Autoinhibition Mechanism of the Ubiquitin-Conjugating Enzyme UBE2S by Autoubiquitination

Liess, A.K.L.; Kucerova, Alena; Schweimer, Kristian; Lu, Yu; Roumeliotis, Theodoros I.; Diebold, Mathias; Dybkov, Olexandr; Sotriffer, Christoph A.; Urlaub, Henning; Choudhary, Jyoti S.; Mansfeld, Jörg; Lorenz, Sonja

doi:10.1016/j.str.2019.05.008

Cited by 22 publications

(37 citation statements)

References 103 publications

(148 reference statements)

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“…Samples were analyzed by SDS-PAGE and fluorescence scan using a Typhoon FLA 9500 laser scanner (GE Healthcare). All ubiquitination reactions with Ubc1 were performed with K93R mutants to block autoubiquitination and thus autoinhibition (Liess et al, 2019). For kinetic analyses, reactions were performed in triplicate.…”

Section: In Vitro Ubiquitination Experimentsmentioning

confidence: 99%

The UBA domain of conjugating enzyme Ubc1/Ube2K facilitates assembly of K48/K63‐branched ubiquitin chains

et al. 2021

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The assembly of a specific polymeric ubiquitin chain on a target protein is a key event in the regulation of numerous cellular processes. Yet, the mechanisms that govern the selective synthesis of particular polyubiquitin signals remain enigmatic. The homologous ubiquitin-conjugating (E2) enzymes Ubc1 (budding yeast) and Ube2K (mammals) exclusively generate polyubiquitin linked through lysine 48 (K48). Uniquely among E2 enzymes, Ubc1 and Ube2K harbor a ubiquitin-binding UBA domain with unknown function. We found that this UBA domain preferentially interacts with ubiquitin chains linked through lysine 63 (K63). Based on structural modeling, in vitro ubiquitination experiments, and NMR studies, we propose that the UBA domain aligns Ubc1 with K63linked polyubiquitin and facilitates the selective assembly of K48/ K63-branched ubiquitin conjugates. Genetic and proteomics experiments link the activity of the UBA domain, and hence the formation of this unusual ubiquitin chain topology, to the maintenance of cellular proteostasis.

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Section: In Vitro Ubiquitination Experimentsmentioning

confidence: 99%

The UBA domain of conjugating enzyme Ubc1/Ube2K facilitates assembly of K48/K63‐branched ubiquitin chains

et al. 2021

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“…The ubiquitin proteasome pathway, conserved from yeast to human, is required for the targeted degradation of most proteins in cells [21]. Traditional ubiquitin modification is an ATP-dependent process carried out by three classes of enzymes including ubiquitin activating enzyme (E1), ubiquitin conjugating enzyme (E2), and ubiquitin ligase (E3) [22,23]. However, sometimes this process is not strictly followed.…”

Section: Introductionmentioning

confidence: 99%

O-GlcNAc-Modification of NSL3 at Thr755 Site Maintains the Holoenzyme Activity of MOF/NSL Histone Acetyltransferase Complex

Zhao

Wei

et al. 2019

IJMS

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Both OGT1 (O-linked β-N-acetylglucosamine (O-GlcNAc) transferase isoform 1) and NSL3 (nonspecific lethal protein 3) are crucial components of the MOF (males absent on the first)/NSL histone acetyltransferase complex. We previously described how global histone H4 acetylation levels were modulated by OGT1/O-GlcNAcylation-mediated NSL3 stability. However, the specific modification site of NSL3 and its molecular mechanism of protein stability remain unknown. Here, we present evidence from biochemical experiments arguing that O-GlcNAcylation of NSL3 at Thr755 is tightly associated with holoenzyme activity of the MOF/NSL complex. Using in vitro O-GlcNAc-transferase assays combined with mass spectrometry, we suppose that the residue Thr755 on NSL3 C-terminus is the major site O-GlcNAc-modified by OGT1. Importantly, O-GlcNAcylation of this site is involved in the regulation of the ubiquitin-degradation of NSL3, because this site mutation (T755A) promotes the ubiquitin-mediated degradation of NSL3. Further in-depth research found that ubiquitin conjugating enzyme E2 S (UBE2S) accelerated the degradation of NSL3 via direct binding to it. Interestingly, OGT1 and UBE2S competitively bind to NSL3, suggesting the coordination of OGT1–UBE2S in regulating NSL3 stability. Furthermore, O-GlcNAcylation of NSL3 Thr755 site regulates the histone H4 acetylation levels at lysine 5, 8, and 16, suggesting that the O-GlcNAcylation of NSL3 at Thr755 is required for maintaining the integrity and holoenzyme activity of the MOF/NSL complex. In colony formation assays, we found that the integrity of the complex impacts the proliferation of the lung carcinoma type II epithelium-like A549 cells. Taken together, our results provide new insight into the elucidation of the molecular mechanism of the MOF/NSL complex.

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“…Precise regulation is achieved through the control of 2 E1, more than 30 E2, and more than 600 E3 enzymes (Buetow and Huang, 2016). Lorenz and coworkers demonstrate that the cell cycle regulator UBE2S contains a self-inactivation mechanism that may be applicable to nearly a quarter of the E2s (Liess et al, 2019).…”

mentioning

confidence: 99%

“…The latter is postulated to occur through autoubiquitination of a lysine-rich C-terminal extension. In this issue of Structure, Lorenz and colleagues demonstrate that regulation of UBE2S and other E2s deserves more attention as autoubiquitination of the UBC domain results in autoinhibition but not proteasomal degradation, suggesting a dynamic and responsive self-regulatory mechanism (Liess et al, 2019).…”

mentioning

confidence: 99%

“…This poses a challenge for the full mechanistic characterization of the enzyme as it is often unclear whether this autoubiquitination event is relevant or a side product of reconstituted reactions. To tackle this problem, Liess et al (2019) use a suite of methodologies, including in vitro ubiquitination assays, molecular dynamics simulations, NMR, quantitative mass spectrometry, immunoprecipitation, and cell-based assays, to investigate the significance of a specific autoubiquitination event on UBE2S (Liess et al, 2019).…”

mentioning

confidence: 99%

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UBE2S Learns Self-Control

Bodrug

Brown

2019

Structure

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Autoinhibition Mechanism of the Ubiquitin-Conjugating Enzyme UBE2S by Autoubiquitination

Cited by 22 publications

References 103 publications

The UBA domain of conjugating enzyme Ubc1/Ube2K facilitates assembly of K48/K63‐branched ubiquitin chains

The UBA domain of conjugating enzyme Ubc1/Ube2K facilitates assembly of K48/K63‐branched ubiquitin chains

O-GlcNAc-Modification of NSL3 at Thr755 Site Maintains the Holoenzyme Activity of MOF/NSL Histone Acetyltransferase Complex

UBE2S Learns Self-Control

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