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Structure of the mitotic checkpoint complex

Nature volume 484, pages 208–213 (2012)Cite this article

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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 p31comet (also known as MAD2L1-binding protein) compete for the same C-Mad2 interface, which explains how p31comet disrupts MCC assembly to antagonize the SAC. This study shows how APC/C inhibition is coupled to degron recognition by co-activators.

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Figure 1: Structure of S. pombe MCC trimer.
Figure 2: Details of Cdc20–Mad2–Mad3 interactions.
Figure 3: The KEN box binds to a conserved surface at the centre of the top side of the WD40 domain.
Figure 4: The D box binds in an extended conformation to a conserved inter-blade channel on the Cdc20 WD40 domain.
Figure 5: Pseudo-atomic structure of human APC/CMCC.

Accession codes

Primary accessions

Protein Data Bank

Data deposits

The MCC coordinates have been deposited in the Protein Data Bank under accession number 4aez.

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Acknowledgements

This work was funded by a Cancer Research UK grant to D.B. and an ICR studentship to W.C.H.C. We thank J. Yang and J. He for advice, and staff at the Diamond Light Source beamline I02 for help with data collection.

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Author notes

  1. William C. H. Chao and Kiran Kulkarni: These authors contributed equally to this work.

Authors and Affiliations

  1. Division of Structural Biology, Institute of Cancer Research, Chester Beatty Laboratories, 237 Fulham Road, London, SW3 6JB, UK,

    William C. H. Chao, Kiran Kulkarni, Ziguo Zhang, Eric H. Kong & David Barford

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Contributions

W.C.H.C. cloned, purified and crystallized the S. pombe MCC, and performed the biochemistry and mutagenesis experiments. W.C.H.C., K.K. and D.B. collected the X-ray diffraction data. K.K. determined the complex structure and modelled the D-box and KEN–D peptides. Z.Z. advised on cloning strategies of the MCC. E.H.K. provided purified endogenous APC/C. W.C.H.C., K.H.K. and D.B. docked crystal coordinates into the human APC/CMCC electron-microscope map. W.C.H.C. and D.B. wrote the manuscript and the others authors provided editorial input.

Corresponding author

Correspondence to David Barford.

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The authors declare no competing financial interests.

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Chao, W., Kulkarni, K., Zhang, Z. et al. Structure of the mitotic checkpoint complex. Nature 484, 208–213 (2012). https://doi.org/10.1038/nature10896

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