Alternative splicing of GnRH2 and GnRH2-associated peptide plays roles in gonadal differentiation of the rice field eel, Monopterus albus

The rice field eel, Monopterus albus, is a protogynous hermaphrodite fish, in which the gonads are initially female ovaries which then transform into male testes. The exact mechanisms governing sex reversal in the rice field eel are unknown. In this study, a novel alternative splicing variant of GnRH2 (GnRH2-SV), retaining the second intron, was discovered in the gonad of the rice field eel. Compared to GnRH2, GnRH2-SV may give rise to a novel truncated GnRH2-associated peptide (New GAP2). The normal transcript of GnRH2 was primarily expressed in the brain, and could also be detected in the liver, spleen, ovary, and testis. However, GnRH2-SV was only expressed in the ovary and testis. During sex reversal, GnRH2 expression levels increased significantly at late stages; however, expression levels of GnRH2-SV were lower in ovary than in ovotestis and testis. We also examined the effect of three peptides (GnRHa, GAP2, and New GAP2) on gonadal sex differentiation during the third stage of ovarian development of the rice field eel. Compared to the control group, the expression of amh increased significantly following incubation with each of the three peptides. However, only New GAP2 stimulated the expression of sox9a1 mRNA in vitro. After intraperitoneal injection of GAP2, the expression of amh, foxl2, and cyp19a1a increased significantly after 12 h; the concentration of serum 11-KT was also significantly increased at the 12 h time point. Treatment with New GAP2 significantly increased the expression of amh, dmrt1a, and sox9a1, and also increased the concentration of serum 11-KT. After treated with GnRHa, the expression of amh, dmrt1a, sox9a1, cyp19a1a, and foxl2 increased significantly, as did the level of serum E2. These results indicated that both GAP2 and New GAP2 play a crucial role in inducing expression changes of sex-differentiation related genes, and may be involved in the gonadal development and sex reversal in the rice field eel.