scholarly journals Cyclin B3 activates the Anaphase-Promoting Complex/Cyclosome in meiosis and mitosis

PLoS Genetics ◽  
2020 ◽  
Vol 16 (11) ◽  
pp. e1009184
Author(s):  
Damien Garrido ◽  
Mohammed Bourouh ◽  
Éric Bonneil ◽  
Pierre Thibault ◽  
Andrew Swan ◽  
...  
Keyword(s):  
Spindle Assembly Checkpoint ◽  
Regulatory Subunit ◽  
Cyclin Dependent Kinase ◽  
Anaphase Promoting Complex ◽  
Loss Of Function ◽  
Metaphase Arrest ◽  
Phosphatase 2A ◽  
Cyclin B3 ◽  
Mitosis And Meiosis

In mitosis and meiosis, chromosome segregation is triggered by the Anaphase-Promoting Complex/Cyclosome (APC/C), a multi-subunit ubiquitin ligase that targets proteins for degradation, leading to the separation of chromatids. APC/C activation requires phosphorylation of its APC3 and APC1 subunits, which allows the APC/C to bind its co-activator Cdc20. The identity of the kinase(s) responsible for APC/C activation in vivo is unclear. Cyclin B3 (CycB3) is an activator of the Cyclin-Dependent Kinase 1 (Cdk1) that is required for meiotic anaphase in flies, worms and vertebrates. It has been hypothesized that CycB3-Cdk1 may be responsible for APC/C activation in meiosis but this remains to be determined. Using Drosophila, we found that mutations in CycB3 genetically enhance mutations in tws, which encodes the B55 regulatory subunit of Protein Phosphatase 2A (PP2A) known to promote mitotic exit. Females heterozygous for CycB3 and tws loss-of-function alleles lay embryos that arrest in mitotic metaphase in a maternal effect, indicating that CycB3 promotes anaphase in mitosis in addition to meiosis. This metaphase arrest is not due to the Spindle Assembly Checkpoint (SAC) because mutation of mad2 that inactivates the SAC does not rescue the development of embryos from CycB3-/+, tws-/+ females. Moreover, we found that CycB3 promotes APC/C activity and anaphase in cells in culture. We show that CycB3 physically associates with the APC/C, is required for phosphorylation of APC3, and promotes APC/C association with its Cdc20 co-activators Fizzy and Cortex. Our results strongly suggest that CycB3-Cdk1 directly activates the APC/C to promote anaphase in both meiosis and mitosis.

scholarly journals Loss of a Protein Phosphatase 2A Regulatory Subunit (Cdc55p) Elicits Improper Regulation of Swe1p Degradation

2000 ◽  
Vol 20 (21) ◽  
pp. 8143-8156 ◽  
Author(s):  
Haifeng Yang ◽  
Wei Jiang ◽  
Matthew Gentry ◽  
Richard L. Hallberg
Keyword(s):  
Phosphatase Activity ◽  
Protein Phosphatase ◽  
Protein Phosphatase 2A ◽  
Cell Types ◽  
Regulatory Subunit ◽  
Cyclin Dependent Kinase ◽  
Wild Type ◽  
Phosphatase 2A

ABSTRACT CDC55 encodes a Saccharomyces cerevisiaeprotein phosphatase 2A (PP2A) regulatory subunit.cdc55-null cells growing at low temperature exhibit a failure of cytokinesis and produce abnormally elongated buds, butcdc55-null cells producing the cyclin-dependent kinase Cdc28-Y19F, which is unable to be inhibited by Y19 phosphorylation, show a loss of the abnormal morphology. Furthermore,cdc55-null cells exhibit a hyperphosphorylation of Y19. For these reasons, we have examined in wild-type and cdc55-null cells the levels and activities of the kinase (Swe1p) and phosphatase (Mih1p) that normally regulate the extent of Cdc28 Y19 phosphorylation. We find that Mih1p levels are comparable in the two strains, and an estimate of the in vivo and in vitro phosphatase activity of this enzyme in the two cell types indicates no marked differences. By contrast, while Swe1p levels are similar in unsynchronized and S-phase-arrested wild-type and cdc55-null cells, Swe1 kinase is found at elevated levels in mitosis-arrestedcdc55-null cells. This excess Swe1p incdc55-null cells is the result of ectopic stabilization of this protein during G2 and M, thereby accounting for the accumulation of Swe1p in mitosis-arrested cells. We also present evidence indicating that, in cdc55-null cells, misregulated PP2A phosphatase activity is the cause of both the ectopic stabilization of Swe1p and the production of the morphologically abnormal phenotype.

