the Nt-Arg, p62 acts as an autophagic N-recognin that undergoes self-polymerization, facilitating cargo collection and lysosomal degradation of p62-cargo complexes. A chemical mimic of Nt-Arg redirects Ub-conjugated substrates from the UPS to macroautophagy and promotes their lysosomal degradation. Our results suggest that the Nt-Arg proteome of arginylated proteins contributes to reprogramming global proteolytic flux under stresses.

    IOWs, ubiquinated proteins are usually dergaded by the proteasome, but if the proteasoms is in trouble, they are shifted to the macroautophagy pathway, and the switch seems to be N-terminal arginylation.

    This is a complrehensive review about this “alternative” degradation pathway:

    p62/SQSTM1/Sequestosome-1 is an N-recognin of the N-end rule pathway which modulates autophagosome biogenesis

    Abstract:

    Macroautophagy mediates the selective degradation of proteins and non-proteinaceous cellular constituents. Here, we show that the N-end rule pathway modulates macroautophagy. In this mechanism, the autophagic adapter p62/SQSTM1/Sequestosome-1 is an N-recognin that binds type-1 and type-2 N-terminal degrons (N-degrons), including arginine (Nt-Arg). Both types of N-degrons bind its ZZ domain. By employing three-dimensional modeling, we developed synthetic ligands to p62 ZZ domain. The binding of Nt-Arg and synthetic ligands to ZZ domain facilitates disulfide bond-linked aggregation of p62 and p62 interaction with LC3, leading to the delivery of p62 and its cargoes to the autophagosome. Upon binding to its ligand, p62 acts as a modulator of macroautophagy, inducing autophagosome biogenesis. Through these dual functions, cells can activate p62 and induce selective autophagy upon the accumulation of autophagic cargoes. We also propose that p62 mediates the crosstalk between the ubiquitin-proteasome system and autophagy through its binding Nt-Arg and other N-degrons.

    p62, or Sequestosome 1, is an ubiquitin binding protein.

  369. Posttranslational modifications (PTMs) are the lingua franca of cellular communication.

    BOOM!

    – – – – – – – – – – – –

    EDIT: But hey, there’s no evidence of design in biology. 🙂

  370. #369 Gpuccio,

    Wow! Great find!

    Ahaha, Upright BiPed is dancing the Post-Edit, Semiotic dance 😉

    From wiki…

    Lingua Franca:

    A lingua franca, also known as a bridge language, common language, trade language, vehicular language, or link language is a language or dialect systematically used to make communication possible between people who do not share a native language or dialect, particularly when it is a third language that is distinct from both native languages.

    Darwinist, Darwinist, whatcha you gonna do, whatcha gonna do when Design comes for you?

    .

  371. #371 Gpuccio,

    Another great find, those little Degrons are important. Also, Recognins.

    It has been a mystery, however, why studies for the past five decades identified only a handful of Nt-arginylated substrates in mammals, although five of 20 principal amino acids are eligible for arginylation.

    Here, we show that the Nt-Arg functions as a bimodal degron that directs substrates to either the ubiquitin (Ub)-proteasome system (UPS) or macroautophagy depending on physiological states.

    Check Physiological State – Stress Table

    IF Condition-State = Normal, Select Proteasome Pathway, Else
    IF Condition-State = Pertubed, Select ChangePath; MacroAutophagy Pathway

    In normal conditions, the arginylated forms of proteolytic cleavage products, D101-CDC6 and D1156-BRCA1, are targeted to UBR box-containing N-recognins and degraded by the proteasome.

    However, when(IF) proteostasis by the UPS is perturbed, their Nt-Arg redirects these otherwise cellular wastes to macroautophagy through its binding to the ZZ domain of the autophagic adaptor p62/STQSM/Sequestosome-1. Upon binding to the Nt-Arg, p62 acts as an autophagic N-recognin that undergoes self-polymerization, facilitating cargo collection and lysosomal degradation of p62-cargo complexes.

    A chemical mimic of Nt-Arg redirects Ub-conjugated substrates from the UPS to macroautophagy and promotes their lysosomal degradation.

    Our results suggest that the Nt-Arg proteome of arginylated proteins contributes to reprogramming global proteolytic flux under stresses.

    (IF) emphasis mine

    This is a pre-programmed backup response to stress conditions overload checkpoint. If overload conditions exist, tag, reroute for destruction until conditions change back to normal.

    Remember, it has to 1) signal overload or recognize stress conditions, 2) tag and mark for rerouting, 3) reroute and destruct, 3) once stress conditions or physiological conditions are no longer true, turn off tagging and marking signal for rerouting and resume normal condition posture.

    .

  372. Upright BiPed:

    I thought you would like it! 🙂

    This redox code is really interesting, maybe we will have to deepen the issue sometime.

  373. DATCG:

    Redox code, degrons, recognins: ubiquitin is certainly offering us a lot of beautiful gifts! 🙂

  374. #371 Gpuccio, Dionisio, Upright BiPed,

    So, why have a conditional check and backup system for overload?

    Why have two pathway systems for degradation and recyling: Proteasome and Autophagy at all?

    I think a typical Darwin’s response would be, this is wasteful and shows no planning, a result of random mutations and natural selection.

    It’s a “Bad” Design.

    OK, so back to “bad” design argument for Darwinist? Those type of Darwinist arguments failed for Eye Design.

    Returning to PTM-Posttranslational Modification. A nice chart(2013) below follows of pathways for Proteasome, CMA(chaperone-mediated autophagy) and Macroautophagy(or autophagy).

    Or, A pre-programmed Quality Control System for distribution and recycling?

    This answers a question(for me – clarifies it) many days ago on Protein Aggregation and balance. Essentially this is Quality Control Systems. And any good QCS has multiple checks and balances especially during times of stress, or in this case aggregation of unwanted product backlogs.

    Posttranslational Modification and Quality Control – Figures and Tables

    Xuejun Wang, J. Scott Pattison, Huabo Su
    https://doi.org/10.1161/CIRCRESAHA.112.268706
    Circulation Research. 2013;112:367-381
    Originally published January 17, 2013

    Figure 1:
    An illustration of protein quality control in the cell.

    Chaperones help fold nascent polypeptides, unfold misfolded proteins and refold them, and channel terminally misfolded proteins for degradation by the ubiquitin-proteasome system (UPS) or chaperone-mediated autophagy (CMA). When(IF) escaped from targeted degradation, misfolded proteins form aggregates via hydrophobic interactions.

    Aggregated proteins can be selectively targeted by macroautophagy to, and degraded by, the lysosome. hsc indicates heat shock cognate 70; and LAMP-2A, lysosome-associated membrane protein 2A.

    Very interesting. Also note: Tables at bottom of page.

    Examples of Posttranslational Modifications in Intracellular Quality Control

    Targets
    PTMs
    Regulating Enzymes
    Biological Function

    The Research Paper at Circulation Research:
    Posttranslational Modification and Quality Control

    Some concluding remarks from the paper:


    Dissection of the upstream pathways that regulate the PTM will be crucial to identify novel targets or strategies for developing pharmaceutics to improve QC in the cell.

    It is anticipated that comprehensive investigations into intracellular QC in cardiac physiology and pathology will give rise to new therapeutics to better battle heart diseases, the leading cause of death of humans.

    On macroautophagy, there is a good body of evidence supporting that activation of macroautophagy improves PQC and thereby protects the heart23; nonetheless, excessive macroautophagy on certain conditions such as reperfusion may be detrimental.94

    Some studies, but not others, have shown that pharmacologically induced ubiquitous proteasome inhibition protects against I/R injury and pressure-overloaded cardiac hypertrophy.143,144 However, genetically induced moderate proteasome inhibition in cardiomyocytes was recently shown to exacerbate acute I/R injury in mice.18 Furthermore, administration of bortezomib, a proteasome inhibitor, to multiple myeloma patients can cause reversible heart failure.145 Conversely, recent genetic studies reveal that proteasome function enhancement in the cardiomyocytes of diseased hearts can slow down the progression of a bona fide cardiac proteinopathy and minimize acute I/R injury in mice.2 Hence, it is envisioned that enhancing proteasome proteolytic function may be a potential new strategy to treat heart diseases with increased proteolytic stress.

    Question – what’s causing the need for Enhanced “proteasome proteolytic function?” Should not the CAUSE for Increased proteolytic function be target of research?

    People eat bad processed foods, then researchers must “fix” the problematic results on the corrosive downside of abnormal functionality . Instead of correcting the input side, our eating habits.

    GIGO = Garbage In, Garbage Out

    Or, You can’t have your cake and eat it too

    Correct bad eating habits, correct the outcome. Correct the soaring cost of health care.

    This is only one area. Obviously stress conditions can arise unrelated to eating habits.

    .

  375. Gpuccio,

    “Redox code, degrons, recognins: ubiquitin is certainly offering us a lot of beautiful gifts!”

    Beautiful indeed 🙂 Screaming Design once again. Wish I had more time to go through all the different papers you and Dionisio post here.

    Such a great treasure trove of Biological Function and FSOC(Functional Sequence Organized Complexity) and Irreducible Complexity!

    Lets take out Redox Codes. the Bridge of Semiotic Language, Degrons, Recognins between the two systems of Proteasome and Autophagy, and what happens to Quality Control?

  376. From the previous paper on Quality Control I posted, from the Intro…

    “Hence, the cell has evolved intracellular quality control (QC) mechanisms at protein and organelle levels to minimize the level and toxicity of misfolded proteins and defective organelles in the cell.”

    Wow, blind, unguided mutations and natural selection Did It.
    Just wham, bam, thank you mutation Ma’am magically created a Quality Control System.

    This blind, unguided mythology is sheer genius. In doing so, it created a semiotic system of post-translation communication systems and codes to bridge between two other systems control features.

    Darwin is Magic and Magic is Darwin.

  377. Looking at the Magic of Darwin…

    From the Intro of Previous posted Paper @377 on Quality Control Mechanisms and determination of Protein Degradation choices.

    Note: Edited sections for breakout of roles

    Intracellular QC is regulated by several mechanisms:
    transcriptional
    translational
    posttranslational

    Posttranslational modifications (PTMs) are:

    – phosphorylation
    – ubiquitination
    – nitrosylation
    – oxidation
    – and more…

    Posttranslational mechanisms:

    – expand size of the proteome exponentially
    – are pivotal in the regulation of proteins

    in the need for Protein:
    – stability
    – distribution
    – and function

    Emerging evidence supports a major role of PTMs in regulating multiple pathways of intracellular Quality Control.

    Protein Quality Control(PQC) is:

    A set of molecular mechanisms. Ensuring that misfolded and damaged proteins are:

    – repaired or removed in a timely fashion

    Thereby minimizing the toxic effects of misfolded proteins (Figure 1 – see above figure in #377).

    The QC of proteins targeted for the secretary pathway (ie, proteins passing through the Endoplasmic Reticulum[ER]) is performed by ER-associated Protein Quality Control.

    This involves retrotranslocation.

    Where:
    1) misfolded proteins from ER lumen are moved to the cytosol

    2) degradation of them via ER-associated degradation pathways such as the Proteasome for instance

    There is a different Protein Quality Control System for non-ER proteins.

    In both cases:
    – PQC is performed by molecular chaperones and target protein degradation.
    – Chaperones serve as the sensor of misfolded proteins
    – and in some cases attempt to Repair misfolding by unfolding/refolding

    IF repair fails:

    misfolded proteins termed as a terminally misfolded proteins are escorted by chaperones for:

    – degradation primarily by the ubiquitin-proteasome system (UPS)
    – and perhaps by chaperone-mediated autophagy (CMA).

    When(IF) misfolded proteins escape the surveillance of chaperones and target degradation:

    – they tend to form aberrant aggregates.

    The intermediate, highly unstable, soluble species of aggregates are very toxic to the cell.4

    Small aggregates assimilate into larger ones that are insoluble and perhaps less toxic to the cell.

    Finally, with assistance from the microtubule transportation system,

    – small aggregates may be translocated to the microtubule organizing center to fuse with one another to form large inclusion bodies, termed, by some, aggresomes.5

    The insoluble aggregates and aggresomes are:

    – unlikely to be accessible to the proteasome and CMA
    – both of which can only degrade proteins individually

    Hence, the removal of aggregated proteins requires a different mechanism that is capable of bulk degradation of substrates, a role filled by macroautophagy.

    And there’s the reason for multiple recycling functions and larger scale systems of degradation.

    What’s interesting is the aggregation system – aggresomes. This cannot happen by accident either. It must be organized or damaged, misfolded proteins just wonder all over the cytoplasm or ER.

    If individual processing breaks down. To relieve Qaulity Control Systems backlog, the Backlog Checking System springs into action rerouting individual proteins targeted for recycling to Bulk Degradation – autophagy.

    Lovely! My gosh how well Darwin Magic works. 😉

    .

  378. DNA Replication, the Replisome, PCNA Ubiquitination and Quality Control Systems Limitations or the ability of neo-Darwinian Magic to “evolve” Highly Organized Complex Functional Network Regulatory Systems to Halt, Decide, and designate different Repair Mechanisms.

    The Replication Fork: Understanding the Eukaryotic Replication Machinery and the Challenges to Genome Duplication

    Published 2013 Adam R. Leman†* and Eishi Noguchi*

    Abstract

    Abstract
    Eukaryotic cells must accurately and efficiently duplicate their genomes during each round of the cell cycle. Multiple linear chromosomes, an abundance of regulatory elements, and chromosome packaging are all challenges that the eukaryotic DNA replication machinery must successfully overcome. The replication machinery, the “replisome” complex, is composed of many specialized proteins with functions in supporting replication by DNA polymerases. Efficient replisome progression relies on tight coordination between the various factors of the replisome. Further, replisome progression must occur on less than ideal templates at various genomic loci.

