Title

Perilous journey: a tour of the ubiquitin-proteasome system.

Journal

Trends in cell biology

Volume

24

Issue

None

Pages

352-9

Date

2014-06-01

Authors

Kleiger G | Mayor T

These studies demonstrated that both E2s and E3s can affect the conformation of the ubiquitin on the E2 surface to promote its transfer to the substrate

Another key early finding was that the cleavage of the ubiquitin chain from the substrate was ATP-dependent and was coupled to the translocation of the protein substrate into the 20S core

These enzymes, referred to as deubiquitinases (DUBs), cleave ubiquitin from conjugated proteins or edit ubiquitin chains by trimming their lengths [62]

Thus, the binding of substrate to the 26S and its concomitant translocation into the 20S results in the activation of Rpn11 through a conformational change.

For instance, ubiquitin can recruit other factors to mediate various cellular responses such as signaling, gene regulation, endocytosis, macro-autophagy, and DNA repair

This function is crucial for cellular homeostasis because failure to activate ubiquitin, as seen by the chemical inhibition of E1 activity in the cell, results in the almost immediate shutdown of the entire UPS

Specifically, Bag6 recognizes and binds to long hydrophobic stretches of the polypeptide chain of misfolded proteins targeted by the ERAD (endoplasmic reticulum associated protein degradation) pathway [56–58]

This observation suggests that chain binding and cleavage from the protein substrate are tightly coordinated by the proteasome, and may help to prevent situations such as premature chain cleavage by Rpn11, which could result in the release of the substrate before it becomes actively engaged to the proteasome.

An E1 enzyme must first activate ubiquitin, a highly conserved, 76 amino acid polypeptide, in an ATP-dependent manner.

E1s are multidomain enzymes that must activate ubiquitin and efficiently transfer it to the E2 active site

Because Bag6 interacts with both E3 ligases as well as the proteasome [57,58,60], it may promote both the ubiquitination and delivery of misfolded proteins to the proteasome while preventing their aggregation

It was initially believed that the presence of DUBs at the proteasome (such as Uch37/UchL5 and Usp14) would promote the degradation of proteins through facilitating substrate processing (see also the next section)

However, recent studies have shown that, by contrast, upon chemical inhibition of the proteasome-bound DUB Usp14, the degradation of aggregation-prone substrates in mammalian tissue culture cells significantly increased [65]

Finally, the E3 ubiquitin ligase binds to both the E2-bound ubiquitin and the protein substrate, promoting the transfer of ubiquitin onto the substrate

However, earlier biochemical investigations on the SCF (Skp, Cullin, F-box containing) ubiquitin ligase complex, the archetypal member of the cullin–RING ligases, indicated that E3s can also positively influence the rate of ubiquitin transfer from the E2 to the protein substrate [17,18].

Some E3s can also stabilize the conformation between ubiquitin and the E2, thereby accelerating further the rate of conjugation of ubiquitin to proteins

Indeed, recent studies have provided insight into the mechanism of E3-mediated activation of E2s by trapping and either co-crystallizing or characterizing by nuclear magnetic resonance (NMR) the unstable and transient complexes between E2~ubiquitin and E3.

One key take-home message is that many E3 ligases likely evolved to be processive, meaning that they can conjugate multiple ubiquitins onto a substrate before it is released from enzyme

Recent work has shown that the E2s Cdc34 (cell division cycle 34) and Ube2S (ubiquitin-conjugating enzyme E2S) also form noncovalent interfaces with ubiquitin in addition to the covalent thioester bond [23,24], suggesting that E2s catalyze ubiquitin transfer at least in part by holding ubiquitin against an interface on the E2 surface that optimizes the position of the thioester bond in the active site

The thioesterified ubiquitin passes from the E1 active site to the next member of the cascade, the E2 or ubiquitin-conjugating enzyme

In summary, mechanisms of E2 activation involve the binding of ubiquitin to surfaces on the E2 and/or E3, thereby allowing an optimal conformation of ubiquitin on the E2 surface

It was originally shown that the E2 Ubc1 (ubiquitin-conjugating enzyme E2 1) can form a non-covalent interface with a ubiquitin that is thioesterified to the active site [22]

Misfolded proteins can aggregate with time, and protein quality-control pathways are tasked with their ubiquitination and subsequent degradation [46,47].

First, ubiquitinated proteins are docked to the proteasome through receptors associated to the 19S [67,68], including the proteasome proteins Rpn10 (proteasome regulatory particle base subunit; also known as Psmd4 – proteasome 26S subunit, non- ATPase, 4) and Rpn13 (also known as Adrm1, adhesion regulating molecule 1), that bind to the poly-ubiquitin chains [69,70].

Second, Rpn11/Poh1 (also known as Psmd14) is a subunit in the 19S regulatory particle that cleaves the entire, intact ubiquitin chain from the protein substrate [71,72].

This activity promotes both the recycling of chains back into the free cellular pool of ubiquitin and creates space for the protein substrate to enter the 20S core.

Similarly, the inactivation of the evolutionarily related DUB in yeast, Ubp6, increases the cellular fitness of aneuploid cells, which have been shown to incur increased proteotoxic stresses due to chromosomal imbalance [66].

For example, yeast Cdc48 (p97 in mammalian cells) is a conserved multisubunit enzyme that plays a major role in dissociating ubiquitinated proteins from their binding partners to promote their degradation by the proteasome [52]

Cdc48/p97 generally acts downstream of ubiquitin ligases, although its activity may also promote ubiquitination in some cases; a major challenge is to delineate further the mechanism of action of this multipurpose enzyme

Both HECT and RBR E3s are unique in comparison to the RING ligases because they catalyze an additional transthioesterification step, where ubiquitin is transferred from the E2 to the E3 before its conjugation to the protein substrate (Figure 1A)

Indeed, three recent structural studies observed the E2~ubiquitin conformation in the presence of either RING domains or the structurally related U-box domain [19–21]

BEL Commons is developed and maintained in an academic capacity by Charles Tapley Hoyt and Daniel Domingo-Fernández at the Fraunhofer SCAI Department of Bioinformatics with support from the IMI project, AETIONOMY. It is built on top of PyBEL, an open source project. Please feel free to contact us here to give us feedback or report any issues. Also, see our Publishing Notes and Data Protection information.

If you find BEL Commons useful in your work, please consider citing: Hoyt, C. T., Domingo-Fernández, D., & Hofmann-Apitius, M. (2018). BEL Commons: an environment for exploration and analysis of networks encoded in Biological Expression Language. Database, 2018(3), 1–11.