title: Mitotic phosphatase activity is required for MCC maintenance during the spindle checkpoint.
abstract: The spindle checkpoint prevents activation of the anaphase-promoting complex (APC/C) until all chromosomes are correctly attached to the mitotic spindle. Early in mitosis, the mitotic checkpoint complex (MCC) inactivates the APC/C by binding the APC/C activating protein CDC20 until the chromosomes are properly aligned and attached to the mitotic spindle, at which point MCC disassembly releases CDC20 to activate the APC/C. Once the APC/C is activated, it targets cyclin B and securin for degradation, and the cell progresses into anaphase. While phosphorylation is known to drive many of the events during the checkpoint, the precise molecular mechanisms regulating spindle checkpoint maintenance and inactivation are still poorly understood. We sought to determine the role of mitotic phosphatases during the spindle checkpoint. To address this question, we treated spindle checkpoint-arrested cells with various phosphatase inhibitors and examined the effect on the MCC and APC/C activation. Using this approach we found that 2 phosphatase inhibitors, calyculin A and okadaic acid (1 μM), caused MCC dissociation and APC/C activation leading to cyclin A and B degradation in spindle checkpoint-arrested cells. Although the cells were able to degrade cyclin B, they did not exit mitosis as evidenced by high levels of Cdk1 substrate phosphorylation and chromosome condensation. Our results provide the first evidence that phosphatases are essential for maintenance of the MCC during operation of the spindle checkpoint.
authors.affiliation: a Department of Pharmacology and Cancer Biology , Duke University Medical Center , Durham , NC , USA.
authors.forename: Kristen M
authors.initials: KM
authors.lastname: Foss
authors.affiliation: a Department of Pharmacology and Cancer Biology , Duke University Medical Center , Durham , NC , USA.
authors.forename: Alexander C
authors.initials: AC
authors.lastname: Robeson
authors.affiliation: a Department of Pharmacology and Cancer Biology , Duke University Medical Center , Durham , NC , USA.
authors.forename: Sally
authors.initials: S
authors.lastname: Kornbluth
authors.affiliation: a Department of Pharmacology and Cancer Biology , Duke University Medical Center , Durham , NC , USA.|b Division of Medical Oncology , Department of Medicine, Duke University Medical Center , Durham , NC , USA.
authors.forename: Liguo
authors.initials: L
authors.lastname: Zhang
mesh_terms: D000705:Anaphase / Q000187:drug effects*; D064173:Anaphase-Promoting Complex-Cyclosome / Q000235:genetics / Q000378:metabolism; D016203:CDC2 Protein Kinase; D064199:Cdc20 Proteins / Q000235:genetics / Q000378:metabolism; D002875:Chromosomes / Q000737:chemistry / Q000187:drug effects*; D019925:Cyclin A / Q000235:genetics / Q000378:metabolism; D019926:Cyclin B / Q000235:genetics / Q000378:metabolism; D018844:Cyclin-Dependent Kinases / Q000235:genetics / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005786:Gene Expression Regulation; D006367:HeLa Cells; D006801:Humans; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D059566:M Phase Cell Cycle Checkpoints / Q000187:drug effects*; D008387:Marine Toxins; D019319:Okadaic Acid / Q000494:pharmacology; D010080:Oxazoles / Q000494:pharmacology; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D046988:Proteasome Endopeptidase Complex / Q000187:drug effects / Q000378:metabolism; D059748:Proteolysis / Q000187:drug effects; D064249:Securin / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D008941:Spindle Apparatus / Q000737:chemistry / Q000187:drug effects*
chemical_list: D064199:Cdc20 Proteins; D019925:Cyclin A; D019926:Cyclin B; D004791:Enzyme Inhibitors; D007527:Isoenzymes; D008387:Marine Toxins; D010080:Oxazoles; D064249:Securin; C578002:pituitary tumor-transforming protein 1, human; C577488:CDC20 protein, human; D019319:Okadaic Acid; C059041:calyculin A; D064173:Anaphase-Promoting Complex-Cyclosome; D016203:CDC2 Protein Kinase; C528899:CDK1 protein, human; D018844:Cyclin-Dependent Kinases; D010749:Phosphoprotein Phosphatases; D046988:Proteasome Endopeptidase Complex
keywords: APC/C; mitosis; mitotic checkpoint complex; phosphatases; spindle checkpoint