    Here, we describe the functions of the major replisome components, as well as some of the obstacles to efficient DNA replication that the replisome confronts. Together, this review summarizes current understanding of the vastly complicated task of replicating eukaryotic DNA.

    Keywords: DNA replication, replisome, replication fork, genome stability, checkpoint, fork barriers, difficult-to-replicate sites, (PCNA Ubiquitination)

    ( ) emphasis mine

    The DNA Sliding Clamp: PCNA

    DNA sliding clamps have evolved, promoting the processivity of replicative polymerases. In eukaryotes, this sliding clamp is a homotrimer known as Proliferating Cell Nuclear Antigen(PCNA), which form a ring structure.

    The PCNA ring has polarity with a surface that interacts with DNA polymerases and tethers them securely to DNA. PCNA-dependent stabilization of DNA polymerases has a significant effect on DNA replication because it enhances polymerase processivity up to 1,000-fold.

    Various PCNA modifications regulate the replisome through specific circumstances during DNA replication. The modifications of PCNA have dramatic effects on its function. Although there are some species-specific modifications of PCNA throughout eukaryota, the principles remain conserved.

    Upon DNA damage, PCNA is monoubiquitinated, which changes PCNA’s affinity from replicative polymerases to the damage-tolerant translesion synthesis (TLS) polymerases [61,62].

    PCNA ubiquitination is dependent on the DNA damage checkpoint pathway and regulates dynamic changes in the replication fork.

    This process allows for bypass of bulky DNA damage that would otherwise prevent replication fork progression, although this method of damage bypass is error prone [62,63].

    In contrast, polyubiquitination of the same site directs the cell towards DNA damage bypass by poorly characterized, but essentially error-free mechanisms [64,65,66].

    PCNA can also be SUMOylated (small ubiquitin-like modifier) at the same site in yeast, and SUMOylated PCNA exists in vertebrates [67,68].

    This modification is thought to suppress ubiquitination of PCNA, therefore inhibiting TLS and other DNA repair pathways, which are potentially harmful to the cell because they can introduce mutations and genome rearrangements [69,70,71].

    These findings indicate that PCNA modifications play critical roles in controlling pathway selection for DNA damage management during DNA replication.

    Clamp Loaders evolved

    Eukaryotes have evolved multiple clamp loading complexes, each of which appears to function in a separate pathway.

    The canonical clamp loader essential for DNA replication is RFC and includes Rfc1, Rfc2, Rfc3, Rfc4 and Rfc5. At least three RFC-like complexes exist in eukaryotic cells. RFCCtf18, which contains Ctf18 in place of Rfc1, promotes sister chromatid cohesion and regulates replication speed [78,79,80,81]. RFCElg1, which contains Elg1, is thought to unload SUMOylated PCNA in the presence of DNA damage to allow for replication progression through damaged DNA templates [71].

    The RFCRad17/Rad24 clamp does not load PCNA, but loads the 9-1-1 complex at DNA damage sites during the replication checkpoint response [82].

    Thus, DNA replication can be regulated at the level of PCNA clamp loading, in order to accommodate multiple processes that take place during DNA replication (Figure 4).

    Sliding Clamps, Clamp Loaders, Processivity “evolved” along with all the checkpoint mechanisms, signals and ubiquitination for DNA regulation of DNA replication repair.

    Magic Darwin

    .

  379. As a follow-up to #381, PCNA and therefore ubiquitin mono and Poly are invovled in this paper’s coverage below. But do not have access to full paper.

    Behind Paywall…


    The Eukaryotic Replication Machine.

    Zhang D1, O’Donnell M2.
    Author information
    1 The Rockefeller University, New York, NY, United States.
    2 The Rockefeller University, New York, NY, United States; Howard Hughes Medical Institute, The Rockefeller University, New York, NY, United States. Electronic address: odonnel@rockefeller.edu.

    Abstract

    The cellular replicating machine, or “replisome,” is composed of numerous different proteins. The core replication proteins in all cell types include a helicase, primase, DNA polymerases, sliding clamp, clamp loader, and single-strand binding (SSB) protein.

    The core eukaryotic replisome proteins evolved independently from those of bacteria and thus have distinct architectures and mechanisms of action.

    The core replisome proteins of the eukaryote include:
    11-subunit CMG helicase
    DNA polymerase alpha-primase
    leading strand DNA polymerase epsilon
    lagging strand DNA polymerase delta
    PCNA clamp
    RFC clamp loader
    and RPA SSB protein

    There are numerous other proteins that travel with eukaryotic replication forks, some of which are known to be involved in checkpoint regulation or nucleosome handling, but most have unknown functions and no bacterial analogue.

    Recent studies have revealed many structural and functional insights into replisome action. Also, the first structure of a replisome from any cell type has been elucidated for a eukaryote, consisting of 20 distinct proteins, with quite unexpected results.

    This review summarizes the current state of knowledge of the eukaryotic core replisome proteins, their structure, individual functions, and how they are organized at the replication fork as a machine.

  380. More on Replisome and Ubiquitin regulation in Nature: Cell Death and Differentiation

    News and Commentary (Open Access)


    Two Paths to Let the Replisome Go

    Vincenzo D’Angiolella & Daniele Guardavaccaro
    Cell Death and Differentiation(2017)24,1140–1141; doi:10.1038/cdd.2017.75; published online 19 May 2017

    Accurate DNA replication is essential for genome maintenance. Two recent reports have uncovered new molecular mechanisms controlling the termination phase of DNA replication in higher eukaryotes and established crucial roles for the CRL2LRR1 ubiquitin ligase and the p97 segregase in replisome unloading from chromatin.

    Eukaryotic DNA replication can be divided into three distinct steps.

    During licensing, pre-replication complexes (pre-RCs) assemble at DNA replication origins in the G1 phase of the cell cycle.

    This is followed by replication initiation at the G1-S phase transition, when CDKs (cyclin-dependent kinases) and DDKs (DBF4-dependent kinases) promote the recruitment of the GINS complex and CDC45 to assemble an active CMG (Cdc45-MCM-GINS) helicase that initiates bidirectional DNA synthesis.1

    When DNA synthesis is completed, the CMG helicase is disassembled and unloaded from chromatin during replication termination.


    “A multitude of studies have demonstrated that the early phases of DNA replication are regulated by ubiquitylation.”

    For instance, the E3 ubiquitin ligase complexes cullin-RING ligase-1 (CRL1) and cullin-RING ligase-4 (CRL4) prevent re-replication and the occurrence of genome instability by targeting pre-RC components for proteasomal degradation (reviewed in Truong et al.2).

    Cullin-RING ligases (CRLs) constitute a protein family of 200 modular E3s and are composed of eight distinct subfamilies containing different cullins, namely CUL1, CUL2, CUL3, CUL4A, CUL4B, CUL5, CUL7 and CUL9.3

    Cullins work as molecular scaffolds assembling the different complex subunits, that is, a RING-finger protein (RBX1 or RBX2), which interacts with the ubiquitin-conjugating enzyme, an adaptor protein and one of many substrate-receptor subunits.

    The activity of CRLs is primarily controlled at the level of substrate recruitment.

    The direct recognition of the target protein by the substrate-receptor subunit and its recruitment to the core CRL platform are in fact regulated in response to specific stimuli.

    Moreover, all CRLs are activated through the covalent attachment of the ubiquitin-like protein Nedd8 to the cullin subunit.

    Figure 1 Replisome Unloading – 2 Pathways Including Backup Mechanism

    Replisome unloading is controlled by two pathways. (a) During DNA replication termination, the CRL2LRR1 ubiquitin ligase and the p97 segregase trigger replisome unloading from chromatin. (b) An additional backup mechanism that depends on the p97 adaptor UBXN3 drives replisome unloading from mitotic chromatin. See text for details. For the sake of clarity, the different components of the replisome are not shown.

    Gee Gpuccio,
    I’m guessing you had some idea just how far reaching the “Ubiquitin” System was, but it must still be amazing how much is unfolding today, before us in research across multi-discipline areas of disease, functions and applications.

  381. DATCG:

    “Gee Gpuccio,
    I’m guessing you had some idea just how far reaching the “Ubiquitin” System was, but it must still be amazing how much is unfolding today, before us in research across multi-discipline areas of disease, functions and applications.”

    You are perfectly right. While working at this OP and at the following discussion with you and the other friends, I have been constantly surprised and overwhelmed at the ever new complexity, scope and “omnipresence” in the cell of the molecular system I had chosen to study in some detail!

    I suppose that happens in some measure with all molecular systems in the cell, but this time the “measure” is really huge. 🙂

  382. So much to read here, and to catch up on. Excellent OP.

  383. DATCG:

    The Replisome is another huge subject, and it woul probably deserve an OP of its own. We’ll see. 🙂

    Certainly, it’s really surprising that such a basci function like DNA replication should be so different in eukaryotes as compared to prokaryotes. This is further confirmation of the all-round re-engineering that took place at the eukaryotes transition!

    Just as an example, the Mcm 2-7 heterohexamer ring which is an integral part of the CMG complex which serves as helicase to start DNA replication is made of 6 different proteins, Mcm 2-7, about 700 – 900 AAs long, all of them highly conserved in eukaryotes, which at sequence level share only modest homology between them (about 300 bits). Although one homolog is described in Archaea, it is almost completely different at sequence level.

    And this is just part of the starting complex! 🙂

  384. DATCG:

    Well, this is new, too. Memory formation.

    The Ubiquitin-Proteasome System and Memory: Moving Beyond Protein Degradation.

    http://journals.sagepub.com/do.....8418762317

    Abstract:

    Cellular models of memory formation have focused on the need for protein synthesis. Recently, evidence has emerged that protein degradation mediated by the ubiquitin-proteasome system (UPS) is also important for this process. This has led to revised cellular models of memory formation that focus on a balance between protein degradation and synthesis. However, protein degradation is only one function of the UPS. Studies using single-celled organisms have shown that non-proteolytic ubiquitin-proteasome signaling is involved in histone modifications and DNA methylation, suggesting that ubiquitin and the proteasome can regulate chromatin remodeling independent of protein degradation. Despite this evidence, the idea that the UPS is more than a protein degradation pathway has not been examined in the context of memory formation. In this article, we summarize recent findings implicating protein degradation in memory formation and discuss various ways in which both ubiquitin signaling and the proteasome could act independently to regulate epigenetic-mediated transcriptional processes necessary for learning-dependent synaptic plasticity. We conclude by proposing comprehensive models of how non-proteolytic functions of the UPS could work in concert to control epigenetic regulation of the cellular memory consolidation process, which will serve as a framework for future studies examining the role of the UPS in memory formation.

  385. DATCG:

    What about human Embryonic Stem Cells? A very hot topic, I would say.

    Insights into the ubiquitin-proteasome system of human embryonic stem cells

    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5840266/

    Abstract:

    Human embryonic stem cells (hESCs) exhibit high levels of proteasome activity, an intrinsic characteristic required for their self-renewal, pluripotency and differentiation. However, the mechanisms by which enhanced proteasome activity maintains hESC identity are only partially understood. Besides its essential role for the ability of hESCs to suppress misfolded protein aggregation, we hypothesize that enhanced proteasome activity could also be important to degrade endogenous regulatory factors. Since E3 ubiquitin ligases are responsible for substrate selection, we first define which E3 enzymes are increased in hESCs compared with their differentiated counterparts. Among them, we find HECT-domain E3 ligases such as HERC2 and UBE3A as well as several RING-domain E3s, including UBR7 and RNF181. Systematic characterization of their interactome suggests a link with hESC identity. Moreover, loss of distinct up-regulated E3s triggers significant changes at the transcriptome and proteome level of hESCs. However, these alterations do not dysregulate pluripotency markers and differentiation ability. On the contrary, global proteasome inhibition impairs diverse processes required for hESC identity, including protein synthesis, rRNA maturation, telomere maintenance and glycolytic metabolism. Thus, our data indicate that high proteasome activity is coupled with other determinant biological processes of hESC identity.

  386. #385 Upright BiPed,

    “So much to read here…”

    See you next year 😉

    At least for me it will take that long, not including a specialized degree and a lifetime of research. Amazing material covered in this OP by Gpuccio.

    It’s a large task for thousands of scientist 🙂 I knew this OP would be fun and expansive, but had no idea the naming of Ubiquitin was so on target 😉 Or the amount of networked regulatory systems content we would be reviewing.

    To go from one area of specialization to another invites an expanding vocabulary of specific terminology for critical systems interactions of Ubiquitin targeting, tagging, recycling, and/or degradation by UPS.

    Highly specialized researchers across so many areas of discipline are discovering fascinating areas of tightly controlled regulatory networks bound by or regulated by the UPS, DUBs, etc.

    If any of these “tightly controlled” regulatory systems experience “random mutations” or stress conditions with Ubiquitin inteactions, it leads to numerous diseases across a spectrum of human organs and networks with immune systems responses that depend upon Ubiquitin signal and recognition systems for conditions based processing.

    Whew…….

    Random mutations are the enemy, not the blind, unguided builder of such highly integrated, tightly regulated, Functionally Organized & Highly Coordinated Complex Systems.

    The material covered is an avalanche of specified information, overwhelming any highly trained team of scientist and lab techs to keep up with.

    Even specialist in their field must be experiencing an overload trying to keep up with latest research and discovery including epigenetic programs. Physicist are involved at level of Quantum Mechanics as well in inter-disciplinary talks on the Ubiquitin System.

    It’s an intellectual smorgasbord of regulatory systems identification and reverse-engineering. 😉

    OK, I just stated the obvious. But sometimes the obvious must be stated 😉

    But I’m sure, given enough time, Darwin Did It!

  387. #386 Gpuccio,

    oooohhhhh… a future OP, OK, that would be really cool to dissect the Replisome 🙂

    How many future OPs are you entertaining now? I know Dionisio listed several, plus you discussed Missing Procedures.

