Phosphorylation of HSP90 by protein kinase A is essential for the nuclear translocation of androgen receptor

The androgen receptor (AR) is often activated in prostate cancer patients undergoing androgen-ablative therapy because of the activation of cellular pathways that stimulate the AR despite low androgen levels. In many of these tumors, the cAMP-dependent protein kinase A (PKA) pathway is activated. Previous studies have shown that PKA can synergize with low levels of androgen to enhance androgen signaling and consequent cell proliferation, leading to castration-resistant prostate cancer. However, the mechanism by which PKA causes AR stimulation in the presence of low/no androgen is not established yet. Here, using immunofluorescence immunoblotting assays, co-immunoprecipitation, siRNA-mediated gene silencing, and reporter gene assays, we demonstrate that PKA activation is necessary for the phosphorylation of heat shock protein (HSP90) that binds to unliganded AR in the cytoplasm, restricting its entry into the nucleus. We also found that PKA-mediated phosphorylation of the Thr(89) residue in HSP90 releases AR from HSP90, enabling AR binding to HSP27 and its migration into the nucleus. Substitution of the Thr(89) in HSP90 prevented its phosphorylation by PKA and significantly reduced AR transactivation and cellular proliferation. We further observed that the transcription of AR target genes, such as prostate-specific antigen (PSA), is also lowered in the HSP90 Thr(89) variant. These results suggest that using a small-molecule inhibitor against the HSP90 Thr(89) residue in conjunction with existing androgen-ablative therapy may be more effective than androgen-ablative therapy alone in the treatment of prostate cancer patients.