The Bub1-Plk1 kinase complex promotes spindle checkpoint signalling through Cdc20 phosphorylation

Nat Commun. 2016 Feb 25;7:10818. doi: 10.1038/ncomms10818.

Abstract

The spindle checkpoint senses unattached kinetochores and inhibits the Cdc20-bound anaphase-promoting complex or cyclosome (APC/C), to delay anaphase, thereby preventing aneuploidy. A critical checkpoint inhibitor of APC/C(Cdc20) is the mitotic checkpoint complex (MCC). It is unclear whether MCC suffices to inhibit all cellular APC/C. Here we show that human checkpoint kinase Bub1 not only directly phosphorylates Cdc20, but also scaffolds Plk1-mediated phosphorylation of Cdc20. Phosphorylation of Cdc20 by Bub1-Plk1 inhibits APC/C(Cdc20) in vitro and is required for checkpoint signalling in human cells. Bub1-Plk1-dependent Cdc20 phosphorylation is regulated by upstream checkpoint signals and is dispensable for MCC assembly. A phospho-mimicking Cdc20 mutant restores nocodazole-induced mitotic arrest in cells depleted of Mad2 or BubR1. Thus, Bub1-Plk1-mediated phosphorylation of Cdc20 constitutes an APC/C-inhibitory mechanism that is parallel, but not redundant, to MCC formation. Both mechanisms are required to sustain mitotic arrest in response to spindle defects.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Anaphase-Promoting Complex-Cyclosome / metabolism*
  • Cdc20 Proteins / genetics
  • Cdc20 Proteins / metabolism*
  • Cell Cycle Proteins / genetics*
  • Cell Line, Tumor
  • Flow Cytometry
  • Fluorescent Antibody Technique
  • HeLa Cells
  • Humans
  • Immunoblotting
  • Immunoprecipitation
  • In Vitro Techniques
  • Kinetochores / metabolism
  • M Phase Cell Cycle Checkpoints / drug effects
  • M Phase Cell Cycle Checkpoints / genetics*
  • Mad2 Proteins
  • Mutation
  • Nocodazole / pharmacology
  • Phosphorylation
  • Protein-Serine-Threonine Kinases / genetics*
  • Proto-Oncogene Proteins / genetics*
  • Signal Transduction
  • Tubulin Modulators / pharmacology
  • Ubiquitination

Substances

  • Cdc20 Proteins
  • Cell Cycle Proteins
  • MAD2L1 protein, human
  • Mad2 Proteins
  • Proto-Oncogene Proteins
  • Tubulin Modulators
  • CDC20 protein, human
  • Anaphase-Promoting Complex-Cyclosome
  • BUB1 protein, human
  • Protein-Serine-Threonine Kinases
  • polo-like kinase 1
  • Nocodazole