Ubiquitination site preferences in anaphase promoting complex/cyclosome (APC/C) substrates

Min, Mingwei; Mayor, Ugo; Lindon, Catherine

doi:10.1098/rsob.130097

Cited by 41 publications

(51 citation statements)

References 57 publications

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“…This leads us to speculate that a conformationally fluctuating surface/region should increase the probability of fruitful Ub transfer to multiple substrate lysines. The APC/C was shown to prefer lysines in disordered regions for ubiquitination (87). Similar observations were also made by bioinformatics studies, suggesting a bias for degradation-linked ubiquitination sites to be more disordered when compared with unmodified lysines (88).…”

Section: Ubiquitin-acceptor Lysines On Substrates and Correlation Witsupporting

confidence: 67%

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

Guharoy

Bhowmick

Tompa

2016

Journal of Biological Chemistry

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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.Many cellular pathways and regulatory networks require spatial and temporal control of effector protein levels. Regulated degradation mediated by the ubiquitin-proteasome system (UPS) 3 is an important post-translational mechanism that helps to achieve precise fine-tuning of protein levels and is being increasingly linked to more and more pathways. The UPS is the major intracellular degradation pathway that has evolved into a complex system consisting of several hundred dedicated components (1). Important examples of regulated degradation include cell cycle regulatory proteins (e.g. cyclins, cyclin-dependent kinase inhibitors, etc.) that need to be degraded or inactivated before cell cycle checkpoint mechanisms decide upon progress (2, 3). Transcription factors (e.g. mammalian Myc, Jun, E2-F, p53, etc.) that activate gene expression triggered by specific stimuli are usually maintained at low levels (4, 5); further, the ubiquitin system also triggers processing (by limited proteolysis) and activation of transcription factors such as NFB (6). Cell surface growth factor/hormone receptors undergo internalization and degradation to switch off signaling inputs (7-12). The UPS also tightly regulates other key intracellular effectors (e.g. Smad proteins, Bcl-2) of signaling pathways (13-15). Not surprisingly, defects in regulated degradation are being linked to increasing numbers of diseases, including neurodegeneration and cancer, making the UPS very attractive for drug design (16 -19).Ubiquitination is organized as a cascade of E1 (ubiquitinactivating), E2 (ubiquitin-conjugating), and E3 (ubiquitin ligase) enzymes. The pathway is strongly conserved from yeast to mammals. The E1-E2-E3 axis has a pyramidal structure, with 2 E1 proteins (in humans), 30 -40 E2 enzymes, and Ͼ600 E3 ligases (20). E3s are subclassified into major groups based on their subunit organization and domain composition ( Fig. 1) (21,...

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Section: Ubiquitin-acceptor Lysines On Substrates and Correlation Witsupporting

confidence: 67%

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

Guharoy

Bhowmick

Tompa

2016

Journal of Biological Chemistry

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“…Furthermore, given the modular architecture and flexibility of disordered regions, acceptor lysines and degrons may not need to be in the same region of the protein. Sequence context of the ubiquitinated lysine might also contribute to the rate of substrate modification (Williamson et al, 2011; Min et al, 2013; Mattiroli and Sixma, 2014). Given the numerous enzymes that can modify lysines, blocking lysine accessibility through competitive modification of sites can be a powerful mechanism to control substrate degradation.…”

Section: Substrate Ordering By the Apc/cmentioning

confidence: 99%

Building a Regulatory Network with Short Linear Sequence Motifs: Lessons from the Degrons of the Anaphase-Promoting Complex

2016

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Summary The anaphase-promoting complex or cyclosome (APC/C) is a ubiquitin ligase that polyubiquitinates specific substrates at precise times in the cell cycle, thereby triggering the events of late mitosis in a strict order. The robust substrate specificity of the APC/C prevents the potentially deleterious degradation of non-APC/C substrates, and also averts the cell cycle errors and genomic instability that could result from mistimed degradation of APC/C targets. The APC/C recognizes short linear sequence motifs, or degrons, on its substrates. The specific and timely modification and degradation of APC/C substrates is likely to be modulated by variations in degron sequence and context. We discuss the extensive affinity, specificity and selectivity determinants encoded in APC/C degrons, and we describe some of the extrinsic mechanisms that control APC/C-substrate recognition. As an archetype for protein motif-driven regulation of cell function, the APC/C-substrate interaction provides insights into the general properties of post-translational regulatory systems.

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“…KIFC1-Venus electroporated into bio Ub cells synchronized in M and C phases as in C was subjected to GFP-Trap® pulldown as described in Ref. 57. Anti-GFP immunoblot revealed unmodified KIFC1-Venus (in green), whereas anti-biotin immunoblot revealed KIFC1-Venus-associated ubiquitin conjugates (in red).…”

Section: New Functions For Apc/c Targeting During Mitotic Exit-mentioning

confidence: 99%

“…Ubiquitination of cellular components during mitotic exit. A, enrichment analysis of the D-box and the KEN box in mitotic-exitspecific hits compared with non-phase-specific hits and a dataset of known APC/C substrates curated from the literature (57). The sequence of each protein was scanned for the presence of potential D-box (RXXL) and KEN motifs.…”

Section: Fig 5 Apc/c Suppresses Cytoplasmic Racgap1 To Promote Cellmentioning

confidence: 99%

Using in Vivo Biotinylated Ubiquitin to Describe a Mitotic Exit Ubiquitome from Human Cells

Min

Mayor

Dittmar

et al. 2014

Molecular & Cellular Proteomics

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Mitotic division requires highly regulated morphological and biochemical changes to the cell. Upon commitment to exit mitosis, cells begin to remove mitotic regulators in a temporally and spatially controlled manner to bring about the changes that reestablish interphase. Ubiquitindependent pathways target these regulators to generate polyubiquitin-tagged substrates for degradation by the 26S proteasome. However, the lack of cell-based assays to investigate in vivo ubiquitination limits our knowledge of the identity of substrates of ubiquitin-mediated regulation in mitosis. Here we report an in vivo ubiquitin tagging system used in human cells that allows efficient purification of ubiquitin conjugates from synchronized cell populations. Coupling purification with mass spectrometry, we have identified a series of mitotic regulators targeted for polyubiquitination in mitotic exit. We show that some are new substrates of the anaphase-promoting complex/ cyclosome and validate KIFC1 and RacGAP1/Cyk4 as two such targets involved respectively in timely mitotic spindle disassembly and cell spreading. We conclude that in vivo biotin tagging of ubiquitin can provide valuable information about the role of ubiquitin-mediated regulation in processes required for rebuilding interphase cells. Molecular & Cellular

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Ubiquitination site preferences in anaphase promoting complex/cyclosome (APC/C) substrates

Cited by 41 publications

References 57 publications

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

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

Building a Regulatory Network with Short Linear Sequence Motifs: Lessons from the Degrons of the Anaphase-Promoting Complex

Using in Vivo Biotinylated Ubiquitin to Describe a Mitotic Exit Ubiquitome from Human Cells

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