title: Role of phosphorylation of Cdc20 in p31(comet)-stimulated disassembly of the mitotic checkpoint complex.
abstract: The mitotic checkpoint system delays anaphase until all chromosomes are correctly attached to the mitotic spindle. When the checkpoint is turned on, it promotes the formation of the mitotic checkpoint complex (MCC), which inhibits the ubiquitin ligase anaphase-promoting complex/cyclosome (APC/C). MCC is composed of the checkpoint proteins BubR1, Bub3, and Mad2 bound to the APC/C activator Cdc20. When the checkpoint is satisfied, MCC is disassembled and APC/C becomes active. Previous studies have shown that the Mad2-binding protein p31(comet) promotes the dissociation of Cdc20 from BubR1 in MCC in a process that requires ATP. We now show that a part of MCC dissociation is blocked by inhibitors of cyclin-dependent kinases (Cdks) and that purified Cdk1-cyclin B stimulates this process. The mutation of all eight potential Cdk phosphorylation sites of Cdc20 partially prevented its release from BubR1. Furthermore, p31(comet) stimulated Cdk-catalyzed phosphorylation of Cdc20 in MCC. It is suggested that the binding of p31(comet) to Mad2 in MCC may trigger a conformational change in Cdc20 that facilitates its phosphorylation by Cdk, and that the latter process may promote its dissociation from BubR1.
authors.affiliation: Unit of Biochemistry, Rappaport Faculty of Medicine,  Technion-Israel Institute of Technology, Haifa 31096,  Israel.
authors.forename: Shirly
authors.initials: S
authors.lastname: Miniowitz-Shemtov
authors.forename: Esther
authors.initials: E
authors.lastname: Eytan
authors.forename: Dvora
authors.initials: D
authors.lastname: Ganoth
authors.forename: Danielle
authors.initials: D
authors.lastname: Sitry-Shevah
authors.forename: Elena
authors.initials: E
authors.lastname: Dumin
authors.forename: Avram
authors.initials: A
authors.lastname: Hershko
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism; D064173:Anaphase-Promoting Complex-Cyclosome; D016203:CDC2 Protein Kinase / Q000378:metabolism; D002135:Calcium-Binding Proteins / Q000378:metabolism; D064199:Cdc20 Proteins; D018797:Cell Cycle Proteins / Q000378:metabolism*; D056744:Cyclin B1 / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D059566:M Phase Cell Cycle Checkpoints / Q000502:physiology*; D064246:Mad2 Proteins; D008938:Mitosis / Q000502:physiology; D046912:Multiprotein Complexes / Q000378:metabolism; D009687:Nuclear Proteins / Q000378:metabolism*; D010766:Phosphorylation / Q000502:physiology; D000075223:Poly-ADP-Ribose Binding Proteins; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D012097:Repressor Proteins / Q000378:metabolism; D008941:Spindle Apparatus / Q000378:metabolism*; D043743:Ubiquitin-Protein Ligase Complexes / Q000378:metabolism*
chemical_list: D048868:Adaptor Proteins, Signal Transducing; C489444:BUB3 protein, human; C534699:CCNB1 protein, human; D002135:Calcium-Binding Proteins; D064199:Cdc20 Proteins; D018797:Cell Cycle Proteins; D056744:Cyclin B1; C577697:MAD2L1 protein, human; C470309:MAD2L1BP protein, human; D064246:Mad2 Proteins; D046912:Multiprotein Complexes; D009687:Nuclear Proteins; D000075223:Poly-ADP-Ribose Binding Proteins; D012097:Repressor Proteins; C577488:CDC20 protein, human; D000255:Adenosine Triphosphate; D043743:Ubiquitin-Protein Ligase Complexes; D064173:Anaphase-Promoting Complex-Cyclosome; C584277:BUB1 protein, human; C090464:Bub1 spindle checkpoint protein; D017346:Protein Serine-Threonine Kinases; D016203:CDC2 Protein Kinase