Although RE inhibits the G(1)-S transition, the mechanism through which RE prevents cell cycle advancement remains unidentified. To delineate the mechanism(s) utilized by RE to exert its anti-proliferative activity, constitutively active RE proteins (which cannot be inactivated by phosphorylation) or p16ink4a (which prevents RE inactivation) were utilized. Both proteins inhibited the G(1)-S transition, whereas wildtype RE did not. We show that active RE acts to attenuate cyclin A promoter activity, and that overexpression of cyclin E reverses RE-mediated repression of the cyclin A promoter. Although cyclin A is an E2F-regulated gene, and it has been long hypothesized that RE 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 RE 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 RE 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 RE-mediated G(1) arrest. The findings presented suggest a pathway wherein cyclin A is a downstream target of RE, and cyclin E functions to antagonize this aspect of RE-mediated G(1)-S inhibition.
Cyclin A is a functional target of retinoblastoma tumor suppressor protein-mediated cell cycle arrest
Knudsen, K. E.; Fribourg, A. F.; Strobeck, M. W.; Blanchard, J. M.; Knudsen, E. S.
Journal of Biological Chemistry
1999-09-24 / vol 274 / pages 27632-27641
gene-expression; transcriptional repression; s-phase; phosphorylation; e2f; carcinoma-cells; dependent kinases; ectopic expression; family proteins; growth suppression