title: Role of cyclin B1/Cdc2 up-regulation in the development of mitotic prometaphase arrest in human breast cancer cells treated with nocodazole.
abstract: 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.
  
  
  RESULTS
  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
  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.
authors.affiliation: Department of Pharmacology, Toxicology and Therapeutics, School of Medicine, University of Kansas Medical Center, Kansas City, Kansas, United States of America.
authors.forename: Hye Joung
authors.initials: HJ
authors.lastname: Choi
authors.forename: Masayuki
authors.initials: M
authors.lastname: Fukui
authors.forename: Bao Ting
authors.initials: BT
authors.lastname: Zhu
mesh_terms: D000970:Antineoplastic Agents / Q000494:pharmacology*; D015153:Blotting, Western; D001943:Breast Neoplasms / Q000378:metabolism* / Q000473:pathology; D016203:CDC2 Protein Kinase; D045744:Cell Line, Tumor; D019926:Cyclin B / Q000378:metabolism*; D056744:Cyclin B1 / Q000378:metabolism*; D018844:Cyclin-Dependent Kinases; D005260:Female; D005434:Flow Cytometry; D006801:Humans; D008856:Microscopy, Fluorescence; D008938:Mitosis / Q000187:drug effects*; D015739:Nocodazole / Q000494:pharmacology*; D049468:Prometaphase / Q000187:drug effects*; D034741:RNA, Small Interfering
chemical_list: D000970:Antineoplastic Agents; D019926:Cyclin B; D056744:Cyclin B1; D034741:RNA, Small Interfering; D016203:CDC2 Protein Kinase; C528899:CDK1 protein, human; D018844:Cyclin-Dependent Kinases; D015739:Nocodazole