Cdk1/cyclin B plays a key role in mitotic arrest-induced apoptosis by phosphorylation of Mcl-1, promoting its degradation and freeing Bak from sequestration.
Mcl-1 is one of the major anti-apoptotic members of the Bcl-2 family of apoptotic regulatory proteins. In this study we investigated the role of Mcl-1 in mitotic arrest-induced apoptosis. Vinblastine treatment of KB-3 cells initially resulted in a phosphatase-sensitive mobility shift in Mcl-1 and then subsequent loss of Mcl-1 protein expression which was prevented by MG132, suggesting that phosphorylation triggered proteosome-mediated degradation. Mcl-1 phosphorylation/degradation was a specific response to microtubule inhibition and did not occur in response to lethal concentrations of DNA damaging agents. Vinblastine treatment caused degradation of Mcl-1 in cells in which apoptosis was blocked by Bcl-xL overexpression, indicating that Mcl-1 degradation was not a consequence of apoptosis. A partial reversible phosphorylation of Mcl-1 was observed in synchronized cells traversing mitosis, whereas more extensive phosphorylation and subsequent degradation of Mcl-1 was observed if synchronized cells were treated with vinblastine. Mcl-1 phosphorylation closely paralleled cyclin B expression, and specific cyclin-dependent kinase (Cdk) inhibitors blocked vinblastine-induced Mcl-1 phosphorylation, its subsequent degradation, and improved cell viability after mitotic arrest. Co-immunoprecipitation studies indicated that Mcl-1 was complexed with Bak, but not Bax or Noxa, in untreated cells, and that Bak became activated in concert with loss of Mcl-1 expression. These results suggest that Cdk1/cyclin B plays a key role in mitotic arrest-induced apoptosis via Mcl-1 phosphorylation, promoting its degradation and subsequently releasing Bak from sequestration.
Department of Biochemistry and Molecular Biology, University of Arkansas for Medical Sciences, Little Rock, AR 72205, USA.
Rong
R
Chu
David T
DT
Terrano
Timothy C
TC
Chambers
D017209:Apoptosis / Q000502:physiology*; D016203:CDC2 Protein Kinase / Q000502:physiology*; D059447:Cell Cycle Checkpoints / Q000502:physiology; D045744:Cell Line, Tumor; D019926:Cyclin B / Q000502:physiology*; D006801:Humans; D008938:Mitosis / Q000502:physiology*; D064549:Myeloid Cell Leukemia Sequence 1 Protein; D010766:Phosphorylation / Q000502:physiology; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism*; D050998:bcl-2 Homologous Antagonist-Killer Protein / Q000378:metabolism*; D051028:bcl-2-Associated X Protein / Q000378:metabolism
C494772:BAK1 protein, human; C494826:BAX protein, human; D019926:Cyclin B; D064549:Myeloid Cell Leukemia Sequence 1 Protein; C498702:PMAIP1 protein, human; D019253:Proto-Oncogene Proteins c-bcl-2; D050998:bcl-2 Homologous Antagonist-Killer Protein; D051028:bcl-2-Associated X Protein; D016203:CDC2 Protein Kinase