    Certainly, it’s really surprising that such a basci function like DNA replication should be so different in eukaryotes as compared to prokaryotes. This is further confirmation of the all-round re-engineering that took place at the eukaryotes transition!

    “… really surprising” Surprising for Darwinist or Design Theorist or both? I was thinking it might be expected considering the enormous amount of Epigenetic information and regulatory systems.

    Maybe, or would it be beneficial for a future OP on Jumps in Functional Complexity across systems from prokaryote to eukaryote transition? A Summary OP of Information Jumps, if you will allow such a description, based on your past OP research and any others you’d like to include.

    This is where I disagreed with Arthur Hunt and thought BLASTing information was critical in reviewing informational jumps. Why he was critical of it, remains a mystery. Perhaps because it hits close to home.

    I know you’ve covered this in other OPs and systems reviews. Really enjoyed learning that aspect of your OPs. Including learning to use BLAST for these type of searches.

  388. #387 Gpuccio,

    Another great find and aspect of UPS regulatory network. I’d looked briefly at other papers on Parkisons, Alzheimers and other diseases and roles of UPS and aggregate protein accumulations.

    That paper’s behind a paywall, but found some previous papers by the author at Research Gate. That paper is so new he’s not listed yet on his Bio Page! 😉

    Thought it interesting to look at his area of research as well.

    Dr. Timothy Jarome Bio – Research Area

    Research in the Jarome lab is focused on elucidating the cellular and molecular mechanisms of memory formation and storage, with an emphasis on understanding how stressful or traumatic events alters brain chemistry that drives future behavioral and physiological responses.

    These future responses are often maladaptive, resulting in a variety health concerns, and can be passed to future generations through “epigenetic” mechanisms.

    The lab focuses on mechanisms of initial memory storage and those involved in memory modification following retrieval (recall). Currently, the lab has several areas of interest:

    – An epigenetic role for the ubiquitin-proteasome system in fear memory formation

    – Epigenetic mechanisms of fear memory modification following retrieval

    To address these topics, we combine a traditional rodent behavioral paradigm (Pavlovian fear conditioning) with a variety of traditional and modern molecular biology and neuroscience techniques.

    This includes using in vivo pharmacology, siRNA-mediated gene knockdown, and CRIPSR-dCas9 transcriptional editing to manipulate specific genes and/or cellular processes during learning or memory retrieval and analyzing the effects of these manipulations on the cellular memory storage process using western blotting, qRT-PCR, chromatin immunoprecipitation, methylated DNA immunoprecipitation, bisulfite sequencing and other molecular biology methods.

    Students who join the lab will have the opportunity to learn these techniques and, as they advance, will have the opportunity to take projects in new directions or initiate new topics.

    So what’s happening here? Besides discovery of Ubiquitin and UPS mechanisms for regulatory role, we see once again Epigenetics roles emerging at the forefront of knowledge on disease control, including in this case Memory Storage and Retrieval, associated with FEAR complex, etc.

    I wish we had a way to measure all the latest Epigenetic Research and Discovery of Function in formerly labeled “JUNK” DNA zones by the Darwinist.

    Is it fair to say and borrow the term Ubiquitous for Epigenetic Roles? As in Epigenetic Roles are Ubiquitous through out Eukaryotes? Or, is that expanding the nature of the Epigenome to fast before evidence?

    This is a bit off-topic, but when we see a research scientist involved in this one area, of Ubiquitin and Epigenetic roles, then …

    1) We know Ubiquitin is Network Wide across all areas of Function and space in the Human genome

    2) We know of this area – Memory – and many others where Epigenetic regulatory roles work with Ubiquitin systems. Or as this scientist has stated, “an Epigenetic role for UPS” !!! 🙂

    Does it logically follow that Ubiquitin is fully dependent upon Epigenetic layers of meta-code to function in all these different areas covered so far in this one OP?

    Is there an areas where Ubiquitin functions without Epigenetic layers or Epigenetic regulatory systems involved?

    Just a thought. I’m not sure we can extrapolate this to mean that ENCODE 80% functionality claim is strengthened by this, but sure seems like a good, informed guess?

  389. Continued from #391 … offtopic a bit, but looking ahead.

    On possible links between Ubiqutin and Epigenetic roles – does this lead to decimation of Dan Graur’s argument for large amount of “Junk” DNA sill existing? According to his last paper? In his last retort he states at least 75% of our DNA must be JUNK.

    I’m not entertaining that the entire Genome is functional with Epigenomic regions.

    But, I do think this is a key area where the bell tolls for Neo-Darwinian evolutionist like Graur, who said, “If ENCODE is right, Evolution is wrong”

    If you look carefully at what he’s stated, much of it is still based upon neo-Darwinian assumptions. That were in the past based largely upon ignorance and today seems to be a stubborn adherence to antiquated beliefs.

    None of this includes recent projects in the last several years unfolding since ENCODE.

    4D Nucleome Network Project Overview at Nature published September 2017

    4D Nucleome Project Consortium North America

    4D Nucleome European Initiative

    1) Ubiquitin System Wide Role
    2) Epigenetic Role of UPS(Ubiquitin Proteome System)
    3) Epigenetic Research turns up new roles and fucntions every day
    4) 4D Nucleome Project will not help neo-Darwinist and can only hurt stubborn Darwinist like Graur

    From the EU initiative for 4D Nucleom research…

    Recent technological advances in high resolution and live microscopy, high-throughput genomics/cell biology approaches and modelling, coupled with increased awareness of the importance of genome organization will soon allow to perform precision analysis of our genomic organization and its dynamic translations from one epigenome to another, as cells differentiate, age, and respond to the environment.

    This a perfect time to launch a concerted effort towards characterizing the dynamic organization of the genome, the epigenome, and the rules that govern determination and maintenance of cell types in face of both internal and external stress linked to disease.

    We can now envisage having a complete 3D atlas in time (4D) of nuclei within the many cell types that form our body. The huge challenge before us is to take the one dimensional genome sequence provided by the Human Genome Project, decorated with the valuable annotations provided by the ENCODE project, and create an integrated 4D understanding of the complexity of this incredible, living, breathing machine that holds the secret of life.

    Semiosis, Rules, Meta-Layers of Code upon Code, Dynamic Post-Translation Modifications, Organization and Functional Networked Systems of Tightly Controlled, Interdependent Systems, Coordination and Coherence.

    To have a Rule, there must be a Rule-maker. Recognition that Rules exist in symbolic representations is by definition a teleological argument for Design.

    Otherwise, stop calling them Rules. Yet, they cannot stop doing so because there’s no other way to logically and coherently describe the process.

  390. #387 Gpuccio,

    back on topic Memory and UPS, a research review by Timothy Jarome in a paper he published 2014. Open Access.

    Ubiquitin Role in Long Term Memory Formation with Protein Degradation and Synthesis

    REVIEW ARTICLE
    Front. Mol. Neurosci., 26 June 2014 | https://doi.org/10.3389/fnmol.2014.00061

    Timothy J. Jarome and Fred J. Helmstetter

    Long-term memory (LTM) formation requires transient changes in the activity of intracellular signaling cascades that are thought to regulate new gene transcription and de novo protein synthesis in the brain. Consistent with this, protein synthesis inhibitors impair LTM for a variety of behavioral tasks when infused into the brain around the time of training or following memory retrieval, suggesting that protein synthesis is a critical step in LTM storage in the brain. However, evidence suggests that protein degradation mediated by the ubiquitin-proteasome system (UPS) may also be a critical regulator of LTM formation and stability following retrieval.

    This requirement for increased protein degradation has been shown in the same brain regions in which protein synthesis is required for LTM storage.

    (balance and trade-off of keeping memory retainable across time)

    Additionally, increases in the phosphorylation of proteins involved in translational control parallel increases in protein polyubiquitination and the increased demand for protein degradation is regulated by intracellular signaling molecules thought to regulate protein synthesis during LTM formation.

    (balance must be maintained again)

    In some cases inhibiting proteasome activity can rescue memory impairments that result from pharmacological blockade of protein synthesis, suggesting that protein degradation may control the requirement for protein synthesis during the memory storage process.

    Amazing, balance

    Results such as these suggest that protein degradation and synthesis are both critical for LTM formation and may interact to properly “consolidate” and store memories in the brain.

    Here, we review the evidence implicating protein synthesis and degradation in LTM storage and highlight the areas of overlap between these two opposing processes.

    (Opposing Processes = Balancing Act)

    We also discuss evidence suggesting these two processes may interact to properly form and store memories. LTM storage likely requires a coordinated regulation between protein degradation and synthesis at multiple sites in the mammalian brain.

    Amazing stuff, this seesaw of regulation, synthesis and degradation.

    Recently, attention has turned to the potential role of protein degradation in learning-dependent synaptic plasticity. Indeed, there is now convincing evidence that UPS-mediated protein degradation is likely involved in various different stages of memory storage.

    However, while some studies have suggested potential roles for protein degradation in long-term memory (LTM) formation and storage (Kaang and Choi, 2012), one intriguing question is whether protein degradation is linked to the well-known transcriptional and translational alterations thought to be critical for memory storage in the brain (Johansen et al., 2011).

    Here, we discuss evidence demonstrating a role for protein degradation and synthesis in the long-term storage of memories in the mammalian brain, highlighting instances in which a requirement for protein degradation correlates with a requirement for protein synthesis.

    Additionally, we discuss evidence suggesting that both protein degradation and synthesis may be regulated by CaMKII signaling during LTM formation. Collectively, we propose that LTM storage requires coordinated changes in protein degradation and synthesis in the brain, which may be primarily controlled through a CaMKII-dependent mechanism.

    Had not read down this far but oh so cool…

    What Comes First, Degradation or Synthesis?

    A majority of the studies discussed here reveal a strong correlation between protein degradation and synthesis during LTM formation.

    This leads to one important question: Which comes first?

    While the exact relationship between protein degradation and synthesis during memory formation currently remains equivocal, the available evidence suggests that protein degradation likely regulates protein synthesis.

    For example, fear conditioning leads to an increase in polyubiquitinated proteins being targeted for degradation by the proteasome (Jarome et al., 2011).

    While a majority of the proteins being targeted by the proteasome for degradation remain unknown, the RNAi-induced Silencing Complex (RISC) factor MOV10 has been identified as a target of the proteasome during increases in activity-dependent protein degradation in vitro (Banerjee et al., 2009) and following behavioral training and retrieval in vivo (Jarome et al., 2011).

    Increases in the degradation of MOV10 are associated with increased protein synthesis in vitro, suggesting that the proteasome could regulate protein synthesis during LTM formation through the removal of translational repressor proteins such as various RISC factors.

    However, it is currently unknown if the selective degradation of MOV10, or any RISC factor, is critical for memory formation in neurons. Nonetheless, studies such as these provide indirect evidence that protein degradation by the UPS could regulate protein synthesis during memory formation in the brain.

    The balancing act and pre-programmed responses to conditioning possibly exert factors on enhanced protein synthesis processing.

    Some of the best evidence that protein degradation may be upstream of protein synthesis during memory storage comes from studies examining memory reconsolidation following retrieval.

    For example, inhibiting proteasome activity can prevent the memory impairments that normally result from post-retrieval blockade of protein synthesis in the hippocampus, amygdala, and nucleus accumbens (Lee et al., 2008; Jarome et al., 2011; Ren et al., 2013) as well as during LTF in aplysia (Lee et al., 2012), suggesting that protein degradation is upstream of protein synthesis during memory reconsolidation.

    This remains some of the best evidence directly linking protein degradation to protein synthesis during memory storage, but it is possible that the rescue of memory impairments in the face of protein synthesis inhibition may occur as an indirect consequence of blocking protein degradation rather than a direct effector.

    Now, how is ubiquitin code and protocol utilized(or inhibited, modified) in Addictions, Drug Abuse or legal drugs for reducing pain?

    I’ll move this to a new comment.

  391. Part of Drug Abuse and Addiction is Conditioning. Habit forming over time. While certainly dealing with different aspects of neural development and different areas related to addiction, I’d now expect to find UB or UPS, and DUBS in some role.

    So, searched on UB and Addiction. And oddly enough found the Review Article below in the same journal…

    Roles of the ubiquitin proteasome system in the effects of drugs of abuse

    Front. Mol. Neurosci., 06 January 2015 | https://doi.org/10.3389/fnmol.2014.00099
    Nicolas Massaly, Bernard Francès and Lionel Moulédous

    What I did not expect immediately to see, but makes absolute sense is the overlapping of Memory in conjunction with addiction and UB.

    Because of its ability to regulate the abundance of selected proteins the ubiquitin proteasome system (UPS) plays an important role in neuronal and synaptic plasticity.

    As a result various stages of learning and memory depend on UPS activity. Drug addiction, another phenomenon that relies on neuroplasticity, shares molecular substrates with memory processes.

    However, the necessity of proteasome-dependent protein degradation for the development of addiction has been poorly studied. Here we first review evidences from the literature that drugs of abuse regulate the expression and activity of the UPS system in the brain.

    We then provide a list of proteins which have been shown to be targeted to the proteasome following drug treatment and could thus be involved in neuronal adaptations underlying behaviors associated with drug use and abuse. Finally we describe the few studies that addressed the need for UPS-dependent protein degradation in animal models of addiction-related behaviors.

    Interesting, regulation after Drug Exposure(B) in Figure 1

    UPS components regulated(see B) after Drug Exposure


    Figure 1. The Ubiquitin Proteasome System and its components regulated after drug exposure.

    (A) Schematic representation of the Ubiquitin Proteasome System. The external and internal rings constitute the 20S proteasome. The lid and base constitute the 19S regulatory complex. In some cases, it can be replaced by the PA28 or 11S regulatory complex, constituted of a single ring of 7 subunits.

    (B) Classification of the UPS components found to be regulated after drug exposure.

