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Molecular cloning and expression during spermatogenesis of a cDNA encoding testicular 11β‐hydroxylase (P450 11β ) in rainbow trout ( Oncorhynchus mykiss )
Author(s) -
Kusakabe Makoto,
Kobayashi Tohru,
Todo Takashi,
Mark Lokman P.,
Nagahama Yoshitaka,
Young Graham
Publication year - 2002
Publication title -
molecular reproduction and development
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.745
H-Index - 105
eISSN - 1098-2795
pISSN - 1040-452X
DOI - 10.1002/mrd.10145
Subject(s) - biology , complementary dna , rainbow trout , endocrinology , medicine , spermatogenesis , microbiology and biotechnology , northern blot , gene , genetics , fishery , fish <actinopterygii>
Abstract Cytochrome P450 1β‐hydroxylase (P450 11β ) is an important steroidogenic enzyme for glucocorticoid and mineralocorticoid production in vertebrates. In teleosts, P450 11β also plays a role in the production of 11‐ketotestosterone (11‐KT), the predominant androgen in male fish. In this study, we cloned a cDNA encoding rainbow trout ( Oncorhynchus mykiss ) testicular P450 11β . The cDNA contains 1,740 nucleotides that encode a protein of 551 amino acids which shares 65.2% homology with testicular P450 11β from Japanese eel, and 33–45% homology with adrenal P450 11β from rat, human, and frog. HEK293 cells transfected with an expression vector containing the rainbow trout P450 11β cDNA open reading frame showed 11β‐hydroxylating activity in the presence of exogenous testosterone. Analysis of tissue distribution by RT‐PCR showed great abundance of P450 11β mRNA in testis and head kidney. In order to clarify the sites of P450 11β gene expression, cRNA in situ hybridization analysis was performed. Hybridization signals were detected in Leydig cells and head kidney inter‐renal cells. The results of Northern blot analysis indicated a single 1.8‐kb transcript encoding P450 11β in testis and in head kidney, suggesting that the testicular form of P450 11β may also be involved in cortisol production by inter‐renal cells. Seasonal changes in total P450 11β mRNA levels in testes during spermatogenesis showed a pattern similar to that of plasma androgens. These results suggest that androgen production in male rainbow trout is partially regulated by changes in abundance of P450 11β mRNA. Mol. Reprod. Dev. 62:456–469, 2002. © 2002 Wiley‐Liss, Inc.