FORSKOLIN-INDUCED DEPHOSPHORYLATION OF THE ANDROGEN RECEPTOR IMPAIRS LIGAND-BINDING

When androgen receptor containing cells are cultured in the presence o f the PKA stimulator forskolin, a rapid dephosphorylation of the andro gen receptor occurs resulting in a decrease in the amount of 112 kDa a ndrogen receptor isoform and an increase in 110 kDa androgen receptor isoform on SDS-PAGE. To establish which amino acid residues in the and rogen receptor were phosphorylated in control and forskolin-treated ce lls, trypsin-digested androgen receptors were subjected to RP-HPLC ana lysis and subsequently to Edman degradation. It was observed that seri ne residues 505, 641, and 653 were potentially phosphorylated in contr ol cells, while after forskolin treatment strong evidence was obtained that phosphorylation of serines 641 and 653 was significantly reduced . When the dephosphorylated androgen receptor was analyzed for its tra nscription activation capacity, it was observed that androgen-induced transcriptional regulation of two endogenous genes (PSA and beta 1-sub unit of Na,K-ATPase), in cells cultured in the presence of forskolin, was inhibited as compared to the control situation. The observation th at the dephosphorylated androgen receptor was transcriptionally less a ctive was further strengthened by the finding that the dephosphorylate d androgen receptor was markedly impaired in ligand binding (B-max was found to be reduced by approximately 40%). The current investigations show for the first time a clear function for the rapid phosphorylatio n which occurs directly after synthesis of the androgen receptor, name ly, effective ligand binding.