REGULATION OF CYTOCHROME P450(SCC) SYNTHESIS AND ACTIVITY IN THE OVINE CORPUS-LUTEUM

The rate-limiting step in luteal biosynthesis of progesterone consists of cleavage of the side chain of cholesterol by mitochondrial cytochrome P450 side-chain cleavage enzyme (P450(scc)) to form pregnenolone. Luteal mRNA encoding P450(scc), quantitated on selected days of the 16-day ovine estrous cycle, was similar on days 3 and 6, increased by 2-fold on day 9 (P < 0.05) and remained elevated on day 15. Levels of P450(scc) mRNA on day 15 of pregnancy were not different from those found on any day of the cycle (P < 0.05). To determine whether levels of mRNA encoding P450(scc) are hormonally regulated, ewes on day 10 of the estrous cycle were injected with hCG or prostaglandin F-2 alpha (PGF(2 alpha)). P450(scc) mRNA was not increased for up to 36 h after injection of hCG, nor decreased within 8h after injection of PGF(2 alpha) (P < 0.05). An assay for P450(scc) activity was developed which utilized ovine small and large luteal cells in the presence of 22R-hydroxycholesterol and ovine high density lipoprotein. Enzyme activity was quantitated by measurement of progesterone production. In small luteal cells activation of the protein kinase A (PKA) second-messenger system by treatment with LH resulted in 910% increase in progesterone production without altering activity of P450(scc). Activation of the protein kinase C (PKC) second-messenger system with phorbol 12-myristate 13-acetate caused a 51% reduction in progesterone secretion from large luteal cells but did not alter activity of P450(scc). These findings suggest that in mature luteal tissue steady state levels of mRNA encoding P450(scc), and enzyme activity are independent of acute regulation by activation of PKA or PKC second-messenger systems.