Characteristics of Neurokinin-3 Receptor and Its Binding Sites by Mutational Analysis

Abstract

NKB (Neurokinin B) is already known to play a crucial role in fish reproduction, but little is known about the structure and function of NKB receptors. Based on an in silico model of the tilapia NKB receptor Tachykinin 3 receptor a (tiTac3Ra) found in the current study, we determined the key residues involved in binding to tilapia NKB and its functional homologue NKF (Neurokinin F). Despite studies in humans suggesting the crucial role of F2516.44 and M2897.43 in NKB binding, no direct peptide interaction was observed in tilapia homologs. In-silico, Ala mutations on residues F2516.44 and M2897.43 did not influence binding affinity, but significantly affected the stability of tiTac3Ra. Moreover, in vitro studies indicated them to be critical to tiNKB/tiNKF-induced receptor activity. The binding of NKB antagonists to tiTac3Ra both in-vitro and in vivo inhibits FSH (follicle stimulating hormone) and LH (luteinizing hormone) release and sperm production in mature tilapia males. Non-peptide NKB antagonist SB-222200 had a strong inhibitory effect on the Tac3Ra activation. SB-222200 also decreased LH plasma levels; two hours post intraperitoneal injection, changed sperm volume and the ratios of the different stages along the spermatogenesis in tilapia testes.

Tachykinins, new players in the control of reproduction and food intake: A comparative review in mammals and teleosts

“…An autocrine/paracrine action is also possible in teleosts as both NKB and NK3R are expressed in the testis [zebrafish (71); tilapia (72)]. In tilapia, recent use of NKB antagonists by ip injections on adult males reduced the number of spermatozoa, leading to lower fertility (251), an effect which can be direct as male tilapia have significant amounts of TACR3 in the testis (72).…”