Components of the Spindle Assembly Checkpoint Regulate the Anaphase-Promoting Complex During Meiosis in Caenorhabditis elegans

ABSTRACT Polo-like kinase 1 (Plk1) plays a role in numerous events in mitosis, but how the multiple functions of Plk1 are separated is poorly understood. We studied regulation of Plk1 through two putative phosphorylation residues, Ser-137 and Thr-210. Using phospho-specific antibodies, we found that Thr-210 phosphorylation precedes Ser-137 phosphorylation in vivo, the latter occurring specifically in late mitosis. We show that expression of two activating mutants of these residues, S137D and T210D, results in distinct mitotic phenotypes. Whereas expression of both phospho-mimicking mutants as well as of the double mutant leads to accelerated mitotic entry, further progression through mitosis is dramatically different: the T210D mutant causes a spindle assembly checkpoint-dependent delay, whereas the expression of the S137D mutant or the double mutant results in untimely activation of the anaphase-promoting complex/cyclosome (APC/C) and frequent mitotic catastrophe. Using nonphosphorylatable Plk1-S137A and Plk1-T210A mutants, we show that both sites contribute to proper mitotic progression. Based on these observations, we propose that Plk1 function is altered at different stages of mitosis through consecutive posttranslational events, e.g., at Ser-137 and Thr-210. Furthermore, our data show that uncontrolled Plk1 activation can uncouple APC/C activity from spindle assembly checkpoint control.

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

The Journal of Cell Biology ◽

10.1083/jcb.200801049 ◽

2008 ◽

Vol 183 (2) ◽

pp. 267-277 ◽

Cited By ~ 42

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.

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Regulated Separation of Sister Centromeres depends on the Spindle Assembly Checkpoint but not on the Anaphase Promoting Complex/Cyclosome

Cell Cycle ◽

10.4161/cc.4.11.2146 ◽

2005 ◽

Vol 4 (11) ◽

pp. 1561-1575 ◽

Cited By ~ 21

Author(s):

Juan F. Gimenez-Abian ◽

Laura A. Díaz-Martínez ◽

Karin G. Wirth ◽

Catherine A. Andrews ◽

Gonzalo Giménez-Martín ◽

...

Keyword(s):

Spindle Assembly Checkpoint ◽

Spindle Assembly ◽

Anaphase Promoting Complex

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Spindle Assembly Checkpoint Protein Cdc20 Transcriptionally Activates Expression of Ubiquitin Carrier Protein UbcH10

Journal of Biological Chemistry ◽

10.1074/jbc.m110.160671 ◽

2011 ◽

Vol 286 (18) ◽

pp. 15666-15677 ◽

Cited By ~ 12

Author(s):

Somsubhra Nath ◽

Taraswi Banerjee ◽

Debrup Sen ◽

Tania Das ◽

Susanta Roychoudhury

Keyword(s):

Chromosomal Instability ◽

Spindle Assembly Checkpoint ◽

Spindle Assembly ◽

Regulatory Function ◽

Physical Interaction ◽

Anaphase Promoting Complex ◽

Transcriptional Regulatory ◽

Subsequent Degradation ◽

Ubiquitin Conjugating Enzyme ◽

Wd40 Domain

The spindle assembly checkpoint (SAC) ensures accurate segregation of chromosomes by monitoring kinetochore attachment of spindles during mitosis. Proper progression of mitosis depends on orderly ubiquitination and subsequent degradation of various mitotic inhibitors. At the molecular level, upon removal of SAC, Cdc20 activates E3 ubiquitin ligase anaphase-promoting complex/cyclosome that, along with E2 ubiquitin-conjugating enzyme UbcH10, executes this function. Both Cdc20 and UbcH10 are overexpressed in many cancer types and are associated with defective SAC function leading to chromosomal instability. The precise mechanism of correlated overexpression of these two proteins remains elusive. We show that Cdc20 transcriptionally up-regulates UbcH10 expression. The WD40 domain of Cdc20 is required for this activity. Physical interaction between Cdc20 and anaphase-promoting complex/cyclosome-CBP/p300 complex and its subsequent recruitment to the UBCH10 promoter are involved in this transactivation process. This transcriptional regulatory function of Cdc20 was observed to be cell cycle-specific. We hypothesize that this co-regulated overexpression of both proteins contributes to chromosomal instability.

