scholarly journals The HECT E3 ligase Smurf2 is required for Mad2-dependent spindle assembly checkpoint

2008 ◽  
Vol 183 (2) ◽  
pp. 267-277 ◽  
Author(s):  
Evan C. Osmundson ◽  
Dipankar Ray ◽  
Finola E. Moore ◽  
Qingshen Gao ◽  
Gerald H. Thomsen ◽  
...  
Keyword(s):  
Ubiquitin Ligase ◽  
Spindle Assembly Checkpoint ◽  
Spindle Checkpoint ◽  
E3 Ligase ◽  
Spindle Assembly ◽  
Cyclin B ◽  
Carboxyl Terminus ◽  
Anaphase Promoting Complex ◽  
Mitotic Spindles ◽  
Anaphase Onset

Activation of the anaphase-promoting complex/cyclosome (APC/C) by Cdc20 is critical for the metaphase–anaphase transition. APC/C-Cdc20 is required for polyubiquitination and degradation of securin and cyclin B at anaphase onset. The spindle assembly checkpoint delays APC/C-Cdc20 activation until all kinetochores attach to mitotic spindles. In this study, we demonstrate that a HECT (homologous to the E6-AP carboxyl terminus) ubiquitin ligase, Smurf2, is required for the spindle checkpoint. Smurf2 localizes to the centrosome, mitotic midbody, and centromeres. Smurf2 depletion or the expression of a catalytically inactive Smurf2 results in misaligned and lagging chromosomes, premature anaphase onset, and defective cytokinesis. Smurf2 inactivation prevents nocodazole-treated cells from accumulating cyclin B and securin and prometaphase arrest. The silencing of Cdc20 in Smurf2-depleted cells restores mitotic accumulation of cyclin B and securin. Smurf2 depletion results in enhanced polyubiquitination and degradation of Mad2, a critical checkpoint effector. Mad2 is mislocalized in Smurf2-depleted cells, suggesting that Smurf2 regulates the localization and stability of Mad2. These data indicate that Smurf2 is a novel mitotic regulator.

scholarly journals Sumoylation promotes optimal APC/C activation and timely anaphase

eLife ◽  
2018 ◽  
Vol 7 ◽  
Author(s):  
Christine C Lee ◽  
Bing Li ◽  
Hongtao Yu ◽  
Michael J Matunis
Keyword(s):  
Spindle Assembly Checkpoint ◽  
E3 Ligase ◽  
Spindle Assembly ◽  
Mitotic Exit ◽  
Regulatory Role ◽  
Anaphase Promoting Complex ◽  
Catalytic Core ◽  
Sumo Modification ◽  
Ubiquitin E3 Ligase ◽  
Anaphase Onset

The Anaphase Promoting Complex/Cyclosome (APC/C) is a ubiquitin E3 ligase that functions as the gatekeeper to mitotic exit. APC/C activity is controlled by an interplay of multiple pathways during mitosis, including the spindle assembly checkpoint (SAC), that are not yet fully understood. Here, we show that sumoylation of the APC4 subunit of the APC/C peaks during mitosis and is critical for timely APC/C activation and anaphase onset. We have also identified a functionally important SUMO interacting motif in the cullin-homology domain of APC2 located near the APC4 sumoylation sites and APC/C catalytic core. Our findings provide evidence of an important regulatory role for SUMO modification and binding in affecting APC/C activation and mitotic exit.

scholarly journals Slip slidin’ away of mitosis with CRL2Zyg11

2016 ◽  
Vol 215 (2) ◽  
pp. 143-145 ◽  
Author(s):  
Michael Brandeis
Keyword(s):  
Spindle Assembly Checkpoint ◽  
E3 Ligase ◽  
Spindle Assembly ◽  
Cyclin B1 ◽  
Anaphase Promoting Complex ◽  
Cell Biol ◽  
Mitotic Slippage ◽  
Mitotic Cells

The spindle assembly checkpoint arrests mitotic cells by preventing degradation of cyclin B1 by the anaphase-promoting complex/cyclosome, but some cells evade this checkpoint and slip out of mitosis. Balachandran et al. (2016. J. Cell Biol. http://dx.doi.org/10.1083/jcb.201601083) show that the E3 ligase CRL2ZYG11 degrades cyclin B1, allowing mitotic slippage.

scholarly journals Recruitment of Cdc20 to the Kinetochore Requires BubR1 but Not Mad2 in Drosophila melanogaster