    All drugs of abuse can thus affect the expression and abundance of key UPS proteins.

    However, the data reported above are only descriptive. Moreover, UPS components are affected differently depending on the drug type, its method of administration, the duration of the treatment and the cell type or brain region considered (Table 1). Complementary studies have also found that drugs of abuse modify the activity of the UPS in parallel with changes in the expression of its various components.

    Indeed morphine was demonstrated to inhibit the activity of the 20S proteasome in human neuroblastoma cells, with neuroprotective consequences (Rambhia et al., 2005).

    On the contrary, PKC-dependent inhibition of the UPS was linked to the autophagy-mediated toxicity of methamphetamine in dopaminergic neurons (Lin et al., 2012).

    In addition it has been proposed that the higher toxicity of methamphetamine compared to cocaine was due to its long inhibitory effect on proteasome activity (Dietrich et al., 2005).

    Finally, a recent study demonstrated that chronic ethanol induces toxicity in mice through a Toll-like receptor 4-dependent impairment of the UPS (Pla et al., 2014).

    Balance between Protein Synthesis and Degradation again

    This deleterious effect of UPS blockade on long term changes in neurons has been suggested to be due to an alteration in the balance between protein synthesis and degradation (Fonseca et al., 2006). Indeed the authors showed that the deleterious effects produced by inhibiting either protein synthesis or degradation on LTP can be reversed by inhibition of the two processes at the same time.

    In addition to synaptic proteins the UPS is also involved in the regulation of the activity of transcription factors, thus revealing a close relationship between protein synthesis and proteasome action.

    For example I?B and CREM (cAMP-responsive element modulator), repressors of the transcription factors NF-?B and CREB (cAMP response element binding) respectively, can be ubiquitinated and degraded by the UPS (Woo et al., 2010; Liu and Chen, 2011).

    In that sense the UPS clearly plays a major role in the regulation of protein turnover implicated in neuronal plasticity acting directly through the degradation of some proteins and indirectly through the modulation of transcriptional activity and protein synthesis.

    Oh, the authors recognize Jarome, et al., previous work in UPS regulation and memory(LTM), but I’m moving on to another paragraph.

    The precise mechanisms underlying the involvement of the proteasome in memory are just beginning to be discovered but it is now clearly established that, in addition to protein synthesis, neuronal protein degradation by the UPS is a mandatory process to create, store and maintain memories and in that sense participates to adaptive behaviors of mammals.

    Since drug addiction shares common mechanisms with memory processes (Hyman et al., 2006; Milton and Everitt, 2012) it is important to question the role of the UPS in the long term effects of drugs of abuse such as opioids, stimulants, ethanol, nicotine and cannabinoids.

    Indeed, very interesting material.

    In the case of opioids, it was shown in a cellular model that a prolonged 72 h morphine treatment modifies the abundance of two proteasome subunits (?3 and ?6) (Neasta et al., 2006). In vivo, intra-cerebro-ventricular (icv) infusion of morphine for 72 h results in an increase in the tyrosine-phosphorylated form of the ?4 subunit in the rat frontal cerebral cortex (Kim et al., 2005).

    A longer intermittent treatment (2 weeks) produces a decrease in the amount of the DUB Ubiquitin C-terminal hydrolase L-1 in the nucleus accumbens (Nacc) (Li et al., 2006).

    4 days after morphine withdrawal, the quantity of this enzyme, as well as that of the ?3 subunit of the proteasome, increases in rat dorsal root ganglia (Li et al., 2009). Similarly, chronic treatment (90 days) and drug withdrawal have been shown to have opposite effects on the amount of ?5 subunit in the Nacc of rhesus monkeys (Bu et al., 2012).


    The levels of Ubiquitin-conjugating enzyme E2 and of Ubiquitin C-terminal hydrolase L-3 are also modulated in this model.

    Finally, in a morphine-induced conditioned place preference (CPP) paradigm which tests the rewarding properties of the drug, both development, extinction and re-instatement are accompanied by a down-regulation of several DUBs and ? and ? subunits (Lin et al., 2011).

    Hmmmm, now, what of legal pharmaceuticals? Not to beat up on Big Pharma, but what of psychotropic medicines intended for good that are suddenly removed from a patient?

    What happens with build up and changes in different areas?

    How does the brain and neural network balance after sudden removal, including UPS, DUBS and other regulatory systems involved at cognition, perception and memory?

    What remains unchanged? What is inherited Epigenetic changes passed down to offspring?


    Table 1. UPS-related molecular and cellular consequences of the treatment with drugs of abuse.

    That’s a load of UPS regulatory functions and consequences by drugs of abuse. Wonder if similar studies exist for legal drugs showing similar areas of changes and modifications for public access.

    .

  392. DATCG at #389:

    “Random mutations are the enemy, not the blind, unguided builder of such highly integrated, tightly regulated, Functionally Organized & Highly Coordinated Complex Systems.”

    Of course. The idea that a complex regulation network may arise from random mutations and natural selection is ridiculous!

    Especially if that network works by coded symbols, like the different types of signals implemented by ubiquitin chains.

    Especially if the network is made by hubdreds and hundreds of specific sub-networks.

    Especially is the network control not one, but tons of different complex functions, practically every function we can imagine.

    So, how is it that no one from the other field has writeen one sinlge word here to try to explain how random variation and natural selection can do this? 🙂

    “The material covered is an avalanche of specified information, overwhelming any highly trained team of scientist and lab techs to keep up with.”

    You bet! Just the 600+ E3 ligases are an example of thousands and thousands, maybe hundreds of thousands, of functional bits whose function is to recognize all the specific target proteins, thousands of them, and tag them in the correct way in each appropriate condition.

    Let’s remember that about 5% of the whole protein coding genome is implied in the ubuquitin network!

    “OK, I just stated the obvious. But sometimes the obvious must be stated”

    Absolutely! 🙂

    When no one seems to have the courage to deny the absurd, stating the obvious is probably the only salvation. 🙂

  393. DATCG at #390:

    “How many future OPs are you entertaining now? ”

    Indeed, I am thinking about 3 or 4 different possibilities. In the end, I will probably follow some sudden “inspiration”! 🙂

    The “prokaryote to eukaryote transition” is a fascinating issue. What a pity that we have no precise idea of when it happened, and least of all a reasonable early tree of eukaryotes!

    I think that both my OP on the spliceosome and this one about ubiquitin are good examples of highly specific eukaryotic machinery. But of course, there are many others! 🙂

    “This is where I disagreed with Arthur Hunt and thought BLASTing information was critical in reviewing informational jumps. Why he was critical of it, remains a mystery. Perhaps because it hits close to home.”

    I disagree with Arthur Hunt too, as much as it is possible to disagree with someone who has not really expressed his thoughts. 🙂

    BLAST is a wonderful tool for us IDists. Neo-darwinists use it mostly to find vague distant homologies. But we can and do use it to detect functional information, which is much more interesting!

    OK, it’s late now here in Italy. I will come back tomorrow.

  394. Simply outstanding work by the two of you. I’m trying to catch up, but it seems almost impossible. Great job.

  395. Gpuccio @395

    “So, how is it that no one from the other field has writeen one sinlge word here to try to explain how random variation and natural selection can do this?”

    Good question, where are the neo-Darwinist?

    I suspect they stay away for purposes of not making your excellent OPs legitimate – as in recognized not only by themselves, but in eyes of their own followers.

    Think of it. If they actually engage you – they can lose. And their followers might see your logic as correct.

    They cannot bear that possible outcome.

    And I suspect Hunt got a hint to back away. I could be wrong, but am surprised he’d back off for any other reason.

    Surely he can mount a defense of group II introns and spliceasome evolution, right?

    By not engaging, they hope Intelligent Design goes away. It’s not, it’s only growing. And more bright minds are learning every day a new way of seeing life as a result of Design.

    Discovery Institute Summer Seminars on Intelligent Design July 6-14

    .

  396. Walking thru time on the 4D Nucleome Project, I ventured out a bit to see if I could find Ubiquitin involvement in different areas.

    Here’s an interesting related area to review…

    Drosophila Casein Kinase I Alpha Regulates Homolog Pairing and Genome Organization by Modulating Condensin II Subunit Cap-H2 Levels
    PLoS Genet. 2015 Feb;11(2): e1005014.Published online 2015 Feb 27. doi: 10.1371/journal.pgen.1005014

    Huy Q. Nguyen, Jonathan Nye, Daniel W. Buster, Joseph E. Klebba, Gregory C. Rogers,and Giovanni Bosco,
    R. Scott Hawley, Editor

    Abstract

    The spatial organization of chromosomes within interphase nuclei is important for gene expression and epigenetic inheritance.

    Although the extent of physical interaction between chromosomes and their degree of compaction varies during development and between different cell-types, it is unclear how regulation of chromosome interactions and compaction relate to spatial organization of genomes. Drosophila is an excellent model system for studying chromosomal interactions including homolog pairing.

    Recent work has shown that condensin II governs both interphase chromosome compaction and homolog pairing and condensin II activity is controlled by the turnover of its regulatory subunit Cap-H2.

    Specifically, Cap-H2 is a target of the SCFSlimb E3 ubiquitin-ligase which down-regulates Cap-H2 in order to maintain homologous chromosome pairing, chromosome length and proper nuclear organization.

    Here, we identify Casein Kinase I alpha(CK1-alpha) as an additional negative-regulator of Cap-H2. CK1alpha-depletion stabilizes Cap-H2 protein and results in an accumulation of Cap-H2 on chromosomes. Similar to Slimb mutation, CK1alpha depletion in cultured cells, larval salivary gland, and nurse cells results in several condensin II-dependent phenotypes including dispersal of centromeres, interphase chromosome compaction, and chromosome unpairing.

    Moreover, CK1alpha loss-of-function mutations dominantly suppress condensin II mutant phenotypes in vivo. Thus, CK1alpha facilitates Cap-H2 destruction and modulates nuclear organization by attenuating chromatin localized Cap-H2 protein.

    Introduction

    Interphase genome organization in eukaryotic cells is non-random [1,2,3].

    Indeed, organization of the genome is crucial because it influences nuclear shape and processes such as DNA repair and replication, as well as gene expression [4, 5, 6].

    While chromosomes are highly organized within the nucleus, they must also remain extremely dynamic. Chromosome dynamics facilitate events that occur not only during cell division, but also during interphase, when cells respond to developmental and environmental cues that require changes in gene expression.

    Interphase events include trans-interactions such as homolog pairing, chromosome remodeling and compaction, and DNA looping. Although numerous studies using Fluorescent In-Situ Hybridization (FISH), live cell imaging, and chromosome conformation capture techniques have revealed the three-dimensional (3D) organization of genomes, much remains to be discovered regarding the factors that govern the overall conformation of interphase chromosomes. An equally important task is to identify the molecular mechanisms that regulate and maintain specific 3D genome organizational states.

    Condensin complexes are highly conserved from bacteria to humans [7,8,9] and have been identified as key drivers of genome organization [10].

    Eukaryotes have two condensin complexes, condensin I and II, which share the core SMC2 and SMC4 (Structural Maintenance of Chromosomes) subunits but differ in their non-SMC Chromosome Associated Protein (CAP) subunits. Condensins have long been known to play vital roles in shaping mitotic chromosomes.

    While condensin I promotes lateral chromosome compaction, condensin II promotes axial compaction; both of which are necessary for faithful mitotic condensation and chromosome segregation [11]. Condensins also display different localization patterns: condensin I only associates with mitotic chromosomes, whereas condensin II is present in the nucleus, where it is bound to chromatin throughout the cell cycle [12,13,14,15]

    Fascinating. So where Ubiquitin? Here, Slimb E3 ligase…

    Ubiquitin E3 coordination with Phosphorlation and Degradation

    note: ? = alpha subunit

    Moreover, Cap-H2 protein levels are controlled by the SCFSlimb ubiquitin-ligase, maintaining low levels of Cap-H2 in vivo and in cultured Drosophila cells [20].

    Interestingly, Slimb(E3) recognizes its target proteins through a phosphodegron motif [29], suggesting that one or more kinases must phosphorylate Cap-H2 before Slimb can target it for destruction.

    A Slimb-binding site consensus sequence (DSGXXS) exists in the extreme C-terminus of Cap-H2 and deletion of this region renders Cap-H2 non-degradable [20].

    As expected for a Slimb substrate, Cap-H2 protein mobility on SDS-PAGE was sensitive to phosphatase treatment, suggesting that Cap-H2 is phosphorylated [20].

    Given that Cap-H2 protein levels may be regulated by its phosphorylation state, we set out to identify kinases that target Cap-H2 for Slimb recognition and that lead to its degradation.

    We show that in Drosophila cultured S2 cells, Casein Kinase I alpha (CK1?) depletion results in the hypercondensation of interphase chromatin in a condensin II-dependent manner.

    We also found that CK1? and condensin II genetically interact in vivo, and that CK1? depletion leads to Cap-H2 protein enrichment on polytene and cultured cell chromosomes.

    Similar to Slimb(E3) depletion [20], CK1? depletion also results in stabilization of Cap-H2 protein in cultured cells. Our findings further elucidate the mechanism by which Cap-H2, and thus condensin II, is regulated and contribute significantly to our understanding of how interphase genome organization, homolog pairing, and chromosome compaction is modulated.

    Results
    Casein Kinase I alpha is required for interphase chromatin reorganization

    Previously, we discovered that the Cap-H2 subunit of condensin II is a SCFSlimb ubiquitination-target in Drosophila cells [20].

    In a whole genome RNAi screen, Slimb was also identified as a homolog pairing-promoting factor, and it was shown to affect pairing in a Cap-H2 dependent manner[18].

