Down-regulation of estrogen receptor alpha (ER alpha) transcriptional activity by p27 is mediated by inhibition of ERa alpha nuclear localization and modulation of the ER alpha transcriptional complex

Abstract

To investigate the role of p27 on estrogen receptor (ER)a-mediated transcription, we generated MCF-7 cells with knocked down p27 via retroviral delivery of p27 shRNA. Suppression of p27 expression in MCF-7 cells resulted in up-regulation of ER alpha-mediated transcription by estradiol compared to the levels in control MCF-7 cells. Accordingly, transient transfection studies in 293T cells revealed that overexpression of p27 reduced ER alpha-mediated transcription. The effect of p27 on ER alpha transcriptional activity was independent of cell cycle arrest by p27, as cell cycle arrest induced by serum starvation did not significantly affect ER alpha-mediated transcription. Further, we observed that p27 inhibited nuclear localization of ER alpha, and that p27 was associated with ER alpha in the cytoplasm. We also investigated the role of p27 in the modulation of ER alpha transcriptional activity in the nucleus. We found that p27 negatively modulated ERa transcriptional activity by inhibiting association of cyclin D1 with ERa and recruiting BRCA1 to ER alpha transcriptional complex. Taken together, these data suggest that p27 inhibits ER alpha transcriptional activity by two independent mechanisms, namely, physical nuclear exclusion of ER alpha, and modulation of the ERa transcriptional complex.