scholarly journals Cyclin B3 activates the Anaphase-Promoting Complex/Cyclosome in meiosis and mitosis

2020 ◽  
Author(s):  
Damien Garrido ◽  
Mohammed Bourouh ◽  
Éric Bonneil ◽  
Pierre Thibault ◽  
Andrew Swan ◽  
...  
Keyword(s):  
Embryonic Development ◽  
Chromosome Segregation ◽  
Ubiquitin Ligase ◽  
Anaphase Promoting Complex ◽  
Cells In Culture ◽  
Cyclin B3 ◽  
Mitosis And Meiosis ◽  
In Cells

ABSTRACTIn mitosis and meiosis, chromosome segregation is triggered by the Anaphase-Promoting Complex/Cyclosome (APC/C), a multi-subunit ubiquitin ligase that targets proteins for degradation, leading to the separation of chromatids. APC/C activation requires phosphorylation of its APC3 and APC1 subunits, which allows the APC/C to bind its Cdc20 co-activator. The identity of the kinase(s) responsible for APC/C activation in vivo is unclear. Cyclin B3 is required for meiotic anaphase in flies, worms and vertebrates, but whether it activates the APC/C is unclear. We found that Drosophila Cyclin B3 (CycB3) collaborates with PP2A-B55/Tws in embryonic development, indicating that CycB3 also promotes anaphase in mitosis. Moreover, CycB3 promotes APC/C activity and anaphase in cells in culture. We show that CycB3 physically associates with the APC/C, is required for phosphorylation of APC3, and promotes APC/C association with its co-activators. We propose that CycB3-Cdk1 directly phosphorylates the APC/C to activate it in both meiosis and mitosis.

scholarly journals A Wee1 checkpoint inhibits anaphase onset

2013 ◽  
Vol 201 (6) ◽  
pp. 843-862 ◽  
Author(s):  
Noel Lianga ◽  
Elizabeth C. Williams ◽  
Erin K. Kennedy ◽  
Carole Doré ◽  
Sophie Pilon ◽  
...  
Keyword(s):  
Budding Yeast ◽  
Regulatory Subunit ◽  
Anaphase Promoting Complex ◽  
Mitotic Entry ◽  
Anaphase Onset ◽  
Phosphatase 2A ◽  
Wee1 Kinase ◽  
Morphogenesis Checkpoint

Cdk1 drives both mitotic entry and the metaphase-to-anaphase transition. Past work has shown that Wee1 inhibition of Cdk1 blocks mitotic entry. Here we show that the budding yeast Wee1 kinase, Swe1, also restrains the metaphase-to-anaphase transition by preventing Cdk1 phosphorylation and activation of the mitotic form of the anaphase-promoting complex/cyclosome (APCCdc20). Deletion of SWE1 or its opposing phosphatase MIH1 (the budding yeast cdc25+) altered the timing of anaphase onset, and activation of the Swe1-dependent morphogenesis checkpoint or overexpression of Swe1 blocked cells in metaphase with reduced APC activity in vivo and in vitro. The morphogenesis checkpoint also depended on Cdc55, a regulatory subunit of protein phosphatase 2A (PP2A). cdc55Δ checkpoint defects were rescued by mutating 12 Cdk1 phosphorylation sites on the APC, demonstrating that the APC is a target of this checkpoint. These data suggest a model in which stepwise activation of Cdk1 and inhibition of PP2ACdc55 triggers anaphase onset.

scholarly journals The Role of Cdh1p in Maintaining Genomic Stability in Budding Yeast

