Role of cyclin B1/Cdc2 up-regulation in the development of mitotic prometaphase arrest in human breast cancer cells treated with nocodazole

PLoS One. 2011;6(8):e24312. doi: 10.1371/journal.pone.0024312. Epub 2011 Aug 30.


Background: During a normal cell cycle, the transition from G₂ phase to mitotic phase is triggered by the activation of the cyclin B1-dependent Cdc2 kinase. Here we report our finding that treatment of MCF-7 human breast cancer cells with nocodazole, a prototypic microtubule inhibitor, results in strong up-regulation of cyclin B1 and Cdc2 levels, and their increases are required for the development of mitotic prometaphase arrest and characteristic phenotypes.

Methodology/principal findings: It was observed that there was a time-dependent early increase in cyclin B1 and Cdc2 protein levels (peaking between 12 and 24 h post treatment), and their levels started to decline after the initial increase. This early up-regulation of cyclin B1 and Cdc2 closely matched in timing the nocodazole-induced mitotic prometaphase arrest. Selective knockdown of cyclin B1or Cdc2 each abrogated nocodazole-induced accumulation of prometaphase cells. The nocodazole-induced prometaphase arrest was also abrogated by pre-treatment of cells with roscovitine, an inhibitor of cyclin-dependent kinases, or with cycloheximide, a protein synthesis inhibitor that was found to suppress cyclin B1 and Cdc2 up-regulation. In addition, we found that MAD2 knockdown abrogated nocodazole-induced accumulation of cyclin B1 and Cdc2 proteins, which was accompanied by an attenuation of nocodazole-induced prometaphase arrest.

Conclusions/significance: These observations demonstrate that the strong early up-regulation of cyclin B1 and Cdc2 contributes critically to the rapid and selective accumulation of prometaphase-arrested cells, a phenomenon associated with exposure to microtubule inhibitors.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Antineoplastic Agents / pharmacology*
  • Blotting, Western
  • Breast Neoplasms / metabolism*
  • Breast Neoplasms / pathology
  • CDC2 Protein Kinase
  • Cell Line, Tumor
  • Cyclin B / metabolism*
  • Cyclin B1 / metabolism*
  • Cyclin-Dependent Kinases
  • Female
  • Flow Cytometry
  • Humans
  • Microscopy, Fluorescence
  • Mitosis / drug effects*
  • Nocodazole / pharmacology*
  • Prometaphase / drug effects*
  • RNA, Small Interfering


  • Antineoplastic Agents
  • Cyclin B
  • Cyclin B1
  • RNA, Small Interfering
  • CDC2 Protein Kinase
  • CDK1 protein, human
  • Cyclin-Dependent Kinases
  • Nocodazole