ISOLATION AND EXPRESSION OF RAINBOW-TROUT (ONCORHYNCHUS-MYKISS) OVARIAN CDNA-ENCODING 3-BETA-HYDROXYSTEROID DEHYDROGENASE/DELTA(5-4)-ISOMERASE

A distinct shift in steroidogenesis from testosterone to 17alpha-hydroxyprogesterone occurs in the salmonid ovarian thecal cell layers immediately prior to oocyte maturation, and is a prerequisite for the production of 17alpha,20beta-dihydroxy-4-pregnen-3-one (maturation-inducing hormone of salmonid fishes) by granulosa cells during oocyte maturation. 17alpha-Hydroxylase/17,20-lyase cytochrome P-450 (P-450(17alpha)) and 3beta-hydroxysteroid dehydrogenase/DELTA5-4-isomerase (3beta-HSD) are the two major steroidogenic enzymes involved in the production of 17alpha-hydroxyprogesterone and testosterone. Using mammalian cDNA probes, we isolated and characterized full-length cDNAs encoding these two enzymes from a rainbow trout (Oncorhynchus mykiss) ovarian thecal cell cDNA library. The cloning of 2.4-kilobase cDNA encoding P-450(17alpha) and transient expression of this clone in nonsteroidogenic monkey kidney tumor COS-1 cells have recently been reported (Sakai et al. 1992). We have isolated a 1.4-kilobase cDNA which is hybridized to the mammalian 3beta-HSD cDNAs. Expression of this cDNA in COS-1 cells led to the production of an enzyme which is capable of converting dehydroepiandrosterone to androstenedione. In this study, enzymatic activities and expression of rainbow trout ovarian P-450(17alpha) and 3beta-HSD are discussed in relation of the steroidogenic shift occurring in the ovarian follicle layers.