PROTEIN-KINASE-A AND PROTEIN-KINASE-C DIFFERENTIALLY REGULATE STEROIDOGENESIS IN HUMAN OVARIAN THECAL TUMOR-CELLS

The stimulatory role of protein kinase A in thecal cell steroidogenesi s is well documented, whereas the role of protein kinase C is not well defined. In this study, using monolayer cultures of human ovarian tum or cells that are steroidogenically similar to thecal cells, we examin ed the effects of the protein kinase C activator tetradecanoylphorbol- 13-acetate (TPA) on steroidogenesis and the expression of 17 alpha-hyd roxylase cytochrome P450 (P450c17), 3 beta-hydroxysteroid dehydrogenas e (3 beta HSD), and cholesterol side-chain cleavage cytochrome P450 (P 450scc). Cells were uniformly plated and grown to confluence prior to experimental treatment in serum-free medium. Treatments were control, forskolin (10 mu M), TPA (0.01-1000 nM), and TPA with forskolin. Treat ment with TPA alone for 24 h had little effect on basal steroid produc tion, enzyme activities, or mRNA levels. However, when added with fors kolin, TPA augmented progesterone production in a concentration-depend ent manner. In contrast, TPA inhibited forskolin stimulation of andros tenedione production and P450c17 activity. To define better the mechan ism of TPA action, Northern analysis of P450c17, P450scc, and 3 beta H SD mRNA was accomplished using total RNA isolated from cells treated f or 24 h. 3 beta HSD mRNA was increased by forskolin and was not signif icantly inhibited by treatment with TPA. P450c17 mRNA, however, was su ppressed to near undetectable levels by TPA at doses as low as 1 nM. I n addition, P450scc mRNA expression was inhibited in a manner similar to that seen for P450c17 mRNA. In summary, activation of the protein k inase A pathway increases expression of 3 beta HSD, P450c17, and P450s cc in this thecal cell model. Simultaneous activation of protein kinas e A and protein kinase C enhances progesterone production while decrea sing androstenedione production and the levels of mRNA encoding P450c1 7 and P450scc. This differential regulation of steroidogenesis suggest s that protein kinase C may play a role in decreased androstenedione p roduction during thecal cell luteinization.