17β-HSD type 12-like is responsible for maturation-inducing hormone synthesis during oocyte maturation in masu salmon

Abstract

The maturation-inducing hormone 17α,20β-dihydroxy-4-pregnen-3-one (DHP) was first identified in the amago salmon. Although carbonyl reductase-like 20β-hydroxysteroid dehydrogenase (CR/20β-HSD) was reported to convert 17α-hydroxyprogesterone (17OHP) to DHP in rainbow trout, we previously found that CR/20β-HSD messenger RNA (mRNA) was not upregulated in stimulated granulosa cells from masu salmon, which suggested that DHP is synthesized by a different enzyme. Accordingly, the current study aimed to identify the specific 20β-hydroxysteroid dehydrogenase (20β-HSD) responsible for DHP production by granulosa cells during final oocyte maturation in masu salmon. RNA sequencing was performed on granulosa layers that were isolated from ovarian follicles at 1 month before ovulation and incubated with or without forskolin, which was used to mimic luteinizing hormone, and ∼12 million reads were obtained, which yielded 71,062 contigs of >100 bp. tBlastx analysis identified 1 contig (#f103496) as similar to 17β-hydroxysteroid dehydrogenase type 12 (hsd17β12); however, because the full-length #f103496 sequence was different from hsd17β12, it was termed hsd17β12-like (hsd17β12l). We found that mammalian cells transfected with full-length hsd17β12l exhibited considerable 20β-HSD activity, as indicated by efficient conversion of exogenous 17OHP to DHP. In addition, we found that hsd17β12l mRNA levels were consistently low in follicles during vitellogenic growth; however, the levels increased significantly during final oocytematuration. The levels of hsd17β12lmRNA were also considerably increased in granulosa layers in which 20β-HSD activity was induced by salmon pituitary extract. Therefore, we suggest that hsd17β12l, not CR/20β-HSD, is the 20β-HSD responsible for DHP production by granulosa cells in masu salmon during final oocytematuration.