Genetics ◽  
2003 ◽  
Vol 165 (2) ◽  
pp. 489-503 ◽  
Author(s):  
Karen E Ross ◽  
Orna Cohen-Fix
Keyword(s):  
Cell Cycle ◽  
Chromosome Segregation ◽  
Spindle Assembly Checkpoint ◽  
Chromosome Loss ◽  
Cyclin Dependent Kinase ◽  
Anaphase Promoting Complex ◽  
Growth Defects ◽  
Genes Encoding ◽  
Specificity Factor

Abstract Cdh1p, a substrate specificity factor for the cell cycle-regulated ubiquitin ligase, the anaphase-promoting complex/cyclosome (APC/C), promotes exit from mitosis by directing the degradation of a number of proteins, including the mitotic cyclins. Here we present evidence that Cdh1p activity at the M/G1 transition is important not only for mitotic exit but also for high-fidelity chromosome segregation in the subsequent cell cycle. CDH1 showed genetic interactions with MAD2 and PDS1, genes encoding components of the mitotic spindle assembly checkpoint that acts at metaphase to prevent premature chromosome segregation. Unlike cdh1Δ and mad2Δ single mutants, the mad2Δ cdh1Δ double mutant grew slowly and exhibited high rates of chromosome and plasmid loss. Simultaneous deletion of PDS1 and CDH1 caused extensive chromosome missegregation and cell death. Our data suggest that at least part of the chromosome loss can be attributed to kinetochore/spindle problems. Our data further suggest that Cdh1p and Sic1p, a Cdc28p/Clb inhibitor, have overlapping as well as nonoverlapping roles in ensuring proper chromosome segregation. The severe growth defects of both mad2Δ cdh1Δ and pds1Δ cdh1Δ strains were rescued by overexpressing Swe1p, a G2/M inhibitor of the cyclin-dependent kinase, Cdc28p/Clb. We propose that the failure to degrade cyclins at the end of mitosis leaves cdh1Δ mutant strains with abnormal Cdc28p/Clb activity that interferes with proper chromosome segregation.

scholarly journals Dual-mode regulation of the APC/C by CDK1 and MAPK controls meiosis I progression and fidelity

2014 ◽  
Vol 204 (6) ◽  
pp. 891-900 ◽  
Author(s):  
Ibtissem Nabti ◽  
Petros Marangos ◽  
Jenny Bormann ◽  
Nobuaki R. Kudo ◽  
John Carroll
Keyword(s):  
Protein Kinase ◽  
Spindle Assembly Checkpoint ◽  
Mitogen Activated Protein Kinase ◽  
Cyclin Dependent Kinase ◽  
Anaphase Promoting Complex ◽  
Female Meiosis ◽  
Dual Mode ◽  
Mitogen Activated Protein ◽  
Meiosis I

Female meiosis is driven by the activities of two major kinases, cyclin-dependent kinase 1 (Cdk1) and mitogen-activated protein kinase (MAPK). To date, the role of MAPK in control of meiosis is thought to be restricted to maintaining metaphase II arrest through stabilizing Cdk1 activity. In this paper, we find that MAPK and Cdk1 play compensatory roles to suppress the anaphase-promoting complex/cyclosome (APC/C) activity early in prometaphase, thereby allowing accumulation of APC/C substrates essential for meiosis I. Furthermore, inhibition of MAPK around the onset of APC/C activity at the transition from meiosis I to meiosis II led to accelerated completion of meiosis I and an increase in aneuploidy at metaphase II. These effects appear to be mediated via a Cdk1/MAPK-dependent stabilization of the spindle assembly checkpoint, which when inhibited leads to increased APC/C activity. These findings demonstrate new roles for MAPK in the regulation of meiosis in mammalian oocytes.

scholarly journals Uncoupling Anaphase-Promoting Complex/Cyclosome Activity from Spindle Assembly Checkpoint Control by Deregulating Polo-Like Kinase 1

