IMMEDIATE AND TRANSIENT STIMULATION OF PROTEIN-TYROSINE PHOSPHORYLATION BY ESTRADIOL IN MCF-7 CELLS

Estradiol stimulates protein phosphorylation on tyrosine in human brea st cancer MCF-7 cells under conditions of estradiol-stimulated cell gr owth. The stimulatory effect of estradiol has been observed by P-32-la beling of cells followed by purification of proteins using antiphospho tyrosine antibody coupled to agarose and confirmed by immunoblotting a nalysis with antiphosphotyrosine antibody. This stimulation is immedia te (maximal in 10 s) and transient. In addition, it is receptor-mediat ed since estradiol stimulation is prevented by two well-known antiestr ogens, OH-Tamoxifen and ICI 164 384. Estradiol fails to stimulate tyro sine protein phosphorylation of Cos cells which do not express the est radiol receptor. Two substrates of the estrogen stimulated phosphoryla tion on tyrosine with approximate mol wt of 55 and 60 kDa interact wit h a polyclonal antibody raised against amino acids 527-533 of pp60c-sr c (anti-cst.1 antibody). Tyrosine kinase activity of immunoprecipitate s made using either anti cst.1 antibody or the monoclonal 327 antibody specific for pp60c-src shows that kinase(s) strongly related to pp60c -src are immediately and transiently stimulated by estradiol treatment of cells. The present findings provide the first demonstration that a steroid hormone rapidly stimulates tyrosine phosphorylation of target cells and induces functional modifications of substrates of this phos phorylation. These modifications might initiate the estradiol action o n cell growth.