Preliminary analysis of H3K4me3 in the gonads during gonadal differentiation in the olive flounder Paralichthys olivaceus

Abstract

Abstract Histone methylation is one of the most important epigenetic modifications. However, there are few studies on its role in fish gonadal differentiation. In this study, we analyzed the patterns of H3K4me3 and the expression of modifying- and de-modifying enzymes in the gonads of the juvenile olive flounder Paralichthys olivaceus during the gonadal differentiation process. Histological analysis showed that phenotypic female or male flounder were obtained by treatment with 17β-estradiol (E2) or testosterone (T), respectively. Real-time quantitative PCR (qPCR) results presented that the expression of kmt2b at 2 and 6 cm total length (TL) was higher in the T group than in the E2 group. The expression of kdm5a and kdm5c was higher in the E2 group at 10 cm TL. The results of in situ hybridization (ISH) indicated that kmt2b was mainly distributed in oocytes of the ovary at 10 cm TL. Western blot analysis showed that, H3K4me3 levels were higher in the T group than in the E2 group at 2 and 6 cm TL. As shown with Immunohistochemistry analysis, H3K4me3 was mainly located in oocytes in the ovaries. The signal in spermatogonia was weak in the testes at 10 cm TL. After incubation with an H3K4 methylase inhibitor (5’-deoxy-5’-methylthioadenosine, MTA), the levels of H3K4me3 in the gonads were significantly decreased. The qPCR results indicated that in the MTA group, the expression of sox9a and dmrt1 was significantly decreased in the testis. These results provide the basic data that H3K4me3 may be involved in fish gonadal differentiation.