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J. Biol. Chem., Vol. 266, Issue 6, 3734-3738, 02, 1991

Hormone-dependent androgen receptor phosphorylation is accompanied by receptor transformation in human lymph node carcinoma of the prostate cells

JH van Laar, CA Berrevoets, J Trapman, ND Zegers and AO Brinkmann
Department of Biochemistry II, Medical Faculty, Erasmus University Rotterdam, The Netherlands.

Phosphorylation of the androgen receptor was investigated in the absence of hormone as well as during and after transformation of the receptor to the tight nuclear binding form. Human prostate tumor cells (LNCaP) were labeled for 4 h with [32P]orthophosphate in the presence or absence of steroid. Subsequently, androgen receptors were immunoprecipitated either from total cell lysates or from nuclear extracts using a specific monoclonal antibody. The immunoprecipitated receptor preparations were analyzed by sodium dodecyl sulfate- polyacrylamide gel electrophoresis, Western blotting, using a polyclonal antiserum, and autoradiography. It was observed that the androgen receptor is already phosphorylated in the absence of hormone, but undergoes a hormone-induced additional phosphorylation. After administration of 10 nM R1881, a 1.8-fold increase in phosphorylation over nonstimulated control cells was reached. Moreover, the amount of nuclear extractable androgen receptor was increased; the acquisition of tight nuclear binding capacity was accompanied by hormone-induced receptor phosphorylation.
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