2010 ◽  
Vol 30 (13) ◽  
pp. 3384-3395 ◽  
Author(s):  
Deyu Li ◽  
Gary Morley ◽  
Michael Whitaker ◽  
Jun-Yong Huang
Keyword(s):  
Drosophila Melanogaster ◽  
Rna Interference ◽  
Spindle Assembly Checkpoint ◽  
Spindle Assembly ◽  
Cyclin B ◽  
Stable Complex ◽  
S2 Cells ◽  
Anaphase Promoting Complex ◽  
Surveillance Mechanism ◽  
Initial Binding

ABSTRACT To prevent aneuploidy, cells require a mitotic surveillance mechanism, the spindle assembly checkpoint (SAC). The SAC prevents metaphase/anaphase transition by blocking the ubiquitylation and destruction of cyclin B and securin via the Cdc20-activated anaphase-promoting complex or cyclosome (APC/C)-mediated proteolysis pathway. This checkpoint involves the kinetochore proteins Mad2, BubR1, and Cdc20. Mad2 and BubR1 are inhibitors of the APC/C, but Cdc20 is an activator. Exactly how the SAC regulates Cdc20 via unattached kinetochores remains unclear; in vertebrates, most current models suggest that kinetochore-bound Mad2 is required for initial binding to Cdc20 to form a stable complex that includes BubR1. Here, we show that the Mad2 kinetochore dimerization recruitment mechanism is conserved and that the recruitment of Cdc20 to kinetochores in Drosophila requires BubR1 but not Mad2. BubR1 and Mad2 can bind to Cdc20 independently, and the interactions are enhanced after cells are arrested at mitosis by the depletion of Cdc27 using RNA interference (RNAi) in S2 cells or by MG132 treatment in syncytial embryos. These findings offer an explanation of why BubR1 is more important than Mad2 for SAC function in flies. These findings could lead to a better understanding of vertebrate SAC mechanisms.

scholarly journals How cyclin A destruction escapes the spindle assembly checkpoint

2010 ◽  
Vol 190 (4) ◽  
pp. 501-509 ◽  
Author(s):  
Barbara Di Fiore ◽  
Jonathon Pines
Keyword(s):  
Ubiquitin Ligase ◽  
Spindle Assembly Checkpoint ◽  
Spindle Assembly ◽  
Cyclin A ◽  
Cyclin Dependent Kinase ◽  
Anaphase Promoting Complex ◽  
N Terminus ◽  
Specific Proteins ◽  
Necessary And Sufficient

The anaphase-promoting complex/cyclosome (APC/C) is the ubiquitin ligase essential to mitosis, which ensures that specific proteins are degraded at specific times to control the order of mitotic events. The APC/C coactivator, Cdc20, is targeted by the spindle assembly checkpoint (SAC) to restrict APC/C activity until metaphase, yet early substrates, such as cyclin A, are degraded in the presence of the active checkpoint. Cdc20 and the cyclin-dependent kinase cofactor, Cks, are required for cyclin A destruction, but how they enable checkpoint-resistant destruction has not been elucidated. In this study, we answer this problem: we show that the N terminus of cyclin A binds directly to Cdc20 and with sufficient affinity that it can outcompete the SAC proteins. Subsequently, the Cks protein is necessary and sufficient to promote cyclin A degradation in the presence of an active checkpoint by binding cyclin A–Cdc20 to the APC/C.

scholarly journals Coupling of Cdc20 inhibition and activation by BubR1

2020 ◽  
Author(s):  
Jamin Hein ◽  
Dimitriya H Garvanska ◽  
Isha Nasa ◽  
Arminja Kettenbach ◽  
Jakob Nilsson
Keyword(s):  
Ubiquitin Ligase ◽  
Cross Talk ◽  
Spindle Assembly Checkpoint ◽  
Spindle Assembly ◽  
Cyclin B1 ◽  
Mitotic Checkpoint ◽  
Anaphase Onset ◽  
Mitotic Checkpoint Complex

Tight regulation of the APC/C-Cdc20 ubiquitin ligase that targets Cyclin B1 for degradation is important for mitotic fidelity. The spindle assembly checkpoint (SAC) inhibits Cdc20 through the mitotic checkpoint complex (MCC). In addition, phosphorylation of Cdc20 by Cyclin B1-Cdk1 independently inhibits APC/C-Cdc20 activation. This creates a conundrum for how Cdc20 gets activated prior to Cyclin B1 degradation. Here we show that the MCC component BubR1 harbours both Cdc20 inhibition and activation activities, allowing for cross-talk between the two Cdc20 inhibition pathways. Specifically BubR1 acts as a substrate specifier for PP2A-B56 to enable efficient Cdc20 dephosphorylation in the MCC. A mutant Cdc20 mimicking the dephosphorylated state escapes a mitotic checkpoint arrest arguing that restricting Cdc20 dephosphorylation to the MCC is important. Collectively our work reveals how Cdc20 can be dephosphorylated in the presence of Cyclin B1-Cdk1 activity without causing premature anaphase onset.

