Cell-cycle-dependent regulation of androgen receptor function.

Abstract

The androgen receptor (AR) is a critical oncogene in prostate cancer (PCa) development and progression. In this study, we demonstrate cell-cycle-dependent regulation of AR activity, localization, and phosphorylation. We show that for three AR-target genes, androgen-stimulated AR transactivation is highest during the G1 phase, decreased during S-phase, and abrogated during G2/M. This change in AR transactivation parallels changes in AR localization and phosphorylation. A combination of imaging techniques and quantitative analysis reveals nuclear AR localization during interphase and the exclusion of the majority, but not all, AR from chromatin during mitosis. Flow cytometry analyses using a phospho-S308 AR-specific antibody in asynchronous and chemically enriched G2/M PCa cells revealed ligand-independent induction of S308 phosphorylation in mitosis when CDK1 is activated. Consistent with our flow cytometry data, IP-western blotting revealed an increase in S308 phosphorylation in G2/M, and the results of an in vitro kinase assay indicated that CDK1 was able to phosphorylate the AR on S308. Pharmacological inhibition of CDK1 activity resulted in decreased S308 phosphorylation in PCa cells. Importantly, using a combination of anti-total AR and phospho-S308-specific antibodies in immunofluorescence experiments, we showed that the AR is excluded from condensed chromatin in mitotic cells when it was phosphorylated on S308. In summary, we show that the phosphorylation of the AR on S308 by CDK1 during mitosis regulates AR localization and correlates with changes in AR transcriptional activity. These findings have important implications for understanding the function of AR as an oncogene.