Molecular cloning and characterization of 3beta-hydroxysteroid dehydrogenase/Delta5-Delta4 isomerase cDNAs from Japanese eel ovary.
3beta-hydroxysteroid dehydrogenase/Delta(5)-Delta(4) isomerase (3beta-HSD) is a crucial steroidogenic enzyme which catalyzes an essential step in the biosynthesis of all classes of steroid hormones. Two closely related cDNAs, encoding Japanese eel ovarian types I and II 3beta-HSD, were cloned and characterized. Both cDNAs putatively encoded 375 amino acid residues sharing high sequence homology with those of rainbow trout (71%) and mammalian (approximately 45-50%) 3beta-HSD. Transient expression of types I and II 3beta-HSD in COS-7 cells revealed that both proteins possess 3beta-hydroxysteroid dehydrogenase as well as Delta(5)-Delta(4) isomerase activity for both pregnenolone and dehydroepiandrosterone, with the preference of pregnenolone over dehydroepiandrosterone as substrate, although the type I protein is more active than the type II. By northern blot analysis, a single band of the 3beta-HSD transcript of approximately 1.5kb in length was observed in ovarian tissue and the total transcript abundance of both 3beta-HSDs remained constant throughout ovarian development artificially induced by gonadotropin-rich salmon pituitary homogenate. This lack of change in 3beta-HSD transcript abundance during ovarian development did not correlate with the fluctuation of its enzymatic activity reported previously, which may suggest that changes in 3beta-HSD activity during ovarian development may be, in part, post-transcriptionally regulated in the Japanese eel ovary.
Kazeto
Yukinori
Y
Division of Marine Biosciences, Graduate School of Fisheries Science, Hokkaido University, 3-1-1 Minato-cho, Hakodate, Hokkaido 041-8611, Japan. kazeto@umbi.umd.edu
Ijiri
Shigeho
S
Matsubara
Hajime
H
Adachi
Shinji
S
Yamauchi
Kohei
K
D015067:17-Hydroxysteroid Dehydrogenases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D000595:Amino Acid Sequence; D000735:Androstenedione / Q000378:metabolism; D000811:Anguilla / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D019556:COS Cells; D002522:Chlorocebus aethiops; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D005260:Female; D018507:Gene Expression Regulation, Developmental; D015971:Gene Expression Regulation, Enzymologic; D008969:Molecular Sequence Data; D010053:Ovary / Q000201:enzymology* / Q000254:growth & development; D011374:Progesterone / Q000378:metabolism; D012313:RNA / Q000737:chemistry / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016415:Sequence Alignment
D018076:DNA, Complementary; D000735:Androstenedione; D011374:Progesterone; D012313:RNA; D015067:17-Hydroxysteroid Dehydrogenases; C044256:3 (or 17)-beta-hydroxysteroid dehydrogenase