2005 ◽  
Vol 25 (5) ◽  
pp. 2031-2044 ◽  
Author(s):  
Barbara C. M. van de Weerdt ◽  
Marcel A. T. M. van Vugt ◽  
Catherine Lindon ◽  
Jos J. W. Kauw ◽  
Marieke J. Rozendaal ◽  
...  
Keyword(s):  
Double Mutant ◽  
Spindle Assembly Checkpoint ◽  
Spindle Assembly ◽  
Mitotic Catastrophe ◽  
Anaphase Promoting Complex ◽  
Checkpoint Control ◽  
Mitotic Progression ◽  
Specific Antibodies ◽  
Mitotic Entry

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.

scholarly journals Spindle checkpoint proteins Mad1 and Mad2 are required for cytostatic factor–mediated metaphase arrest

2003 ◽  
Vol 163 (6) ◽  
pp. 1231-1242 ◽  
Author(s):  
Brian J. Tunquist ◽  
Patrick A. Eyers ◽  
Lin G. Chen ◽  
Andrea L. Lewellyn ◽  
James L. Maller
Keyword(s):  
Cell Cycle ◽  
Spindle Assembly Checkpoint ◽  
Spindle Checkpoint ◽  
Anaphase Promoting Complex ◽  
Metaphase Arrest ◽  
Checkpoint Proteins ◽  
Cycle Arrest ◽  
Egg Extracts ◽  
Cytostatic Factor ◽  
Sustained Inhibition

In cells containing disrupted spindles, the spindle assembly checkpoint arrests the cell cycle in metaphase. The budding uninhibited by benzimidazole (Bub) 1, mitotic arrest-deficient (Mad) 1, and Mad2 proteins promote this checkpoint through sustained inhibition of the anaphase-promoting complex/cyclosome. Vertebrate oocytes undergoing meiotic maturation arrest in metaphase of meiosis II due to a cytoplasmic activity termed cytostatic factor (CSF), which appears not to be regulated by spindle dynamics. Here, we show that microinjection of Mad1 or Mad2 protein into early Xenopus laevis embryos causes metaphase arrest like that caused by Mos. Microinjection of antibodies to either Mad1 or Mad2 into maturing oocytes blocks the establishment of CSF arrest in meiosis II, and immunodepletion of either protein blocked the establishment of CSF arrest by Mos in egg extracts. A Mad2 mutant unable to oligomerize (Mad2 R133A) did not cause cell cycle arrest in blastomeres or in egg extracts. Once CSF arrest has been established, maintenance of metaphase arrest requires Mad1, but not Mad2 or Bub1. These results suggest a model in which CSF arrest by Mos is mediated by the Mad1 and Mad2 proteins in a manner distinct from the spindle checkpoint.

scholarly journals The Zds proteins control entry into mitosis and target protein phosphatase 2A to the Cdc25 phosphatase

2011 ◽  
Vol 22 (1) ◽  
pp. 20-32 ◽  
Author(s):  
Sidonie Wicky ◽  
Hendri Tjandra ◽  
David Schieltz ◽  
John Yates ◽  
Douglas R. Kellogg
Keyword(s):  
Protein Phosphatase ◽  
Protein Phosphatase 2A ◽  
Regulatory Subunit ◽  
Cyclin Dependent Kinase ◽  
Cdc25 Phosphatase ◽  
Phosphatase 2A ◽  
Wee1 Kinase ◽  
Cycle Dependent ◽  
Related Proteins ◽  
Stoichiometric Complex

The Wee1 kinase restrains entry into mitosis by phosphorylating and inhibiting cyclin-dependent kinase 1 (Cdk1). The Cdc25 phosphatase promotes entry into mitosis by removing Cdk1 inhibitory phosphorylation. Experiments in diverse systems have established that Wee1 and Cdc25 are regulated by protein phosphatase 2A (PP2A), but a full understanding of the function and regulation of PP2A in entry into mitosis has remained elusive. In budding yeast, entry into mitosis is controlled by a specific form of PP2A that is associated with the Cdc55 regulatory subunit (PP2ACdc55). We show here that related proteins called Zds1 and Zds2 form a tight stoichiometric complex with PP2ACdc55and target its activity to Cdc25 but not to Wee1. Conditional inactivation of the Zds proteins revealed that their function is required primarily at entry into mitosis. In addition, Zds1 undergoes cell cycle–dependent changes in phosphorylation. Together, these observations define a role for the Zds proteins in controlling specific functions of PP2ACdc55and suggest that upstream signals that regulate PP2ACdc55may play an important role in controlling entry into mitosis.

