Kisspeptin, encoded by kiss gene, plays an important role in reproduction, and it has been proven that kisspeptin stimulate the synthesis and releasing of gonadotropins (GtHs) via controlling the secretion of GnRH in vertebrate.As known, reproduction is controlled by the environment and endocrine factors, especially the pulsative release of GtHs around estrous.However, the feedback of estrogen on the pulsatile release of GtHs remains unclear.Interestingly,the GnRH neurons dont expression steroid receptor(ER), but the majority of kisspeptin neurons express ER, androgen receptor (AR), and progestrion receptor (PR) in most vertebrate.So the kiss neurons have been recognized as the "miss link" in the steroid feedback on GnRH release.The kisspeptin system has been identified in various vertebrate, and two paralogous genes of kisspeptin (kiss1 and kiss2) have been identified in several teleosts, implying the multiplicity of their physiological functions in teleosts.In the present study, two kiss genes in orange-spotted grouper have been identified.To investigate the molecular regulations of sex steroids on the two kiss genes, 5flank regions of kiss1 (1777bp) and kiss2 (1726bp) in orange-spotted grouper were cloned, and the presence of putative binding sites, glucocorticoid receptors, Sp 1, AP 1 and estrogen relative element (ERE) also been analyzed in silico analysis.Then the E2-induced two kiss gene promoter activities were asseyed using a luciferase reporter vector on the presence of Ers.While under the presence of ERβ2, E2 significantly reduced kiss1 and kiss2 promoter activities in HEK-293T cells.But when HEK-293T cells were cotransfected with pcDNA-Erα two kiss promoters, no significant variations was observed.The novel molecular mechanisms of ERβ2 mediating E2 down-regulate kiss1 and kiss2 ion in HEK-293T cells remain to be further identified.