Abstract:In order to clarify the physiological roles of gonadotropin-releasing hormone 2 (GnRH2) during ovarian maturation in half-smooth tongue sole (Cynoglossus semilaevis), we first cloned the full-length cDNA of GnRH2 by RT-PCR coupled with RACE. Then, the tissue distribution and changes in gnrh2 mRNA levels during ovarian maturation were evaluated by real-time quantitative PCR. Our data showed that the full-length cDNA of GnRH2 was 538 bp in size, excluding the poly-A tail. It consisted of a 5¢ untranslated region (UTR) of 154 bp, a 3¢ UTR of 126 bp and an open reading frame (ORF) of 258 bp which encoded an 85-amino acids preprohormone with a deduced molecular mass and isoelectric point of 9.69 kDa and 8.55, respectively. GnRH2 precursor contained a signal peptide, a mature decapeptide, a processing site, and a GnRH-associated peptide. Multiple sequence alignments indicated that GnRH2 preprohormones were highly conserved among teleosts, especially the decapeptide (QHWSHGWYPG) motif. Tongue sole GnRH2 precursor displayed the highest sequence identities with the order Perciformes (89.41%–90.59%), followed by Pleuronectiformes, Salmoniformes, and Tetraodontiformes (78.82%– 85.88%), and the lowest sequence identities with Cypriniformes (61.18%–71.76%). The tissue distribution showed that gnrh2 transcripts could be detected at the highest levels in the brain and at lower levels in the pituitary and other peripheral tissues. The developmental stages of ovaries were divided into five stages (Ⅱ, Ⅲ, Ⅳ, Ⅴ, and Ⅵ) by histological analysis and the expression patterns of gnrh2 mRNA during ovarian maturation were also investigated. In the brain, the mRNA expression of gnrh2 peaked at stage Ⅲ, declined sharply at stage Ⅳ and reached a minimum at stage Ⅴ. However, the brain gnrh2 mRNA levels ascended again at stage Ⅵ. The pituitary gnrh2 mRNA levels declined gradually, except for a temporary increase at stage Ⅴ. These data indicate that brain GnRH2 appears to be involved in ovarian maturation in the half-smooth tongue sole, and this study extends our knowledge of the roles of GnRH2 on the regulation of reproduction in fish.