    In cultured S2 and Kc cells, depletion of SCFSlimb components Slimb, Cul-1 and SkpA prevents Cap-H2 degradation and leads to condensin II hyperactivation during interphase and the remodeling of each chromosome into a compact globular structure (Fig. 1A-C). Based on their overall appearance, we refer to these hypercondensed chromosomes as “chromatin-gumballs” (Fig. 1A).

    Overexpression of a GFP tagged wild type Cap-H2 also induces this phenotype [20].

    Since phosphorylation of the Slimb-binding domain within its substrates is required for Slimb binding [29], we reasoned that depletion of a kinase involved in this pathway would also stabilize Cap-H2 and phenocopy the effect on chromatin remodeling observed after Slimb depletion.

    CK1alpha Highly Conserved Kinase

    CK1?=CK1alpha

    CK1? is a highly conserved serine/threonine kinase involved in Wnt signaling pathways, DNA repair, cell cycle progression, and mRNA metabolism [35,47,48]. Identification of CK1? furthers our understanding of the mechanisms by which condensin II is regulated.

    The chromodomain protein Mrg15 is involved in the loading of Cap-H2, while the E3 Ubiquitin ligase, SCFSlimb ubiquitylates Cap-H2, removing it from chromatin and targeting it for proteasomal degradation [20,21]. Phosphorylation is known to be a prerequisite for Slimb recognition of its target proteins

    Q: if CK1alpha is highly conserved, then is UBLigase-E3 Slimb highly conserved with it?

    It is tempting to speculate that cytokine signaling could trigger the activation of a condensin II antagonist, leading to the decrease in condensin II activity.

    This would lead to decondensation of chromatin allowing STAT5 access to DNA.

    Our findings in the Drosophila model suggest that similar interphase condensin II functions may be at play, and CK1? along with Slimb are critical regulators of this condensin II activity.

    However, at present it is not known if mammalian condensin II activity is regulated by Slimb or CK1?, and it should be noted that mouse and human Cap-H2 do not have clear Slimb binding consensus sequences.

    It will be of great value to identify additional kinases that may collaborate with CK1? and Slimb to negatively regulate Drosophila condensin II activity, and to further elucidate the biological significance of this interphase condensin II function in Drosophila and other species.

    OK, so Condensin II, next up a cool video. Packaging Pathways.

    .


  397. Packing a Genome, Step-by-Step – Condensin II

    Just to cool.

    While the video or “steps” do not say it. Somewhere, where there’s regulation, there’s Ubiquitination.
    Remember Condensin II and E3-ligase Slimb regulation in coordination with phosphorlation.

    – Ooops @399, sorry – missed a closing Bold Font highlight.

  398. Especially if that network works by coded symbols, like the different types of signals implemented by ubiquitin chains.

    Especially if the network is made by hundreds and hundreds of specific sub-networks.

    Especially if the network controls not one, but tons of different complex functions, practically every function we can imagine.

    Terrifying — if it’s your job to make sure there are never enough dissenters, that they might change the paradigm. 🙂

    Going to need some extra dogma. I suspect some shame, threats, and group enforcement will come in handy as well.

  399. DATCG at #391:

    “Does it logically follow that Ubiquitin is fully dependent upon Epigenetic layers of meta-code to function in all these different areas covered so far in this one OP?”

    Well, probably almost everything is under the control of epigenetic layers, because all transcription is fully dependent upon them. But ubuquitin has a definite role on epigenetic layers, as shown for example by its many roles with histones.

    It’s not a cse that we find ever more often the term “cross-talk” in biological papers. One thing is astonishingly clear: the cell has many, many independent layers of regulations, and all of them are constantly influencing one another and exchanging information. I think this is unprecedented, even in human programming and engineering.

  400. DATCG at #392:

    “If ENCODE is right, Evolution is wrong”

    What a pity! If it had been the other way round:

    “If Evolution is wrong, ENCODE is right”

    we could be certain that ENCODE is right! 🙂

    However, I agree with you that maybe some non coding DNA could be non functional, but it’s certainly not 75%!

    This 4D Nucleome Project is extremely interesting. I am sure that the 3D dynamic structure of chromatin and its constant modifications in time are one of the most important keys to understand something which goes beyond a mere accumulation od fetails.

    Hi-C (and its variants) is a really promising technique. The real primary aim is to understand how TFs work, their combinatorial nature, their ability to form chromatin loops and to connect distant parts of the genome in functional complexes.

    At that level, we are really just beginning to understand things.

  401. Upright BiPed at # 397:

    “I’m trying to catch up, but it seems almost impossible.”

    I can understand you. Sometimes it seems almost impossible to me to catch up with myself! 🙂

    However, it seems that adding DATCG to myself works combinatorially in fully unexpected ways. The results are really scary! 🙂

  402. #401-403 Gpuccio,

    Thanks, that illuminates the field of Epigenetics.
    I’d not considered Transcription’s dependence on Epigenetic factors.

    Then there’s so many other networks dependent upon Epigenetics.

    The 75% threshold by Dan Graur was precarious from the start. But I think he built that artificial wall based upon what he must have for neo-Darwinian faith to continue. By creating this in his anger and stubborn attitude he’s erected what appears to be a Humpty Dumpty Wall made of cards.

    and…

    “If Evolution is wrong, ENCODE is right”

    ah 🙂 Now you have juxtaposed a good turnabout is fair play.

    Dan and his mirror… as ENCODE proceeds and non-coded regions are explored with new functions found every day around the world.

    Mirror, mirror on the wall, who is right after all?
    Is it Darwin, is it Dan? Do unguided lots make up plans?
    Mirror, mirror on the wall, is neo-Darwinism due to fall?
    Humpty Dumpty Darwin’s game, does blind search fall in shame?

  403. #404 Gpuccio,

    and Dionisio contributions add even more! I’ve not come close to reviewing all of his papers! Simply not enough time.

    It’s a cornucopia of Ubiquitin fruit 😉

  404. GP at 404

    Agreed — and Dio’s contributions as well.

    I don’t know about this DATCG character. We’ll have to keep our eyes on him/her. He/she appears to be exceptionally bright. Clearly not his/her first rodeo in this area.

    🙂 🙂 🙂

  405. Upright BiPed:

    All of you, DATCG, Dionisio and you, have given great contributions! 🙂

    I was just focusing on DATG because his comments have become really prominent in the last phase of the discussion…

  406. DATCG at #393:

    Fascinating facts about long term memory.

    I think that this field is in great expansion, and maybe we can see something more specific in a short time.

    The working of the brain and nervous system is probably to be explained, as far as that is possible, at two different levels:

    a) The network of connections between neurons (and other cell types). This is amazing, if we think that we have about 10^11 neurons, and maybe 10^15 neuron connections. Those are big numbers, indeed. Any expert in hardware and software engineering knows all to well how important it is to have the right connections. And neuronal connections are dynamic, they can change and be rewired.

    b) But even more amazing is the biochemical plasticity of all that happens in neurons, and especially in synapses. And out friend ubiquitin, as you have shown, is critically linked to all this.

    I think that the only luck for neo-darwinists as far as the central nervous system is concerned is that we really understand too litlle of how it works. At least for the moment.

  407. DATCG at #393:

    Nice stuff about ubiquitin and addiction.

    By the way, the linked Table 1 is really amazing! 🙂

  408. DATCG at #399:

    “Q: if CK1alpha is highly conserved, then is UBLigase-E3 Slimb highly conserved with it?”

    Yes they are both highly conserved in metazoa.

    CK1alpha (337 AAs): the human protein shows 71% identities and 84% positives in Fungi (483 bits, 1.433 baa), and reaches a practically complete homology (99% identities, 100% positives) in Cartilaginous Fish. Amazing!

    Slimb (542 AAs) is just a little “slower”: 40% idenitites and 62% positives in Fungi, 91% and 93% in Cartilaginous Fish.

    Fascinating data about the condensin complexes. This issue of chromosome and chromatin structure certainly deserves some in-depth analysis. Let’s put it among our future plans! 🙂

  409. #411 Gpuccio,

    re: CK1alpha and Slimb

    Thanks!

    It seems UB E3 Slimb being a little “slower” makes sense due to species specific needs? The pesky human brain for example?

    On Condensins, Chromosomes, and beautiful DNA packaging and compression…

    “Let’s put it among our future plans!”

    Ahaha! 🙂 Your list is growing Gpuccio!

  410. #410 and #409 Gpuccio,

    Yes! It’s fascinating “stuff” our neural capacity, plasticity and change factors. Including all forms of stress, embryonic development and formation. Long-term memory, epigenetic regulatory systems and well, our little ubiquitin friends.

    “I think that this field is in great expansion, and maybe we can see something more specific in a short time.”

    I agree! Very interested in this area. Hope to devote more time to study these areas in future.

    I’m encouraged by the advancements in research being made at a rapid pace.

  411. #407 UB,

    First, I may have to call the Evergreen SJ Warriors on you for limiting me to a Binary choice Upright Bi-Ped! In fact, did you know many Uprights today consider themselves to be Poly-Peds! Oh my gosh, we need a meeting to review this and a sit in of UD! All Poly-Peds heed the call!

    I’m a guy. 😉
    and I’m stumbling through code as usual 😉

    OK, hope this does not bore you guys,

    I have a background in debugging production problems for large-scale, enterprise solutions years ago.

    Included print stream translations, hexadecimal to binary, etc., EBCDIC, ASCII, IBM, Xerox MetaCode, HP, and a myriad other solutions in imaging technology.

    After multiple input files were reformatted, merged, and post-processed, they were distributed to a multitude of different print streams, bar-coding, more post-processing and shipment, or imaging archival and viewing solutions for our clients.

    In the debugging process, often we had to go through reams of client code and reproduce it. Including bit and byte analysis of imaging and print streams to determine faults locally or upstream.

    As a result of client growth and expansion of many different streaming conventions, I developed a series of steps and solutions to quicken the debugging process.

    Much like the researchers, we might do “knock-outs” or obviously tracking and dumps, etc. We had 24hr windows to turnaround production for our clients.

    I formalized debugging solutions in a series of visual diagrams and checkpoints and gave out to our clients in technical presentations. Found out a decade later they were still using it.

    I guess that background helps. But not much different than most programmers would experience in debugging solutions.

    But mainly I’m fascinated how these cellular processes all work. And believe the Design heuristic holds the most promise going forward. And I’m a bit driven to find out how so much of it works together in such highly coordinated fashion.

    Every research paper we’ve seen posted here shows typical debugging steps to find problems in a multitude of Codes and branching steps or interactions. And researchers are getting better at debugging the different codes so to speak.

    Not to trivialize Life to much, but we are a collection of functional groupings of input/output steps, right?

    As an example, we can think of our skin as the finalized output of input and a cellular process to reformat the input.

    Therefore to know why carcinoma may exist in various forms of skin cells, we must know the cellular steps of:

    Input Processing -> Variable Data Reformatting Process of Organic Life -> Output Processing

    Input: Digestive, Environmental, Solar, etc.
    VDRPOL: Digestive Track, etc.
    Output: cellular function of skin cell production, reproduction and repair mechanisms

    I’m obviously leaving out a lot of steps and communication. This is way over simplification, but if in fact we are designed, it’s how I look at it from a design perspective.

    Start with Simplification of Top Down Structured thinking, then go to each branch, sub-branch, sub, sub, sub and loop backs. Throw in Modular concepts and OOP, networking communications, translations and/or transcribing and Post-Translation Modifications, etc., etc.

    But, the DNA code can be read backwards and forwards – come on! 🙂 LOL! I mean, the compression algorithms blow away anything today by modern methods. Amazing stuff!

    Now, expand by how many Input/Output Cellular Processes there are? I mean literally, you can find Ubiquitin’s role in the Gut 😉 I was going to post a research paper on Gut and Ubiquitin processing earlier, but ran out of time.

    I am enjoying this! Studying molecular biology and cellular processes. Deciphering Codes and OPs like this by Gpuccio!

    But I’m stumbling through it. Thankfully Gpuccio is patient.
    Thanks Gpuccio 🙂

    It’s been years since chemistry and genetics courses in undergrad. There’s so much I’m having to relearn. I switched from mechanical engineering to CS and left any trace of biochemistry and genetics behind.

    But I’ve always loved this area of scientific research.

    Going through massive amount of new terminology on these different Ubiquitin interactions is reminding me just how much I do enjoy it! 🙂

    Have a great weekend guys.

  412. I did not get this post-edit in place for 414…

    Regulatory Network of Epigenetic Processes oversees the following:

    Input: Digestive, Environmental, Solar, etc.
    VDRPOL: Digestive Tract, etc.(note: correction)
    Output: cellular function of skin cell production, reproduction and repair mechanisms

    Which in this OP includes the mighty Ubiquitin System 🙂

    And anyone who can think programmatically through of an input/output process understands that regulatory systems usually dwarf the core process to insure the core process never stops running.

    We are essentially a bunch of highly regulated, walking, talking consciousness of Organic Variable Data Reformatters 😉

    As are plants, trees, leaves, reformatting photons and CO2 for growth and structure, etc.

  413. DATCG at #414

    Beautiful.

    – – – – – – – – – – – – – – –

    (I knew this wasn’t your first rodeo in this area)

    Thanks for sharing.

  414. DATCG at #414:

    Thank you for sharing some of your background. You have really done a lot of brilliant work! 🙂

    I think you share some personal history with Dionisio, in terms of coming from Information Technology but having a deep love and understanding of biology.

    I have followed some different, but in a way specular, path, coming from medicine (and therefore, indirectly, biology) but having always loved, and in some way practiced, informatics. My good experience in medical data analysis and statistics has certainly helped too.

    I think that ID, as a new and revolutionary scientific paradigm, is specially attracting to people like us, who in some way have an interdisciplinary attitude. Maybe it’s also easier for us to be less conditioned by academic dogmas.

    Another thing that, IMO, unites people like you and UB and Dionisio and me is a genuine enthusiasm for ID as a scientific enterprise. I believe that, whatever our personal worldviews, we feel no particular need to overlap our more general beliefs with our scientific approach to facts.