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Slip slidin’ away of mitosis with CRL2Zyg11

The Journal of Cell Biology ◽

10.1083/jcb.201609086 ◽

2016 ◽

Vol 215 (2) ◽

pp. 143-145 ◽

Cited By ~ 1

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.

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EFFECTORS OF THE SPINDLE ASSEMBLY CHECKPOINT BUT NOT THE MITOTIC EXIT NETWORK ARE CONFINED WITHIN THE NUCLEUS OF SACCHAROMYCES CEREVISIAE

10.1101/340091 ◽

2018 ◽

Author(s):

Lydia R Heasley ◽

Jennifer G DeLuca ◽

Steven M Markus

Keyword(s):

Nuclear Envelope ◽

Mammalian Cells ◽

Spindle Assembly Checkpoint ◽

Spindle Assembly ◽

Mitotic Checkpoint ◽

Mitotic Exit ◽

Cell Cycle Checkpoints ◽

Anaphase Promoting Complex ◽

Mitotic Exit Network ◽

And Function

The Spindle Assembly Checkpoint (SAC) prevents erroneous chromosome segregation by delaying mitotic progression when chromosomes are incorrectly attached to the mitotic spindle. This delay is mediated by Mitotic Checkpoint Complexes (MCCs), which assemble at unattached kinetochores and repress the activity of the Anaphase Promoting Complex/Cyclosome (APC/C). The cellular localizations of MCCs are likely critical for proper SAC function, yet remain poorly defined. We recently demonstrated that in mammalian cells, in which the nuclear envelope disassembles during mitosis, MCCs diffuse throughout the spindle region and cytoplasm. Here, we employed binucleate yeast zygotes to examine the localization dynamics of SAC effectors required for MCC assembly and function in budding yeast, in which the nuclear envelope remains intact throughout mitosis. Our findings indicate that in yeast MCCs are confined to the nuclear compartment and excluded from the cytoplasm during mitosis. In contrast, we find that effectors of the Mitotic Exit Network (MEN) - a pathway that initiates disassembly of the anaphase spindle only when it is properly oriented - are in fact freely exchanged between multiple nuclei within a shared cytoplasm. Our study provides insight into how cell cycle checkpoints have evolved to function in diverse cellular contexts.

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Dissection of the NUP107 nuclear pore subcomplex reveals a novel interaction with spindle assembly checkpoint protein MAD1 in Caenorhabditis elegans

Molecular Biology of the Cell ◽

10.1091/mbc.e11-11-0927 ◽

2012 ◽

Vol 23 (5) ◽

pp. 930-944 ◽

Cited By ~ 31

Author(s):

Eduardo Ródenas ◽

Cristina González-Aguilera ◽

Cristina Ayuso ◽

Peter Askjaer

Keyword(s):

Caenorhabditis Elegans ◽

Nuclear Envelope ◽

Molecular Mechanisms ◽

Spindle Assembly Checkpoint ◽

Protein Import ◽

Spindle Assembly ◽

Nucleocytoplasmic Transport ◽

Nuclear Pore ◽

Nuclear Pore Complexes ◽

Pore Complexes

Nuclear pore complexes consist of several subcomplexes. The NUP107 complex is important for nucleocytoplasmic transport, nuclear envelope assembly, and kinetochore function. However, the underlying molecular mechanisms and the roles of individual complex members remain elusive. We report the first description of a genetic disruption of NUP107 in a metazoan. Caenorhabditis elegans NUP107/npp-5 mutants display temperature-dependent lethality. Surprisingly, NPP-5 is dispensable for incorporation of most nucleoporins into nuclear pores and for nuclear protein import. In contrast, NPP-5 is essential for proper kinetochore localization of NUP133/NPP-15, another NUP107 complex member, whereas recruitment of NUP96/NPP-10C and ELYS/MEL-28 is NPP-5 independent. We found that kinetochore protein NUF2/HIM-10 and Aurora B/AIR-2 kinase are less abundant on mitotic chromatin upon NPP-5 depletion. npp-5 mutants are hypersensitive to anoxia, suggesting that the spindle assembly checkpoint (SAC) is compromised. Indeed, NPP-5 interacts genetically and physically with SAC protein MAD1/MDF-1, whose nuclear envelope accumulation requires NPP-5. Thus our results strengthen the emerging connection between nuclear pore proteins and chromosome segregation.

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