EFFECTS OF 12-O-TETRADECANOYLPHORBOL-13-ACETATE ON ESTROGEN-RECEPTOR ACTIVITY IN MCF-7 CELLS

The effects of long term treatment with 12-O-tetradecanoylphorbol-13-a cetate (TPA) on estrogen receptor (ER) expression in the human breast cancer cell line, MCF-7, were studied. This study demonstrates that tr eatment of cells with the phorbol ester blocked estrogen receptor acti vity. Treatment of cells with 100 nM TPA resulted in an 80% decrease i n the level of ER protein and a parallel decrease in ER mRNA and bindi ng capacity. Following removal of TPA from the medium, the level of ER protein and mRNA returned to control values; however, the receptor fa iled to bind estradiol. These cells also failed to induce progesterone receptor in response to estradiol, In addition, TPA treatment blocked transcription from an estrogen response element in transient transfec tion assays and inhibited ER binding to its response element in a DNA mobility shift assay. The estrogen receptor in treated cells was recog nized by two monoclonal anti-ER antibodies and was not quantitatively different from ER in control cells. RNase protection analysis failed t o detect any qualitative changes in the ER mRNA transcript. Mixing exp eriments suggest that TPA induces/activates a factor which interacts w ith the ER to block binding of estradiol. The effects of TPA on ER lev els and binding capacity were concentration-dependent. Low concentrati ons of TPA inhibited estradiol binding without a decrease in the level of protein, whereas higher concentrations were required to decrease t he level of ER protein. The effects of TPA appear to be mediated by ac tivation of protein kinase C since the protein kinase C inhibitors, H- 7 and bryostatin, block the effects of TPA on estradiol induction of p rogesterone receptor. TPA treatment had no effect on the level or bind ing capacity of the glucocorticoid receptor, indicating that the effec ts are not universal for steroid receptors. These data demonstrate tha t activation of the protein kinase C signal transduction pathway modul ates the estrogen receptor pathway. The long term effect of protein ki nase C activation is to inhibit estrogen receptor function through ind uction/activation of a factor which interacts with the receptor.