    Whatever the reasons, I think we make a great team! 🙂

  415. DATCG at #400:

    Great video!

    And here are the last two papers published by the 4D Nucleome Project:

    A pathway for mitotic chromosome formation

    http://science.sciencemag.org/.....6135?rss=1

    Abstract:

    Mitotic chromosomes fold as compact arrays of chromatin loops. To identify the pathway of mitotic chromosome formation, we combined imaging and Hi-C of synchronous DT40 cell cultures with polymer simulations. We show that in prophase, the interphase organization is rapidly lost in a condensin-dependent manner and arrays of consecutive 60 kb loops are formed. During prometaphase ~80 kb inner loops are nested within ~400 kb outer loops. The loop array acquires a helical arrangement with consecutive loops emanating from a central spiral-staircase condensin scaffold. The size of helical turns progressively increases during prometaphase to ~12 Mb. Acute depletion of condensin I or II shows that nested loops form by differential action of the two condensins while condensin II is required for helical winding.

    and:

    Real-time imaging of DNA loop extrusion by condensin

    http://science.sciencemag.org/.....r7831.long

    Abstract:

    It has been hypothesized that Structural Maintenance of Chromosomes (SMC) protein complexes such as condensin and cohesin spatially organize chromosomes by extruding DNA into large loops. Here, we provide unambiguous evidence for loop extrusion by directly visualizing the formation and processive extension of DNA loops by yeast condensin in real-time. We find that a single condensin complex is able to extrude tens of kilobase pairs of DNA at a force-dependent speed of up to 1,500 base pairs per second, using the energy of ATP hydrolysis. Condensin-induced loop extrusion is strictly asymmetric, which demonstrates that condensin anchors onto DNA and reels it in from only one side. Active DNA loop extrusion by SMC complexes may provide the universal unifying principle for genome organization.

    Great work for condensins! 🙂

  416. GPuccio

    Just quickly passing by. Another great bookmark in the browser! I will try to read this OP as soon as I find time. I appreciate your efforts in laying out really hard-code ID stuff.

    I still would like to propose that readers have access to OPs by author on this blog! It would just be a lot more convenient!

    Upright Biped

    You have mail 🙂

  417. Hello Evgeny!

    Good to see you back on UD. With your interest in semiosis, you will certainly enjoy this thread.

    hmm … I do not see any mail !?

  418. Upright Biped, Gpuccio,

    Thanks guys, enjoy learning from you both!

    Gpuccio, your background in medicine helps so much. And I like your detailed analysis.

    This Ubiquitin post of yours sparked my interest more than usual.

    Precisely because it’s a Tagging system. Or Markup Language as an analogy?

    Most of what I worked on dealt with conditional processing of language specific identifiers, imaging systems, document management and packaging.

    Input Processing, Tagging and Rules based systems were created to coordinate a tightly controlled decision tree of subroutines. Built on specific language requirements used across all 50 states. All of it controlled by municipalities, medical boards, state and federal regulations. Lots of legalese w/ medical and beneficiary enrollment plans – healthcare.

    For large corporations and government it was a lengthy process and a possible legal nightmare if a single mistake(mutation) was made.

    Every decision made revolved around Tagging and Rules Based language procedures for Identification and Information processing routines.

    It was seen as fairly revolutionary at the time by a small software startup. What traditionally took six months to a year, even two years in large cases were reduced to mere days or weeks(and that only due to Human reviews). Simpler applications reduced it to mere seconds for request to end users.

    We were doing Markup Language and Tagging before HTML was fully accepted. Looking back realized I worked with some of the brightest in the industry. Visionary developers at that time. Was a great experience.

    These were legacy systems that eventually crossed over to PC and Browsers. Fortunately, I was chosen to bridge the gap for a few clients. So I learned many different platforms and markup languages over the years.

    OK, all this to explain my interest in this post Gpuccio.

    Going back to your OP, you identified Ubiquitin as a Tagging Solution.

    Precisely! In my view this Tagging solution and “Markup Language” or Ubiquitin Code Identification leaped out as Design from the start. Functional information processing systems or Cellular Processing cannot exist without Identification and Tagging or Marking Codes.

    A quick review of your initial post(note: edited)

    The semiosis: the ubiquitin code

    The title of this OP makes explicit reference to semiosis. Let’s try to see why.

    The simplest way to say it is: ubiquitin is a tag. The addition of ubiquitin to a substrate protein marks that protein for specific fates, the most common being degradation by the proteasome.

    Nonproteolytic Functions of Ubiquitin in Cell Signaling

    Abstract:

    In the past few years…, nonproteolytic functions of ubiquitin have been uncovered at a rapid pace. These functions include <b<(Tagging of:) membrane trafficking, protein kinase activation, DNA repair, and chromatin dynamics.

    A common mechanism(Tagging) underlying these functions is that ubiquitin, or polyubiquitin chains, serves as a signal to recruit proteins harboring ubiquitin-binding domains, thereby bringing together ubiquitinated proteins and ubiquitin receptors to execute specific biological functions.

    Another important aspect is that ubiquitin is not one tag, but rather a collection of different tags. IOWs, a tag based code.

    Bingo 🙂 a “Tag Based Code” Or, Markup Language Identifier?

    One area of clarification. Gpuccio you and Dionisio previously highlighted and discussed the missing procedures? Where are the governing Rules and procedures?

    Thinking from a Design interpretation.

    If we look at it through combination of instantiated information processing(substrates, Tags and Markings), we see a series of different Markup Languages and Tagging Identifiers. Mono, Poly-ubiquitin and branch Ubiquitin, etc.

    Can we designate these as external Tags(markups)?

    Based upon an information tagging processes being researched and discovered today of regulatory UB systems.

    But, what is missing? Might it be an internal Rules based, Tagging system?

    Back to Language Markup principles and Design. We used internal Markup Languages and Rules for identification of external tag markers, conditions based or Contextual and systems based, including transformation across different coded networks and languages.

    We internally marked(Tagged) every bit of language in whole document pagkages by Document IDs, pages, sections, paragraphs, down to single words, even characters, and internal translations, including special post-processing moditifications. All of this modular packaging sytems clients could pick and choose for whatever best represented their requirements.

    There was external Markup Language that End Users edited documents with, as identifiers to internal systems processing and tagging for eventual output destinations and other decision-required processing.

    All of it regulated by internal identifiers – Tags and Procedures only Developers could change or update. Overtime we allowed more overrides by customers to speed-up client specializations and less dependency on Developers. A more Open based User friendly markup.

    Q: Are we are on a similar threshold looking in as End Users today across cellular processes?

    Am I applying to many Information Process techniques of Markup languages and Tags to the Ubiquitin System? Does the analogy or application of Markup Language(Tags) make some rudimentary common sense? Or go to far afield from what you guys may be thinking?

    Can we state for example,

    For a Semiotic Code of Life to be interpreted and appropriate responses and actions to take place of any kind, it requires a Rules based, or Procedural Markup language? Both external and internal?

    For many different facets of:
    1) Input
    2) Identification, Tagging
    3) Procedures and Rules based calls
    4) Functional Operations
    – subsets of functions
    5) Interactions and Communications(Bridges) between Functions, Systems and network subsystems.
    6) Error Checking, Maintenance, Stress Management
    7) Final output or result

    Reflecting on Gpuccio’s posting at #369,

    Ube2V2 Is a Rosetta Stone Bridging Redox and Ubiquitin Codes, Coordinating DNA Damage Responses.

    On the scale of Life we see Semiosis. Multiple Codes and “Markup Languages.” MetaCodes, MetaLayers, and “forms” and/or Cellular processing techniques are conserved across millions of years in evolutionary terms, while some are plastic and vary across phyla, kingdoms and domains: Bacteria, Archaea, Eukaryota .

    This returns to Epigenetic Regulatory Code of Life. It seems safe to say, it’s larger than the blueprint itself upon which all core systems processing turns.

    Like any functionally organized, complex system, the blueprint must adhere to a large network of regulatory functions for initial design and importantly future maintenance.

    Poor Dan Graur, as Gpuccio rightly pointed out and turned Graur’s words back on him:

    “If Evolution is wrong, ENCODE is right”

    I wish there was a way to track the artificial boundary set by Dan’s 75% threshold.

    I wonder if ENCODE project is tracking the areas and numbers of Functions, including percentages of formerly declared “JUNK” DNA regions that today show important, tightly controlled functions.

    Since they laid out 80%, it would be in their interest to do so in comparison to Graur’s dogmatic response.

  419. EugeneS:

    Nice to see you here! 🙂

    I hope you will like the discussion about our friend ubiquitin. It has been much wider than each of us expected.

    And, of course, any comments from you will be greatly appreciated! 🙂

  420. DATCG at #421:

    Wow! What a tour the force!

    As usual, it is late now. I will comment on it tomorrow! 🙂

  421. ES, I do not see any mail, my friend.

  422. DATCG at #421:

    “This Ubiquitin post of yours sparked my interest more than usual.”

    Mine too, I must say.

    “Precisely because it’s a Tagging system. Or Markup Language as an analogy?”

    Exactly. The tagging/markup analogy is perfectly justified!

    I would like to add a few thoughts about the nature of the tag.

    Of course, ubiquitin is not the only “tag” here. We have seen that many other systems cooperate, and some of them just give the appropriate signal to the ubiquitin system.

    One good example is phosphorylation, which often serves as a tag to recruit the ubiquitin system to some specific target. So, we have a double specificity here: the ubiquitin system recognizes the target (usually by the E3 ligase), and it also recognizes the tag (phosphorylation). A good example of that mechanism can be found in the OP, where it is mentioned that phosphorylation of I?B? at serines 32 and 36 is the signal for the ubiquitination of the IkB alpha inhibitor (See Fig. 6).

    OK, so what is the main difference between, say, the phosphorylation tag and the ubiquitin tag?

    I would say that it is the fact that ubiquitin is a collection of different tags: IOWs, the system is much richer.

    Phosphorylation is a very powerful tag, but it is one tag, and therefore its symbolic meaning is linked essentially to the positions that are phosporylated in the target porteoin. For example, serines 32 and 36 inb the case of I?B?.

    The same would be true for ubiquitin if only mono-ubiquitination, single or multiple, existed. Then we would have one tag, whcih can assume different meanings according to the positions where it is added.

    But, as we well know, things are much more complex for ubiquitin. Much of the signaling, here, is made not ny mono-ubiquitination, but by ubiquitin chains. So, while the position where the chian is added retains all its symbolic meaning, a new layer of coding is added: the length and nature of the chain.

    In that sense, ubiquitin is really a miraculous peotein. Its special fold provides 8 different switches that can be used to build chains. So we have the following combinatorial degrees of freedom:

    a) The length of the chain can vary

    b) Homogeneous chains can be buith using each of the possible switches.

    c) Heterogeneous chains can be built by mixing different switches.

    That’s simply outstanding! 🙂

  423. DATCG:

    By the way, have you seen that our private party here has got a lot of new visibility, thanks to this kind OP by Barry Arrington? 🙂

    https://uncommondescent.com/intelligent-design/they-wont-dance-they-wont-mourn/

  424. DATCG:

    OK, it had to happen:

    Here are the ubiquitin system and the spliceosome joined together, with additional involvement in DNA repair. And in a very complex way. Mol Cell. 2018 Mar 15:

    Prp19/Pso4 Is an Autoinhibited Ubiquitin Ligase Activated by Stepwise Assembly of Three Splicing Factors

    Abstract
    Human nineteen complex (NTC) acts as a multimeric E3 ubiquitin ligase in DNA repair and splicing. The transfer of ubiquitin is mediated by Prp19-a homotetrameric component of NTC whose elongated coiled coils serve as an assembly axis for two other proteins called SPF27 and CDC5L. We find that Prp19 is inactive on its own and have elucidated the structural basis of its autoinhibition by crystallography and mutational analysis. Formation of the NTC core by stepwise assembly of SPF27, CDC5L, and PLRG1 onto the Prp19 tetramer enables ubiquitin ligation. Protein-protein crosslinking of NTC, functional assays in vitro, and assessment of its role in DNA damage response provide mechanistic insight into the organization of the NTC core and the communication between PLRG1 and Prp19 that enables E3 activity. This reveals a unique mode of regulation for a complex E3 ligase and advances understanding of its dynamics in various cellular pathways.

    The Prp19/nineteen complex (NTC) is a multifunctional protein complex involved in very diverse biological processes, including pre-mRNA splicing and the DNA damage response (DDR)

    Prp19 is one more Prp involved, among other things, in the spliceosome assembly. From Uniprot:

    Ubiquitin-protein ligase which is a core component of several complexes mainly involved pre-mRNA splicing and DNA repair. Core component of the PRP19C/Prp19 complex/NTC/Nineteen complex which is part of the spliceosome and participates in its assembly, its remodeling and is required for its activity. During assembly of the spliceosome, mediates ‘Lys-63’-linked polyubiquitination of the U4 spliceosomal protein PRPF3. Ubiquitination of PRPF3 allows its recognition by the U5 component PRPF8 and stabilizes the U4/U5/U6 tri-snRNP spliceosomal complex (PubMed:20595234). Recruited to RNA polymerase II C-terminal domain (CTD) and the pre-mRNA, it may also couple the transcriptional and spliceosomal machineries (PubMed:21536736). The XAB2 complex, which contains PRPF19, is also involved in pre-mRNA splicing, transcription and transcription-coupled repair (PubMed:17981804). Beside its role in pre-mRNA splicing PRPF19, as part of the PRP19-CDC5L complex, plays a role in the DNA damage response/DDR. It is recruited to the sites of DNA damage by the RPA complex where PRPF19 directly ubiquitinates RPA1 and RPA2. ‘Lys-63’-linked polyubiquitination of the RPA complex allows the recruitment of the ATR-ATRIP complex and the activation of ATR, a master regulator of the DNA damage response (PubMed:24332808). May also play a role in DNA double-strand break (DSB) repair by recruiting the repair factor SETMAR to altered DNA (PubMed:18263876). As part of the PSO4 complex may also be involved in the DNA interstrand cross-links/ICLs repair process (PubMed:16223718). In addition, may also mediate ‘Lys-48’-linked polyubiquitination of substrates and play a role in proteasomal degradation (PubMed:11435423). May play a role in the biogenesis of lipid droplets (By similarity). May play a role in neural differentiation possibly through its function as part of the spliceosome

    So many proteins involved in this extraordinary multi-protein complex:

    Prp19 (504 AAs) (as an homotetramer)
    SPF27 (225 AAs)
    CDC5L (802 AAs)
    PLRG1 (514 AAs)

    All of them highly conserved! 🙂

  425. DATCG:

    And this is about the NineTeen Complex (NTC):

    The NineTeen Complex (NTC) and NTC-associated proteins as targets for spliceosomal ATPase action during pre-mRNA splicing.

