Hormonal and genetic regulation of steroidogenic P450 enzymes in Leydig cells.

Treatment of Leydig cells in culture with cAMP or testosterone causes oxygen-mediated decreases in the amount of P450$\sb{17\alpha}$ which are comparable to decreases in P450$\sb{17\alpha}$ activity. The testosterone-induced decrease in P450$\sb{17\alpha}$ protein is not due to a decrease in de novo synthesis of the protein, but increases the rate of decay of P450$\sb{17\alpha}$. The effect of testosterone is neither a general effect of steroids nor androgen receptor-mediated. Product-induced, oxygen-mediated decreases in cholesterol side-chain cleavage (P450$\sb{\rm scc}$) enzyme protein are also observed and are due to pregnenolone accumulation. The data suggest that pregnenolone and testosterone decrease the activity of steroidogenic P450 enzymes in Leydig cells by enhancing the generation of oxygen-derived free radicals that directly or indirectly damage the enzyme and lead to increased enzyme degradation. The relative amounts of P450$\sb{\rm scc}$, P450$\sb{17\alpha}$ and ferredoxin reductase, as well as maximal testosterone production, were quantitated in Leydig cells and adrenal glands from RF/J, SWR/J, C3H/He and DBA/2 inbred mice. P450$\sb{\rm scc}$ is higher in Leydig cells from RF/J and SWR/J than in DBA/2 and C3H/He, but, no significant difference in P450$\sb{17\alpha}$ can be detected. Ferredoxin reductase is significantly higher in Leydig cells of RF/J, SWR/J and C3H/He than from DBA/2. Maximal testosterone production by Leydig cells from these strains is correlated with the amount of P450$\sb{\rm scc}$, suggesting that the amount of P450$\sb{\rm scc}$ determines the capacity of Leydig cell testosterone production. P450$\sb{17\alpha}$ was undetectable in the adrenal glands. The pattern of expression of ferredoxin reductase and P450$\sb{\rm scc}$ in adrenals differs from Leydig cells. The results indicate that both genetic and tissue-specific factors influence the expression of P450$\sb{\rm scc}$, P450$\sb{17\alpha}$ and ferredoxin reductase in the steroidogenic tissues of mice. Preliminary studies identified a missing Hinf I restriction fragment in the Southern blot hybridization pattern of the P450$\sb{\rm scc}$ gene of C3H/He mice. This variant can be used as a genetic marker in future studies designed to test the relationship between the P450$\sb{\rm scc}$ gene, the amount of P450$\sb{\rm scc}$ per Leydig cell, and maximal testosterone production by Leydig cells.Ph.D.BiochemistryPure SciencesUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/128281/2/8907124.pd