scholarly journals Protein Phosphatase 2A Stabilizes Human Securin, Whose Phosphorylated Forms Are Degraded via the SCF Ubiquitin Ligase

2006 ◽  
Vol 26 (11) ◽  
pp. 4017-4027 ◽  
Author(s):  
Ana M. Gil-Bernabé ◽  
Francisco Romero ◽  
M. Cristina Limón-Mortés ◽  
María Tortolero
Keyword(s):  
Ubiquitin Ligase ◽  
Protein Phosphatase ◽  
Protein Phosphatase 2A ◽  
Anaphase Promoting Complex ◽  
Anaphase Onset ◽  
Chromatid Segregation ◽  
Phosphatase 2A ◽  
F Box Protein

ABSTRACT Sister chromatid segregation is triggered at the metaphase-to-anaphase transition by the activation of the protease separase. For most of the cell cycle, separase activity is kept in check by its association with the inhibitory chaperone securin. Activation of separase occurs at anaphase onset, when securin is targeted for destruction by the anaphase-promoting complex or cyclosome E3 ubiquitin protein ligase. This results in the release of the cohesins from chromosomes, which in turn allows the segregation of sister chromatids to opposite spindle poles. Here we show that human securin (hSecurin) forms a complex with enzymatically active protein phosphatase 2A (PP2A) and that it is a substrate of the phosphatase, both in vitro and in vivo. Treatment of cells with okadaic acid, a potent inhibitor of PP2A, results in various hyperphosphorylated forms of hSecurin which are extremely unstable, due to the action of the Skp1/Cul1/F-box protein complex ubiquitin ligase. We propose that PP2A regulates hSecurin levels by counteracting its phosphorylation, which promotes its degradation. Misregulation of this process may lead to the formation of tumors, in which overproduction of hSecurin is often observed.

scholarly journals Phosphatase 2A Negatively Regulates Mitotic Exit in Saccharomyces cerevisiae

2006 ◽  
Vol 17 (1) ◽  
pp. 80-89 ◽  
Author(s):  
Yanchang Wang ◽  
Tuen-Yung Ng
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mitotic Exit ◽  
Regulatory Subunit ◽  
Temperature Sensitive ◽  
Cyclin Dependent Kinase ◽  
Yeast Saccharomyces Cerevisiae ◽  
Catalytic Subunits ◽  
Phosphatase 2A ◽  
Negative Role

In budding yeast Saccharomyces cerevisiae, Cdc5 kinase is a component of mitotic exit network (MEN), which inactivates cyclin-dependent kinase (CDK) after chromosome segregation. cdc5-1 mutants arrest at telophase at the nonpermissive temperature due to the failure of CDK inactivation. To identify more negative regulators of MEN, we carried out a genetic screen for genes that are toxic to cdc5-1 mutants when overexpressed. Genes that encode the B-regulatory subunit (Cdc55) and the three catalytic subunits (Pph21, Pph22, and Pph3) of phosphatase 2A (PP2A) were isolated. In addition to cdc5-1, overexpression of CDC55, PPH21, or PPH22 is also toxic to other temperature-sensitive mutants that display defects in mitotic exit. Consistently, deletion of CDC55 partially suppresses the temperature sensitivity of these mutants. Moreover, in the presence of spindle damage, PP2A mutants display nuclear localized Cdc14, the key player in MEN pathway, indicative of MEN activation. All the evidence suggests the negative role of PP2A in mitotic exit. Finally, our genetic and biochemical data suggest that PP2A regulates the phosphorylation of Tem1, which acts at the very top of MEN pathway.

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