scholarly journals Chromosome biorientation and APC activity remain uncoupled in oocytes with reduced volume

2017 ◽  
Vol 216 (12) ◽  
pp. 3949-3957 ◽  
Author(s):  
Simon I.R. Lane ◽  
Keith T. Jones
Keyword(s):  
Cell Volume ◽  
Chromosome Segregation ◽  
Spindle Assembly Checkpoint ◽  
Spindle Assembly ◽  
Anaphase Promoting Complex ◽  
Chromosome Missegregation ◽  
Anaphase Onset ◽  
Reduced Volume ◽  
Chromosome Attachment ◽  
Meiosis I

The spindle assembly checkpoint (SAC) prevents chromosome missegregation by coupling anaphase onset with correct chromosome attachment and tension to microtubules. It does this by generating a diffusible signal from free kinetochores into the cytoplasm, inhibiting the anaphase-promoting complex (APC). The volume in which this signal remains effective is unknown. This raises the possibility that cell volume may be the reason the SAC is weak, and chromosome segregation error-prone, in mammalian oocytes. Here, by a process of serial bisection, we analyzed the influence of oocyte volume on the ability of the SAC to inhibit bivalent segregation in meiosis I. We were able to generate oocytes with cytoplasmic volumes reduced by 86% and observed changes in APC activity consistent with increased SAC control. However, bivalent biorientation remained uncoupled from APC activity, leading to error-prone chromosome segregation. We conclude that volume is one factor contributing to SAC weakness in oocytes. However, additional factors likely uncouple chromosome biorientation with APC activity.

scholarly journals p31comet acts to ensure timely spindle checkpoint silencing subsequent to kinetochore attachment

2011 ◽  
Vol 22 (22) ◽  
pp. 4236-4246 ◽  
Author(s):  
Robert S. Hagan ◽  
Michael S. Manak ◽  
Håkon Kirkeby Buch ◽  
Michelle G. Meier ◽  
Patrick Meraldi ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Spindle Assembly Checkpoint ◽  
Spindle Assembly ◽  
Mitotic Exit ◽  
Cyclin B ◽  
Anaphase Promoting Complex ◽  
Mitotic Progression ◽  
Microtubule Attachment ◽  
And Function ◽  
High Cyclin

The spindle assembly checkpoint links the onset of anaphase to completion of chromosome-microtubule attachment and is mediated by the binding of Mad and Bub proteins to kinetochores of unattached or maloriented chromosomes. Mad2 and BubR1 traffic between kinetochores and the cytosol, thereby transmitting a “wait anaphase” signal to the anaphase-promoting complex. It is generally assumed that this signal dissipates automatically upon kinetochore-microtubule binding, but it has been shown that under conditions of nocodazole-induced arrest p31comet, a Mad2-binding protein, is required for mitotic progression. In this article we investigate the localization and function of p31comet during normal, unperturbed mitosis in human and marsupial cells. We find that, like Mad2, p31comet traffics on and off kinetochores and is also present in the cytosol. Cells depleted of p31comet arrest in metaphase with mature bipolar kinetochore-microtubule attachments, a satisfied checkpoint, and high cyclin B levels. Thus p31comet is required for timely mitotic exit. We propose that p31comet is an essential component of the machinery that silences the checkpoint during each cell cycle.

scholarly journals Kinetochore stretching inactivates the spindle assembly checkpoint

2009 ◽  
Vol 184 (3) ◽  
pp. 383-390 ◽  
Author(s):  
Kazuhiko S.K. Uchida ◽  
Kentaro Takagaki ◽  
Kazuki Kumada ◽  
Youko Hirayama ◽  
Tetsuo Noda ◽  
...  
Keyword(s):  
Hela Cell ◽  
Live Cell Imaging ◽  
Spindle Assembly Checkpoint ◽  
Cell Imaging ◽  
Protein A ◽  
Spindle Assembly ◽  
Cyclin B ◽  
Anaphase Promoting Complex ◽  
Hela Cell Line ◽  
Centromere Protein A

