Stable MCC binding to the APC/C is required for a functional spindle assembly checkpoint.
The spindle assembly checkpoint (SAC) delays progression into anaphase until all chromosomes have aligned on the metaphase plate by inhibiting Cdc20, the mitotic co-activator of the APC/C. Mad2 and BubR1 bind and inhibit Cdc20, thereby forming the mitotic checkpoint complex (MCC), which can bind stably to the APC/C. Whether MCC formation per se is sufficient for a functional SAC or MCC association with the APC/C is required remains unclear. Here, we analyze the role of two conserved motifs in Cdc20, IR and C-Box, in binding of the MCC to the APC/C. Mutants in both motifs assemble the MCC normally, but IR motif integrity is particularly important for stable binding to the APC/C. Cells expressing Cdc20 with a mutated IR motif have a compromised SAC, as uninhibited Cdc20 can compete with the MCC for APC/C binding and activate it. We thus show that stable MCC association with the APC/C is critical for a functional SAC.
The Novo Nordisk Foundation Center for Protein Research, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark.
Jamin B
JB
Hein
Jakob
J
Nilsson
D020816:Amino Acid Motifs; D064173:Anaphase-Promoting Complex-Cyclosome / Q000378:metabolism*; D001665:Binding Sites; D064199:Cdc20 Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017124:Conserved Sequence; D006367:HeLa Cells; D006801:Humans; D059566:M Phase Cell Cycle Checkpoints; D064246:Mad2 Proteins / Q000378:metabolism*; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D008941:Spindle Apparatus / Q000378:metabolism*
D064199:Cdc20 Proteins; C577697:MAD2L1 protein, human; D064246:Mad2 Proteins; C577488:CDC20 protein, human; D064173:Anaphase-Promoting Complex-Cyclosome; C584277:BUB1 protein, human; D017346:Protein Serine-Threonine Kinases