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APC/C(Cdc20) controls the ubiquitin-mediated degradation of p21 in prometaphase

Virginia Amador1, Sheng Ge, Patricia G Santamaría

  • 1Department of Pathology, NYU Cancer Institute, New York University School of Medicine, 550 First Avenue MSB 599, New York, NY 10016, USA.

Molecular Cell
|August 7, 2007
PubMed

Insights

The study reveals that Cdc20 targets p21 for degradation during prometaphase, enabling cyclin-dependent kinase 1 (Cdk1) activation. This process is crucial for cell cycle progression and preventing errors during mitosis.

Area of Science:

  • Cell Biology
  • Molecular Biology
  • Biochemistry

Background:

  • p21 proteolysis is regulated during the cell cycle, with Skp2 mediating degradation during G1/S.
  • The precise mechanisms controlling p21 degradation during mitosis, particularly in prometaphase, were previously unknown.

Purpose of the Study:

  • To investigate the role of Cdc20 in p21 degradation during mitosis.
  • To elucidate the molecular mechanisms by which p21 degradation is controlled in prometaphase.
  • To understand the functional consequences of p21 degradation for Cdk1 activity and mitotic progression.

Main Methods:

  • Co-immunoprecipitation to detect protein-protein interactions between Cdc20 and p21.
  • Site-directed mutagenesis to identify the D box motif in p21.
  • Western blotting to assess protein levels and ubiquitylation.
  • RNA interference (siRNA) to silence Cdc20 expression.
  • Cell-based assays using wild-type and p21 knockout cells to evaluate Cdk1 activity and cell cycle progression.

Main Results:

  • Cdc20 binds to p21 in cultured cells, and a D box motif in p21 is essential for APC/C(Cdc20)-mediated ubiquitylation and degradation.
  • Overexpression of Cdc20 or Skp2 destabilizes wild-type p21, but only Skp2 destabilizes a p21 mutant lacking the D box.
  • Silencing Cdc20 leads to p21 accumulation, increased p21 binding to Cdk1, and inhibition of Cdk1 activity in prometaphase cells.
  • These effects are specific to cells with functional p21.

Conclusions:

  • Cdc20 acts as a positive regulator of Cdk1 activity in prometaphase by mediating p21 degradation.
  • APC/C(Cdc20)-mediated degradation of p21 is critical for the full activation of Cdk1, which is necessary for mitotic progression.
  • This degradation pathway helps prevent mitotic slippage, especially when the spindle assembly checkpoint is active.

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