    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4615276/

    Abstract:

    Pre-mRNA splicing is an essential step in gene expression that removes intron sequences efficiently and accurately to produce a mature mRNA for translation. It is the large and dynamic RNA-protein complex called the spliceosome that catalyzes intron removal. To carry out splicing the spliceosome not only needs to assemble correctly with the pre-mRNA but the spliceosome requires extensive remodelling of its RNA and protein components to execute the 2 steps of intron removal. Spliceosome remodelling is achieved through the action of ATPases that target both RNA and proteins to produce spliceosome conformations competent for each step of spliceosome activation, catalysis and disassembly. An increasing amount of research has pointed to the spliceosome associated NineTeen Complex (NTC) of proteins as targets for the action of a number of the spliceosomal ATPases during spliceosome remodelling. In this point-of-view article we present the latest findings on the changes in the NTC that occur following ATPase action that are required for spliceosome activation, catalysis and disassembly. We proposed that the NTC is one of the main targets of ATPase action during spliceosome remodelling required for pre-mRNA splicing.

    Look at Fig. 1 for a “simple” summary. 🙂

  426. DATCG:

    Well, the issue is more complex than I thought.

    It seems that NineTeen complex is involved in the many phases of the spliceosome assembly, with all the activities described in the paper linked at #428.

    However, the complex itself is formed by at least 8 core proteins (in yeats): Prp19, Cef1, Syf1, Syf2, Syf3,
    Snt309, Isy1 and Ntc20, plus about 18 associated proteins.

    See here:

    The function of the NineTeen Complex (NTC) in regulating spliceosome conformations and fidelity during pre-mRNA splicing.

    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4234902/

    Abstract:

    The NineTeen Complex (NTC) of proteins associates with the spliceosome during pre-mRNA splicing and is essential for both steps of intron removal. The NTC and other NTC-associated proteins are recruited to the spliceosome where they participate in regulating the formation and progression of essential spliceosome conformations required for the two steps of splicing. It is now clear that the NTC is an integral component of active spliceosomes from yeast to humans and provides essential support for the spliceosomal snRNPs (small nuclear ribonucleoproteins). In the present article, we discuss the identification and characterization of the yeast NTC and review recent work in yeast that supports the essential role for this complex in the regulation and fidelity of splicing.

    In particular, Table 1

    But the strange point is that:

    The NTC is named after the splicing factor Prp19, which was first identified in 1993 as a splicing factor in the yeast S. cerevisiae [4]. Prp19 is essential for splicing but is not a constituent of any of the individual spliceosomal snRNPs [5,6]. Association of Prp19 with itself to form tetramers provides the basis for the hypothesis that Prp19 provides a scaffold for NTC organisation [7]. Prp19 contains a U-box domain which exhibits E3 ubiquitin ligase activity in vitro [8], however, a target for this activity in the spliceosome is still lacking.

    Emphasis mine.

    IOWs, Prp19 is definitely an E3 ligase, but that activity is not documented in the spliceosome assembly (although it could certainly be present).

    However, the very recent paper referenced at #427 clearly documents the E3 ligase activity, but in relation to DNA repair, not to spliceosome assembly.

    Moreover, the 3 proteins listed at #427 are required for the E3 ligase activity in DNA repair, but are not apparently part of the NTC complex, as I had believed initially (I have corrected my comment in that sense).

    These proteins are really amazing, I would say. 🙂

  427. DATCG:

    Here is another protein regulated by ubiquitin and involved in splicing, Sde2:

    Sde2 is an intron-specific pre-mRNA splicing regulator activated by ubiquitin-like processing.

    http://emboj.embopress.org/content/37/1/89.long

    Abstract
    The expression of intron?containing genes in eukaryotes requires generation of protein?coding messenger RNAs (mRNAs) via RNA splicing, whereby the spliceosome removes non?coding introns from pre?mRNAs and joins exons. Spliceosomes must ensure accurate removal of highly diverse introns. We show that Sde2 is a ubiquitin?fold?containing splicing regulator that supports splicing of selected pre?mRNAs in an intron?specific manner in Schizosaccharomyces pombe. Both fission yeast and human Sde2 are translated as inactive precursor proteins harbouring the ubiquitin?fold domain linked through an invariant GGKGG motif to a C?terminal domain (referred to as Sde2?C). Precursor processing after the first di?glycine motif by the ubiquitin?specific proteases Ubp5 and Ubp15 generates a short?lived activated Sde2?C fragment with an N?terminal lysine residue, which subsequently gets incorporated into spliceosomes. Absence of Sde2 or defects in Sde2 activation both result in inefficient excision of selected introns from a subset of pre?mRNAs. Sde2 facilitates spliceosomal association of Cactin/Cay1, with a functional link between Sde2 and Cactin further supported by genetic interactions and pre?mRNA splicing assays. These findings suggest that ubiquitin?like processing of Sde2 into a short?lived activated form may function as a checkpoint to ensure proper splicing of certain pre?mRNAs in fission yeast.

    And:

    Intron specificity in pre-mRNA splicing

    https://link.springer.com/article/10.1007%2Fs00294-017-0802-8

    Abstract
    The occurrence of spliceosomal introns in eukaryotic genomes is highly diverse and ranges from few introns in an organism to multiple introns per gene. Introns vary with respect to their lengths, strengths of splicing signals, and position in resident genes. Higher intronic density and diversity in genetically complex organisms relies on increased efficiency and accuracy of spliceosomes for pre-mRNA splicing. Since intron diversity is critical for functions in RNA stability, regulation of gene expression and alternative splicing, RNA-binding proteins, spliceosomal regulatory factors and post-translational modifications of splicing factors ought to make the splicing process intron-specific. We recently reported function and regulation of a ubiquitin fold harboring splicing regulator, Sde2, which following activation by ubiquitin-specific proteases facilitates excision of selected introns from a subset of multi-intronic genes in Schizosaccharomyces pombe

    Both of January 2018.

    Strangely, Sde2 is not linked to splicing in Uniprot:

    Involved in both DNA replication and cell cycle control (PubMed:27906959). Unprocessed SDE2 interacts with PCNA via its PIP-box. The interaction with PCNA prevents monoubiquitination of the latter thereby inhibiting translesion DNA synthesis. The binding of SDE2 to PCNA also leads to processing of SDE2 by an unidentified deubiquinating enzyme, cleaving off the N-terminal ubiquitin-like domain. The resulting mature SDE2 is degraded by the DCX(DTL) complex in a cell cycle- and DNA damage dependent manner (PubMed:27906959). Binding of SDE2 to PCNA is necessary to counteract damage due to ultraviolet light induced replication stress. The complete degradation of SDE2 is necessary to allow S-phase progression

    So, its role in intron splicing seems to be a really recent discovery.

    By the way, human Sde2 is not highly conserved, but has a rather slow emergence in evolutionary history, with two dicrete jumps at the vertebrate and mammal transitions.

  428. UB

    “ES, I do not see any mail, my friend”.

    That explains your silence 🙂 I’ll send another one. Thanks for letting me know.

    I always enjoy reading this kind of OPs, as you know. But time is really tight just now. I hope to be able to devote more time to it in due course.

  429. Hey guys, hope everyone had a great weekend, restful and invigorating 🙂

    Gpuccio @425 – 430! 🙂 Wow!

    It’s always fun to read all the spectacular details of what you’ve posted.

    I especially like Splicesome Ubiquitin connections as expected. And I’d love to delve into that area you highlighted, especially a favorite area of introns for me 🙂

    My time is limited however so will focus on the part of your OP on Phosphorylation
    ———————

    On #425… building upon an analogy of Design process and Markup Languages…

    The Phosphorylation Tag as a refined, or fine tuning condition step is very interesting. What is it accomplishing and why precede UPS with a unique tag?

    Whats the reason for this specific pathway? To release a transcription factor? For that matter, why is this a Pre-release state required as a partial degradation for release?

    Weird, huh? Is there a precedent in Design and Coding realms for an Information processing analogy? There’s a reason Bill Gates said, “DNA is like a computer program but far, far more advanced than any software ever created.”

    Well, why would he say that? Because he is intimately aware of how a CPU works with Coding languages. He recognizes Code when he sees it in an operating system.

    So yes, there is a precedent in information processing. Programmers maintain internal information, code, data and append internal tags or external tags in a table or database and strip data or “inhibitors” if you like as Pre-processing requirements are met, based upon different input signals.

    So, if we view the Cytoplasm as another method of large CPU-processing Memory in the Cell, Or, lets call it an “aqueous MotherBoard*” this begins to make sense as an Information Processing analogy.

    Data or in this case, the NF-?B TF Struture is Pre-formatted. That alone is a sign of Design.

    The NF-kB Transcription Factor is ready and waiting for activation to instigate it’s release based upon specific input signal(s) for it’s eventual modification and release into the Nucleus.

    One of my first questions is…
    Why have a pre-formatted structure waiting for use in a blind, unguided neo-Darwinian story?

    This Pre-formatted TF does not fire off for any reason. In fact, if it starts firing off for any reason, it would cause chaos and possible damage downstream. So it’s tightly regulated and only released upon correct Signal(s).

    It must match the signal input. And in turn the pathway has to launch the correct response by a Directive or Rules based procedure code that has locations built-in for eventual locations.

    It’s like a data-table or structure is be preloaded into memory ready for quick access and retrieval by a CPU-nucleus instruction set for transcription.

    This is not a single Input -> Modification -> Output process. These rapid process are going on in parallel, especially for quick reaction immune systems and/or repair mechanisms and any millions of trillions of cellular transactions at any one time in our body, brain, heart, skin, immune, gut, etc. etc., etc.

    But simplifying, we have…

    Input to Aqueous Cytoplasm “Motherboard” for the Release of Transcription Factor :
    -> Signal A
    —> Structure Awaiting Signal A in Wet Memory
    —–> Tag for Modification
    ——-> Alert UPS
    ———> Strip Component Part, freeing TF
    ———–> Send NF-?B TF to Nucleus for processing

    An interesting question arises which I think is even more succinct and descriptive of a Design process.

    Why not ubiquitin Mono, Poly, or Branched tags?
    Why a requirement for a conditional, two-step Tagging process?

    Why not a one-step UPS solution or chaining events? That would satisfy partial degradation and release of TF to the nucleus?

    I need more time to review. Unfortunately I do not have enough background in biochemistry and atomic structures to know why these different Tagging procedures might be required.

    A few searches have not produced answers though I may easily be missing them. Is it a conformational component where Phosphorylation is required to adjust folds?

    If by Design, we should be able to qualify the reasons for conditions of refinement specificity that you mention and Tagging requirements.

    So much to review 🙂

    *Aqueous Motherboard – what else can we call it? Or designate the Cytoplasm as? Other than it’s aqueous structure of floating functions and organelles – it resembles a motherboard. The Cytoplasm functions as a motherboard for active, instant retrieval of Pre-formatted structures, specified functions and specialized processing units(1). These units and floating pre-formatted functions surround the CPU – Core process-Nucleus instruction set for Eukaryotes. This enables high speed throughput, as signals enter and ignite pre-programmed responses.

    (1) specialized-processing units = organelles.

    What am I missing? What more might be added I’m leaving out in symbolic tagging and semiosis? Or processing functionality?

  430. Correction of sentence from 432 above:

    It must match the signal input. And in turn the pathway has to launch the correct response by a Directive or Rules based procedure code that has addresses or addressable-location mechanisms built-in to guide for eventual locations and active response(i.e. inflammatory and/or immune responses must be directed to correct area of inflammation and repair)

  431. Hmmmm…

    “Phosphorylation of IkBalpha on serines 32 and 36 is mediated by IkB kinases(IKKs), whose activity is induced by activators of the NFkB pathway. IKK activity exists as large Cytoplasmic multi-subunit complex(700-900kDAa) containing two kinase subunits, IKK1(IKKalpha) and IKK2(IKKbeta), and regulartory subunit, NEMO…”

    “Sequence analysis revealed that both IKK1 and IKK2 contain a canonical MAP kinase(MAPKK) activation loop motif. This region contains specific sites whose phosphorylation induces a conformational change that results in kinase activation.”

    Hmmm, is that Kinase activation in support of Tagging.

    OK, so this is interesting. Paragraphs above and blockquote below are from google book source. There may be a few typos:
    Regulation of Organelle and Cell Compartment Signaling: Cell Signaling…

    edited by Ralph A. Bradshaw, Edward A. Dennis(1st Edition 2011).

