The Effects of Epidermal Growth Factor on c-Fos mRNA Proto-Oncogene Expression in The Human Endometrial Cancer Cell Line , HEC-1-A

Park, J Y; Park, Y K; Kang, J S

Abstract

Growth factors stimulate cell proliferation and differentiation through binding to specific high affinity receptors. Signalling pathway that mediate the normal functions of growth factors are commonly subverted in cancers. Proto-oncogenes are normal cellular genes whose alteration by mutation or deregulation has been implicated in the process of tumorigenesis and they play a role for control of normal cellular growth and differentiation. Epidermal growth factor is mitogenic in moderately differentiated endometrial adenocarcinoma cell line, HEC-1-A. These studies were performed to determine the effects of EGF on c-fos mRNA proto-oncogene expression, and whether EGF need new protein synthesis, and whether PKC pathway plays a role in the induction of c-fos mRNA expression in EGF treated cells. HEC-1-A cells were grown to confluency and maintained in a serum free media (0.3% BSA) 24 hours prior to treatment. The cells were treated with EGF (0, 0.01, 0.1, 1.0, 10, 100 ng/ml) and PMA (0, 0.001, 0.01, 0.1, 1, 10, 100 1000 nM) at varying times (0, 0.12, 0.25, 0.5, 1, 3, 6, 12, 24 h) and the expression of c-fos mRNA expression examined by northern blot analysis. The expession of c-fos mRNA induced by EGF and PMA was dose dependent and peak induction occurred at 1 hour, and decreased steadily after 1 hour. To determine whether PKC may mediate EGF effects on c-fos mRNA expression, the cells were preincubated without or with PMA (1000 nM) for 48 hours for down regulation of PKC, followed by EGF (100 ng/ml). Following 48 hours preincubation with PMA (1000 nM), EGF increased c-fos mRNA expression, while PMA failed to induce its expression, suggesting that EGF induces c-fos mRNA expression independent of PKC activation. To determine whether EGF need new protein synthesis in the induction of c-fos mRNA, the cells were preincubated with anisomycin, a stringent protein synthesis inhibitor, for 30 minutes followed by EGF(100 ng/ml). Anisomycin induced c-fos mRNA and augmented EGF effects on c-fos mRNA expression. These results suggest that EGF may induce c-fos mRNA expression through PKC independent pathway and the induction does not require new protein synthesis.