Abstract
In mitosis, the spindle assembly checkpoint (SAC) ensures genome stability by delaying chromosome segregation until all sister chromatids have achieved bipolar attachment to the mitotic spindle. The SAC is imposed by the mitotic checkpoint complex (MCC), whose assembly is catalysed by unattached chromosomes and which binds and inhibits the anaphase-promoting complex/cyclosome (APC/C), the E3 ubiquitin ligase that initiates chromosome segregation. Here, using the crystal structure of Schizosaccharomyces pombe MCC (a complex of mitotic spindle assembly checkpoint proteins Mad2, Mad3 and APC/C co-activator protein Cdc20), we reveal the molecular basis of MCC-mediated APC/C inhibition and the regulation of MCC assembly. The MCC inhibits the APC/C by obstructing degron recognition sites on Cdc20 (the substrate recruitment subunit of the APC/C) and displacing Cdc20 to disrupt formation of a bipartite D-box receptor with the APC/C subunit Apc10. Mad2, in the closed conformation (C-Mad2), stabilizes the complex by optimally positioning the Mad3 KEN-box degron to bind Cdc20. Mad3 and p31(comet) (also known as MAD2L1-binding protein) compete for the same C-Mad2 interface, which explains how p31(comet) disrupts MCC assembly to antagonize the SAC. This study shows how APC/C inhibition is coupled to degron recognition by co-activators.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Amino Acid Motifs
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Anaphase-Promoting Complex-Cyclosome
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Cdc20 Proteins
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Cdh1 Proteins
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Cell Cycle Proteins / chemistry*
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Cell Cycle Proteins / metabolism
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Conserved Sequence
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Crystallography, X-Ray
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Humans
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M Phase Cell Cycle Checkpoints*
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Mad2 Proteins
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Models, Molecular
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Multiprotein Complexes / chemistry*
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Multiprotein Complexes / metabolism
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Nuclear Proteins / chemistry*
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Nuclear Proteins / metabolism
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Protein Structure, Quaternary
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Protein Structure, Tertiary
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Saccharomyces cerevisiae Proteins / chemistry
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Saccharomyces cerevisiae Proteins / genetics
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Saccharomyces cerevisiae Proteins / metabolism
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Schizosaccharomyces / chemistry*
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Schizosaccharomyces pombe Proteins / chemistry*
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Schizosaccharomyces pombe Proteins / metabolism
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Spindle Apparatus
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Structure-Activity Relationship
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Substrate Specificity
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Ubiquitin-Protein Ligase Complexes / antagonists & inhibitors
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Ubiquitin-Protein Ligase Complexes / chemistry
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Ubiquitin-Protein Ligase Complexes / metabolism
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Ubiquitin-Protein Ligase Complexes / ultrastructure
Substances
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Cdc20 Proteins
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Cdh1 Proteins
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Cell Cycle Proteins
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Mad2 Proteins
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Multiprotein Complexes
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Nuclear Proteins
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Saccharomyces cerevisiae Proteins
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Schizosaccharomyces pombe Proteins
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mad2 protein, S pombe
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mad3 protein, S pombe
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Ubiquitin-Protein Ligase Complexes
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Anaphase-Promoting Complex-Cyclosome