Mechanism of 17-β-estradiol-induced Erk1/2 activation in breast cancer cells

Activation of mitogen-activated protein kinase (Erk/ MAPK) is a critical signal transduction event for estrogen (E-2)-mediated cell proliferation. Recent studies from our group and others have shown that persistent activation of Erk plays a major role in cell migration and tumor progression. The signaling mechanism(s) responsible for persistent Erk activation are not fully characterized, however. In this study, we have shown that E-2 induces a slow but persistent activation of Erk in MCF-7 breast carcinoma cells. The E-2-induced Erk activation is dependent on new protein synthesis, suggesting that E-2-induced growth factors play a major role in Erk activation. When MCF-7 cells were treated with E-2 in the presence of an anti-HER-2 monoclonal antibody (herceptin), 60-70% of E-2-induced Erk activation is blocked. In addition, when untreated MCF-7 cells were exposed to conditioned medium from E-2-treated cells, Erk activity was significantly enhanced. Furthermore Erk activity was blocked by an antibody against HER-2 or by heregulin (HRG) depletion from the conditioned medium through immunoprecipitation. In contrast, epidermal growth factor receptor (Ab528) antibody only blocked 10-20% of E-2-induced Erk activation, suggesting that E-2-induced Erk activation is predominantly mediated through the secretion of HRG and activation of HER-2 by an autoctine/paracrine mechanism. Inhibition of PKC-delta-mediated signaling by a dominant negative mutant or the relatively specific PKC-delta inhibitor rottlerin blocked most of the E-2-induced Erk activation but had no effect on TGFalpha-induced Erk activation. By contrast inhibition of Ras, by inhibition of farnesyl transferase (Ftase-1) or dominant negative (N17)-Ras, significantly inhibited both E-2- and TGFalpha-induced Erk activation. This evaluation of downstream signaling revealed that E-2-induced Erk activation is mediated by a In HRG/HER-2/PKC-delta/Ras pathway that could be crucial for E-2-dependent growth-promoting effects in early stages of tumor progression.