[Studies on phosphorylation of estrogen receptor from porcine uterus]

Estrogen receptor (ER) complex with estrogen interacts with estrogen responsive element (ERE) in target gene promoters and modulates transcription. ER phosphorylation process might be involved in the transcription regulation. To investigate the ER phosphorylation, the author have purified ER from porcine uterine cytosol through heparin-Sepharose column chromatography followed by immunoprecipitation with monoclonal anti ER antibody and protein A-Sepharose. The ER was phosphorylated with [gamma-32P] ATP without any protein kinase, showing that ER has autophosphorylation activity. 63kDa and 120kDa proteins were mainly detected by SDS-PAGE analysis following autoradiography. ER became phosphorylated in the presence of divalent cation such as Mg2+ or Ca2+ in a time-dependent manner and an ATP concentration-dependent manner. Phosphoamino acid analysis showed phosphorylation of serine residue. Phosphorylation was stimulated by estrogen (E2), and phosphorylated receptor was bound to DNA fragment that contained EREs. It is suggested that ER phosphorylation process might participate in transcription regulation.