The spindle assembly checkpoint (SAC) monitors the attachment of microtubules to the kinetochore and inhibits anaphase when microtubule binding is incomplete. The SAC might also respond to tension; however, how cells can sense tension and whether its detection is important to satisfy the SAC remain controversial. We generated a HeLa cell line in which two components of the kinetochore, centromere protein A and Mis12, are labeled with green and red fluorophores, respectively. Live cell imaging of these cells reveals repetitive cycles of kinetochore extension and recoiling after biorientation. Under conditions in which kinetochore stretching is suppressed, cells fail to silence the SAC and enter anaphase after a delay, regardless of centromere stretching. Monitoring cyclin B levels as a readout for anaphase-promoting complex/cyclosome activity, we find that suppression of kinetochore stretching delays and decelerates cyclin B degradation. These observations suggest that the SAC monitors stretching of kinetochores rather than centromeres and that kinetochore stretching promotes silencing of the SAC signal.

scholarly journals EMB-30: An APC4 Homologue Required for Metaphase-to-Anaphase Transitions during Meiosis and Mitosis in Caenorhabditis elegans

2000 ◽  
Vol 11 (4) ◽  
pp. 1401-1419 ◽  
Author(s):  
Tokiko Furuta ◽  
Simon Tuck ◽  
Jay Kirchner ◽  
Bryan Koch ◽  
Roy Auty ◽  
...  
Keyword(s):  
Positional Cloning ◽  
Spindle Assembly Checkpoint ◽  
Spindle Assembly ◽  
Class I ◽  
Meiotic Spindle ◽  
Anaphase Promoting Complex ◽  
Anaphase Onset ◽  
Polar Bodies ◽  
M Phase ◽  
Set Up

Here we show that emb-30 is required for metaphase-to-anaphase transitions during meiosis and mitosis inCaenorhabditis elegans. Germline-specificemb-30 mutant alleles block the meiotic divisions. Mutant oocytes, fertilized by wild-type sperm, set up a meiotic spindle but do not progress to anaphase I. As a result, polar bodies are not produced, pronuclei fail to form, and cytokinesis does not occur. Severe-reduction-of-function emb-30 alleles (class I alleles) result in zygotic sterility and lead to germline and somatic defects that are consistent with an essential role in promoting the metaphase-to-anaphase transition during mitosis. Analysis of the vulval cell lineages in these emb-30(class I) mutant animals suggests that mitosis is lengthened and eventually arrested when maternally contributed emb-30 becomes limiting. By further reducing maternal emb-30 function contributed to class I mutant animals, we show that emb-30 is required for the metaphase-to-anaphase transition in many, if not all, cells. Metaphase arrest in emb-30 mutants is not due to activation of the spindle assembly checkpoint but rather reflects an essential emb-30 requirement for M-phase progression. A reduction in emb-30 activity can suppress the lethality and sterility caused by a null mutation in mdf-1, a component of the spindle assembly checkpoint machinery. This result suggests that delaying anaphase onset can bypass the spindle checkpoint requirement for normal development. Positional cloning established thatemb-30 encodes the likely C. elegansorthologue of APC4/Lid1, a component of the anaphase-promoting complex/cyclosome, required for the metaphase-to-anaphase transition. Thus, the anaphase-promoting complex/cyclosome is likely to be required for all metaphase-to-anaphase transitions in a multicellular organism.

scholarly journals Coupling of Cdc20 inhibition and activation by BubR1

2021 ◽  
Vol 220 (5) ◽  
Author(s):  
Jamin B. Hein ◽  
Dimitriya H. Garvanska ◽  
Isha Nasa ◽  
Arminja N. Kettenbach ◽  
Jakob Nilsson
Keyword(s):  
Ubiquitin Ligase ◽  
Cross Talk ◽  
Spindle Assembly Checkpoint ◽  
Spindle Assembly ◽  
Cyclin B1 ◽  
Mitotic Checkpoint ◽  
Anaphase Onset ◽  
Mitotic Checkpoint Complex

Tight regulation of the APC/C-Cdc20 ubiquitin ligase that targets cyclin B1 for degradation is important for mitotic fidelity. The spindle assembly checkpoint (SAC) inhibits Cdc20 through the mitotic checkpoint complex (MCC). In addition, phosphorylation of Cdc20 by cyclin B1–Cdk1 independently inhibits APC/C–Cdc20 activation. This creates a conundrum for how Cdc20 is activated before cyclin B1 degradation. Here, we show that the MCC component BubR1 harbors both Cdc20 inhibition and activation activities, allowing for cross-talk between the two Cdc20 inhibition pathways. Specifically, BubR1 acts as a substrate specifier for PP2A-B56 to enable efficient Cdc20 dephosphorylation in the MCC. A mutant Cdc20 mimicking the dephosphorylated state escapes a mitotic checkpoint arrest, arguing that restricting Cdc20 dephosphorylation to the MCC is important. Collectively, our work reveals how Cdc20 can be dephosphorylated in the presence of cyclin B1-Cdk1 activity without causing premature anaphase onset.

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