    Phosphorylation within the activation loop typically occurs through the action of an upstream kinase or through transphosphorylation enabled by regulated proximity between two kinase subunits. IKK2 activation loop mutations, in which serines 177 and 181 were replaced with alanine, render the kinase refarctory to stimulus dependent activiatio. In contrast, replacement of serine 177 and 181 with glutamic acid, to mimic phoserine, yielded a constitutively active kinase, and was capable of cell stimulation. The corresponding mutations in IKK1 did not interfere wiht NFkB activation in response to IL-1 or TNF, providing the first data suggesting that IKK2 plays a more prominent role in NFkB activation in response to proinflammatory cytyokines.

    .

  432. So, is that activity around serines 177 and 181 applicatlbe to serines 32 and 36? I don’t know if same rules apply.

    It is from another report in the book linked in #434, that describes…

    “two potentially novel components of the IKK complex, namely Cdc37 and Hsp90. Apparently, formation of the core IKK complex with Cdc37/Hsp90 is required for TNF induced activation and recruitment of the core IKK complex from the Cytoplasm to the membrane.”

    Different functional requirements may be unrelated.

  433. DATCG:

    First of all, I would like to offer some clarifications about the complex issue of phosphorylation in the NFkB pathway, just to avoid confusion.

    a) The protein which is phosphorylated at serines 32 and 36 is IkB alpha, which is a direct inhibitor of RelA p50 (in the canonocal pathway of activation). The double phopshorylation leads to ubiquitination of the inhibitor and to its degradation in the proteasome, releasing the transcription factor (RelA p50, which is one form of NFkB TF).

    b) The phosphorylation described in a) is done by a big molecular complex, which includes the two IKK1 (or alpha) and IKK2 (or beta) proteins, NEMO and at least 3 other proteins (Hsp90, Cdc37 and ELKS). Many of those proteins must be phosphorylated to be active, in particular IKK1 at Ser 176 and 180, IKK2 at Ser 177 and 181.

    c) Not much is understood of the processes that lead to the phosphorylations described in b). The TAK1 kinase seems to be involved.

    d) However, the formation of the protein complex described in b) is cuased by another protein complex, which adheres to the cell membrane, and includes TRADD, TRAF2 and RIP (this one, too, phosphorylated).

    e) The formation of the complex described in d) is caused by the reaction between a specific membrane receptor and a cytokine (usually Tumor Necrosis Factor, TNF).

    Simple, isn’t it? And this is only the canonical pathway! 🙂

    OK, the above information is not extremely recent, it comes from the following two papers:

    Phosphorylation of NF-kB and IkB proteins: implications in cancer and inflammation

    https://orbi.uliege.be/bitstream/2268/1280/1/21.%20Review%20phosphorylation%20NF-kB%20TIBS.pdf

    See especially Fig. 1.

    And:

    The IKK Complex, a Central Regulator of NF-?B Activation

    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2829958/pdf/cshperspect-NFK-a000158.pdf

    I just wanted to clarify these points to go on with some more general discussion in next post.

  434. DATCG at #432:

    Now, the more general discussion, for which I will use our NFkB as a model, but there are lots of similar systems in the cell.

    In your comment you have touched a very important point. An extremely important point, I would say.

    In brief, I will sum it up as followes:

    Why such complex multiple intertwined regulation systems to control one single pathway?

    I suggest that anyone who reads the following could refer to Fig. 1 in the already quoted paper:

    https://orbi.uliege.be/bitstream/2268/1280/1/21.%20Review%20phosphorylation%20NF-kB%20TIBS.pdf

    That will make the discussion simpler. Please, refer only to the left part of the Figure, which represents the canonical activation pathway.

    Now, the whole process coule be summarized as follows:

    a) A signal arrives at the cell membrane (The TNF cytokine)

    b) A transcription factor is activated to convey the message to the nucleus.

    c) The activated TF interacts with the genome and aucses a series of events there.

    In our Fig. 1, it is easy to identify the eseential actors.

    a) The cytokine (TNF) and its membrane receptor (TNFR1) are well visible at the top of the Figure (left part).

    b) Our TF is the p50-p65 entity (p65 is the same as RelA, so this is the same as RelA-p50 in Fig. 5 of the OP).

    c) The TF interacting with DNA can be seen at the bottom of the Figure.

    OK, so I think that the spontaneous question that everyone is asking is:

    But there are a lot of other things there! Why?

    And there are!

    a) The protein complex which forms at the inner side of the cell membrane (3 proteins, one of which phosphorylated, + SODD which has to be released for the activation)

    b) The big protein complex which phosphorylates the TF to release it from its inhibitor: 6 proteins, many of which phosphorylated.

    c) The inhibitor of the TF (IkB alpha)

    d) The SKP1 – beta TRC complex which ubiquinates the inhibitor after its double phosphorylation (not shown in the figure, see Fig. 7 in the OP): 6 proteins, including ubiquitin.

    e) And, of course, the proteasome.

    And I would bet that a lot of other components are not shown, maybe even not known (for example, the various Kinase systems that phosphorylate many of the mentioned proteins).

    Each of those components has an active role in the regulation and control of the process.

    And again, the question is: why?

    Of course, anyone can see how simpler it would have been in this other way:

    The interaction between TNF and its receptor enzumatically modifies a TF (p50-p65) which was before in some inactive state. The active TF then relocates to the nucleus, where it does what it has to do.

    Simpler, isn’t it?

    And beware, that simpler version could easily be controlled too, for example at the level where the TF is activated by the membrane receptor.

    So, why all the added complexity?

    That is a difficult question. I will try to discuss some aspects in next post.

  435. Ciao Gpuccio @436-437 🙂

    And thanks! Especially on clarifications and this one…

    b) The phosphorylation described in a) is done by a big molecular complex, which includes the two IKK1 (or alpha) and IKK2 (or beta) proteins, NEMO and at least 3 other proteins (Hsp90, Cdc37 and ELKS). Many of those proteins must be phosphorylated to be active, in particular IKK1 at Ser 176 and 180, IKK2 at Ser 177 and 181.

    To see IKK2(important role) at Ser 177 and 181 is interesting. I’ve missed something and thought it was Ser 32, Ser 36.

    Thanks for recognizing the question(however uninformed of biochemical pathways) about processes taking place – which seems like at initial look – to be over regulated for lack of better terminology.

    So, taking neo-Darwinist side, would a hodge-podge of pathways like this lead credence to their theory? Or, upon closer inspection will we find purposes for:

    a) multiple pathways
    b) reasons for phosphorylation steps
    c) ubiquitinylatoin steps
    d) Reason for Pre-Formatted NF-kB Waiting in the Cytoplasm?

    I’d really like a neo-Darwinist to explain D) Pre-formatted TF w/ inhibitor awaiting signals.

    Why? Why is is there? Makes no since for an unguided, blind process to pre-plan actions. Does it? Will neo-Darwinist appeal to bad design as an answer?

    So yes, the question again, is Why? Why for many of these steps.

    I was in the middle of breaking down the two pathways of Figure 1, Classical vs Alternative(TNF vs CD40) and simply do not have enough time at this moment. But will hopefully return later tonight and finish before posting.

    But what we see is Directed Response pathways for TNF and CD40, plus a third pathway which I’m not planning to cover.

    I like how you are detailing what might happen in a simpler pathway – if only we knew all of these processing techniques and other components.

    I think the more we peer in to different Signaling actions and Pathways, starting at the membrane, the more beneficiary it is to a Design hypothesis.

    First, it must distinguish between TNF and CD40. These are conditional signals for appropriate specified and targeted immune responses(ie. viruses in alternative pathway of CD40)

    OK, have very busy week ahead, but I’ll respond when I can! Really enjoying this OP Gpuccio!

    And I hope other readers appreciate all your efforts here.
    And hope they feel OK to ask any questions.

  436. Gpuccio,
    Before I go, restating for my own clarity and maybe others what you have stated re: “simpler” it might be on another pathway.

    “…how simpler it would have been in this other way:

    1) The interaction between TNF and its receptor enzumatically modifies a TF (p50-p65)

    OK, that’s bypassing a huge Complex and many phosphorylation steps. So, am I correct in speculating Phosphorylation is crucial in ways I do not understand yet?

    2) …which was before in some inactive state.

    This is to me is an important Design concept or at least a good question: inactive state

    I question and expanded in previous comment @438.

    Why have NF-kB protein complex hanging around? I designate it as a “Pre-Formatted” state.

    Speed considerations? Quick response and deployment? Is there another Conditional element where different signals change the resulting pathway or proteolytic process?

    Why would a supposedly blind, unguided “process” do it?

    And how would it know it the Protein Complex would ever be utilized or “evolve” to be utilized in such a pathway?

    I’ve not seen previous remarks on this by neo-Darwinist. So if any readers supporting a neo-Darwinian pathway of evolution for this scenario can comment, I’d be happy to see it.

    Why does a blind, unguided series of events decide in the past, millions of years ago even that:

    a) I’m going to leave a Pre-Formatted protein complex like NF-kB waiting around in suspended animation?

    b) So a signal process can cause a chain reactions of Lemony Snicket series of “FORTUNATE” events?

    This does not make sense even for Lemony Snicket.

    3) The active TF(edit) then relocates to the nucleus, where it does what it has to do.

    After a series of unfortunate Lemony Snicket’s events! 😉 it proceeds to the Nucleus where a whole other series of unfortunate Lemony Snicket events translate, transcribe, Post-Modify and transpire to send an immune response to the target of the alert.

    Amazing! 🙂 That Lemony Snicket!

    Simpler, isn’t it?

    And beware, that simpler version could easily be controlled too, for example at the level where the TF is activated by the membrane receptor.

    So, why all the added complexity?

    Because, two kids showed up and created a new pathway for Lemony Snicket to go down, all by random walks, accidental events and “natural selection.”

    .

  437. Note: @439

    Bigger Picture: One question leads to another

    Why does a blind, unguided series of events “decide” in the past, millions of years ago to “evolve” a solution so that:

    a) a Pre-Formatted protein complex like NF-kB waits around in suspended animation for a cascade of events initiated by a signal event through a specific TNF pathway?

    Q: How many other Pre-formatted Protein Complexes are waiting in Suspended Animation of the Cytoplasm?

    The irreducible complexity of such networked systems amounts to multiple interdependent systems coordinating organized interactions while “simultaneously” and “blindly evolving” working solution steps at just the right time for any of these pathways to function.

    OK, now I must go. Will check in later.

    (Edit) and this does not include other possible pathways for this specific NF-kB protein complex for other factors(signals processing) and decisions. Either Pre or Post Phosphorylation and Ubiquitination.

  438. DATCG:

    OK, I would like to go on with the more general discussion starting from what I have already said at #437.

    Why all the added complexity?

    I would like to start with an old friend, Michael Behe, and with one of his first metphors (in Darwin’s black box): the Rude Goldberg machine.

    For example, here is an example:

    https://en.wikipedia.org/wiki/Rube_Goldberg_machine#/media/File:Rube_Goldberg%27s_%22Self-Operating_Napkin%22_(cropped).gif

    Now, it is rather obvious that the scenario we have seen at #437 about the NFkB pathway (see Fig. 1 of the quoted paper) does resemble a Rude Goldberg machine.

    But there is more. Behe offers, in his fundamental book, a couple of important examples of irreducible complexity: the bacterial flagellum and the coagulation cascade.

    But those examples are a little different from out regulation overkill. In a sense, the irreducible complexity is more “understandable” there. For example, in the flagellum the various parts, stator, rotor, filament and so on are parts of a machine. their role, therefore, is immediately obvious.

    In the coagulation cascade, the linear cascade can be explained, in a way, by the need to amplify linearly the signal to get a wide final effect.

    But in out regulation scenario, the explanation is less obvious.

    There is also another feature which can help us in our approach: the regulation network we have seen is not linear.

    If we look at the famous Fig. 1 already quoted, while the main cascade can be considered linear, there are many “cross-talks”. For example, some phosphorylation systems and the ubiquitination step act “at the side” of the cascade.

    Now, I would propose what seems to me the only reasonable explanation for the “added complexity”.

    The reason for the added complexity is that the pathway, like almost all similar pathways that work between the cell membrane and the nucleus, is not an isolated mechanism, but is part of a huge “neural network” which involves all the different pathways which transmit and integrate the communication between outward signals (the cell membrane) and the final transcription regulation in the nucleus.

    My idea is that the many proteins that are involved in the many “redundant” steps in the pathway are regulation nodes, and act as “sensors” which integrate the specific pathway with all that happens in the cytoplasm, receiving information from the other pathways and transmitting information to them.

    That is also an explanation to the more specific answer:

    Why are some apparently simple steps implemented by huge multi-protein complexes?

    For example, why is the double phosphorylation of IkB alpha performed by a structure made of 9 protein blocks, and not by a single kinase?

    One possible answer is: because it must receive and transmit information to other cell pathways, interacting with many other protein structures.

    IOWs, we see here something similar to what happens at the level of the nucleus with the combinatorial working of TFs in big multi-protein structures.

    Or, if we want to push the similitude even further, something similar to what happens in the synapses to integrate many different signals.

    OK, these are just a few tentative thoughts. But, if there is something true in these ideas, then the “neural network” of transmission pathways really deserved a lot of attention.

    And, of course, only a design perspective can help in this kind of issues.

  439. Alan Fox has taken umbrage with the claiming of ubiquitin as evidence for ID. Of course he doesn’t have any idea how blind and mindless processes could have produced it and he doesn’t have any idea how to test the claim that they could. So there.

    the alleged refutation of the concept of ubiquitin as evidence for ID

    Alan talks about alternatives- alternative to what? Alan’s alleged evolutionary theory doesn’t have some theory or hypothesis that could be a testable explanation for the ubiquitin system, to begin with.

  440. ET:

    Oh no! Not another corss-talk with TSZ! 🙂

    OK thanks for the notification. I checked TSZ and found the thread (rather short, for the moment, thanks God!).

    I must say that the paper you link was really mentioned by TomMueller, not Alan Fox (who is instead the author of the OP).

    Well, I am going to answer in next post. 🙂