LOCALIZATION AND HORMONAL-STIMULATION OF PHOSPHORYLATION SITES IN THE LNCAP-CELL ANDROGEN RECEPTOR

Phosphorylation of the androgen receptor in human prostate tumour cells (LNCaP) is increased by addition of androgens to intact cells. Double-label studies, using [S-35]methionine incorporation into receptor protein, and [P-32]P(i) to label metabolically receptor phosphorylation sites, have enabled us to determine the phosphate content, relative to receptor protein, of both non-transformed and transformed androgen receptors generated in intact LNCaP cells. No net change in the phosphorylation of the intact 110 kDa steroid-binding component of the androgen-receptor complex was found upon transformation to the tight nuclear binding form in the intact cell. Partial proteolysis of androgen receptor protein metabolically labelled with [P-32]P(i) and photolabelled with [H-3]R1881 (methyltrienolone) revealed that phosphorylation occurs mainly in the N-terminal trans-activation domain, whereas no phosphorylation was detected in the steroid- and DNA-binding domains. The location of most (> 90 %) of the hormonally regulated phosphorylation sites in the N-terminal trans-activation domain suggests a role of phosphorylation of the androgen receptor in transcription regulation.