Cyclin A is a functional target of retinoblastoma tumor suppressor protein-mediated cell cycle arrest

J Biol Chem. 1999 Sep 24;274(39):27632-41. doi: 10.1074/jbc.274.39.27632.

Abstract

Although RB inhibits the G(1)-S transition, the mechanism through which RB prevents cell cycle advancement remains unidentified. To delineate the mechanism(s) utilized by RB to exert its anti-proliferative activity, constitutively active RB proteins (which cannot be inactivated by phosphorylation) or p16ink4a (which prevents RB inactivation) were utilized. Both proteins inhibited the G(1)-S transition, whereas wild-type RB did not. We show that active RB acts to attenuate cyclin A promoter activity, and that overexpression of cyclin E reverses RB-mediated repression of the cyclin A promoter. Although cyclin A is an E2F-regulated gene, and it has been long hypothesized that RB mediates cell cycle advancement through binding to E2F and attenuating its transactivation potential, cyclin E does not reverse dominant negative E2F-mediated repression of the cyclin A promoter. Although active RB repressed both cyclin A and two other paradigm E2F-regulated promoters, only cyclin A transcription was restored upon co-expression of cyclin E. Additionally, we show that RB but not dominant negative E2F regulates the cyclin A promoter through the CCRE element. These data identify cyclin A as a downstream target of RB-mediated arrest. Consistent with this idea, ectopic expression of cyclin A reversed RB-mediated G(1) arrest. The findings presented suggest a pathway wherein cyclin A is a downstream target of RB, and cyclin E functions to antagonize this aspect of RB-mediated G(1)-S inhibition.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Carrier Proteins*
  • Cell Cycle / physiology*
  • Cell Cycle Proteins*
  • Cell Division
  • Cell Line
  • Cyclin A / genetics*
  • Cyclin A / physiology
  • Cyclin E / genetics
  • Cyclin E / physiology
  • Cyclin-Dependent Kinase Inhibitor p16 / metabolism
  • DNA-Binding Proteins*
  • E2F Transcription Factors
  • G1 Phase
  • Gene Expression Regulation
  • Genes, Reporter
  • Luciferases / genetics
  • Promoter Regions, Genetic
  • Recombinant Fusion Proteins / biosynthesis
  • Retinoblastoma Protein / physiology*
  • Retinoblastoma-Binding Protein 1
  • S Phase
  • Tetrahydrofolate Dehydrogenase / genetics
  • Transcription Factors / metabolism
  • Transfection

Substances

  • Carrier Proteins
  • Cell Cycle Proteins
  • Cyclin A
  • Cyclin E
  • Cyclin-Dependent Kinase Inhibitor p16
  • DNA-Binding Proteins
  • E2F Transcription Factors
  • Recombinant Fusion Proteins
  • Retinoblastoma Protein
  • Retinoblastoma-Binding Protein 1
  • Transcription Factors
  • Luciferases
  • Tetrahydrofolate Dehydrogenase