Urotensin II in GIFT Nile tilapia (Oreochromis niloticus): CDS cloning, tissue distribution, and in vitro regulation of male reproduction

Abstract

The caudal neurosecretory system (CNSS), present in all jawed vertebrates, except sarcopterygians, is considered a major site of urotensin II (UII) secretion. UII, a 12-amino acid peptide with a conserved hexapeptide ring structure, is also secreted by other tissues and found in sarcopterygians. UII has been associated with endocrine regulation, osmoregulation, and several pathophysiological conditions. In this study, CDS of GIFT Nile tilapia (Oreochromis niloticus) UII (tUII) and its receptors UT1 (tUT1) and UT2 (tUT2) were cloned from the CNSS and cerebellum, respectively. Phylogenetic analysis indicated that tUII, tUT1, and tUT2 shared a high homology with the ones of cichlid species, Haplochromis burtoni and Neolamprologus brichardi. Despite variations in precursor peptide sequences, the core sequence of the mature UII peptide remains highly conserved. tUII was predominantly expressed in the CNSS, while tUT1 and tUT2 were widely distributed in the central nervous system (CNS) and peripheral tissues of male and female tilapia. Functional studies revealed that synthetic tUII significantly activated luciferase activity in HEK293T cells transiently transfected with pNFAT-TA-Luc vectors and tUT1 or tUT2. In vitro studies in male GIFT Nile tilapia showed that tUII stimulated mRNA expression of gnrh1, gnrh2, and gnrh3 in a dose-dependent manner by brain fragments, as well as fshβ, lhβ, and gthα by primary culture of pituitary cells. Furthermore, tUII promoted the expression of gnrhr1, gnrhr2, and gnrhr3 in pituitary cells and stimulated mRNA levels of fshr, lhr, arα, cyp11b2, and dmrt1 in testicular tissue. All these stimulatory effects of tUII on gene expression mentioned above were blocked by the non-selective UT antagonist urantide, suggesting for the first time that the actions of tUII were mediated via tUT1 or tUT2. In addition, tUII could significantly stimulate the secretion of testosterone by testis fragments. Taken together, these results suggest that tUII may play a role in reproductive regulation in male GIFT Nile tilapia.