Phorbol esters increase prolactin synthesis and release and inhibit growth hormone production by GH4C1 cells, a clonal strain of rat pituitary cells. We examined the actions of phorbol esters on the binding of peptides that regulate the function of GH4C1 cells and found that phorbol esters decreased the specific binding of epidermal growth factor (EGF), thyrotropinreleasing hormone (TRH), and somatostatin. 12-O-Tetradecanoyl-13-phorbol acetate (TPA) at 100 ng/ml decreased 125I-EGF binding to 15% of control within 1 hr, 3H-TRH binding to 30% of control after 4 hr, and 125I-Tyr1-somatostatin binding to 40% of control after 24 hr. The action of TPA on 125I-EGF and 3H-TRH binding was temperature dependent, occurring at 37° but not at 4°, and was reversed within 72 hr after removal of TPA. By Scatchard analysis of 125I-EGF-binding data, TPA (100 ng/ml) decreased the concentration dependence of 125I-EGF binding (7- to 10-fold) when binding was assayed at 37° and decreased both the apparent affinity (2- to 6-fold) and the number of 125I-EGF-binding sites (by 25 to 50%) when binding was assayed at 4°. The decrease in 3H-TRH binding induced by TPA (100 ng/ml) was due to a 2- to 5-fold decrease in the apparent affinity of binding at 37°. Thus, unlike findings reported for certain other cell lines, phorbol esters induce an alteration in membrane function in GH4C1 cells which decreases the binding to specific receptors for more than one regulatory peptide. Modulation of receptors for regulatory ligands may play a role in the mechanism by which phorbol esters promote neoplastic transformation, but the action on “growth factor” receptors cannot be considered specific in all cell types.
This investigation was supported in part by Research Grant AM 11011 from the National Institute of Arthritis, Diabetes, Kidney and Digestive Diseases, Center Grant ES 00002, and Cooperative Agreement CR 807809 with the Environmental Protection Agency.