摘要
新型的神经内分泌因子kisspeptin与GnIH(gonadotropin-inhibitory hormone)对哺乳动物的生殖轴起着直接而相反的调控作用,kisspeptin刺激脑垂体促性腺激素(GtH,gonadotropin)的合成与释放,而GnIH抑制GtH的合成与释放。然而,在鱼类中,相关研究甚少。本研究室利用比较基因组学技术,通过同线性分析结合分子生物学手段,在斑马鱼(Danio rerio)、金鱼(Carassius auratus)中克隆得到了kisspeptin(kiss1、kiss2)与GnIH以及它们相关受体的基因cDNA序列。氨基酸序列同源比对结果显示:在脊椎动物中,kisspeptin的核心肽相对保守,而GnIH的核心肽在不同物种中则有较大的差异。比较基因组分析显示kisspeptin与GnIH基因在鱼类、两栖类、鸟类和哺乳类都维持着保守的同线性结构;并且,同线性的结果表明,鱼类kiss1与kiss2基因来源于早期的基因组复制,在漫长的进化过程中,哺乳类丢失了kiss2基因。通过配体受体结合实验,证明2种kisspeptins均能激活其相关受体GPR54(GPR54a、GPR54b),启动下游通路,但却存在一定的配体-受体选择性差异。金鱼、斑马鱼组织表达模式研究显示:kisspeptins与GnIH以及它们相关受体在生殖相关组织或区域(下丘脑、垂体、性腺)均有丰富的表达,提示kisspeptins与GnIH可能参与鱼类的生殖调控。通过化学合成金鱼kisspeptins与GnIH的核心肽,在体注射成熟的雌性金鱼,发现kiss1能显著刺激金鱼LH的分泌,并且能诱导金鱼排卵,而kiss2不能;在高剂量的状况下,GnIH亦能有效抑制金鱼LH的分泌。然而,在离体实验中,2种kisspeptin与GnIH对金鱼垂体细胞LH的分泌均没有显著的影响,提示kisspeptins与GnIH对金鱼的LH调控可能发生于下丘脑水平。以上结果表明:与哺乳动物相类似,在鱼类生殖轴中也存在kisspeptins与GnIH的正负调控系统。本文侧重对这项研究以及相关的研究成果进行归纳与分析,旨为深入探讨鱼类生殖内分泌调控相关功能基因提供参考。
Kisspeptin and GnIH （gonadotropin-inhibitory hormone）, the RFamine peptides, play important roles in mammalian reproductive system. Kisspeptin stimulates the GtH （gonadotropin） release and synthesis via control of GnRH （gonadotropin-release hormone ） secretion. In contrast, GnlH inhibits the GtH release and synthesis directly or via control of GnRH secretion. However, much less is known about these two novel neuropeptide in teleost fish. We have isolated kisspeptide （kiss1, kiss2 ）, GnIH and their putative receptors （GPR54a, GPR54b, GnlHR1, GnlHR2, GnlHR3 ） cDNAs from goldfish and zebrafish by a combination of bioinformatics method and PCR cloning. Genomie syteny analysis revealed that kisspeptin and GnlH were preserved in vertebrates during evolution, and indicated that the two Kisspeptin isoforms were created by ancient duplication events. Sequence analysis showed that the functional regions of kisspeptin （kisspeptin-10s）were relatively conserved in vertebrates, while that of GnlH varied greatly. RT-PCR analysis demonstrated that the expression of the kiss 1 s, gpr54s, GnlH and GnlHRs were observed in the neuronal and reproductive related tissues, including the brain, pituitary, and gonads. Subsequent studies revealed that both mature goldfish kisspeptin-10 peptides （gfKissl-10 and gfKiss2-10 ） are biologically active as they could functionally interact with the two goldfish receptors expressed in cultured eukaryotic cells to trigger the down-stream signaling pathways with different potencies. The actions of gfKiss 1-10, gfKiss2-10 and zebrafish GnlH on luteinizing hormone （LH） secretion were further investigated in vitro and in vivo. Intraperitoneal administration of gfKissl-10 to sexually mature female goldfish could increase the serum LH levels, indicating that this peptide is active in vivo. However, this peptide does not significantly influence LH release from goldfish pituitary cells in primary culture, indicating that the peptide does not exert its actions at the pituitary level. Whereas gfKiss2-10 appears to be a much less potent peptide as it exhibits no significant in vivo bioactivity and is also inactive on the primary pituitary cells. On the other hand, Intraperitoneal administration of the mature zebrafish GnIH peptide （ LPXRFa peptide-3 ） could significantly reduce the basal serum LH level in goldfish. These data indicated that kisspeptin/GPR54 and GnIH/ GnIHRs systems were also operative in teleost fish. [Journal of Fishery Sciences of China, 2010, 17 （ 2 ） ： 363-368]
出处
《中国水产科学》
CAS
CSCD
北大核心
2010年第2期363-368,共6页
Journal of Fishery Sciences of China
基金
国家自然科学基金资助项目(30700611
U0631007)
广东省自然科学基金资助项目(9151027501000081)
高等学校博士学科点专项科研基金(20090171110031)
作者简介
张勇（1978-），男，博士，主要从事鱼类生理学研究．
通讯作者：林浩然，教授，主要从事鱼类生理学研究．E-mail：lsslhr@mail．sysu．edu．cn
