大口黑鲈GHRH-LP和GHRH基因序列同源性、基因结构和时序表达研究

COMPARISON OF GENE STRUCTURE, SEQUENCE HOMOLOGY AND EXPRESSION PATTERN OF LARGEMOUTH BASS GHRH-LP AND GHRH

促生长激素释放激素(Growth hormone releasing hormone,GHRH)是生长激素释放调控的重要因子。过去人们一直将鱼类的促生长激素释放激素样多肽(Growth hormone releasing hormone like peptide,GHRH-LP)误认为是GHRH,直到最近才分离出GHRH基因。为了了解GHRH和GHRH-LP基因的结构特征及表达差异,研究克隆了大口黑鲈(Micropterus salmoides)GHRH和GHRH-LP基因,同时应用实时定量PCR技术研究了这两个基因的组织表达及发育时序表达情况。结果显示,大口黑鲈GHRH的成熟肽由27个氨基酸残基组成,GHRH-LP的成熟肽由44个氨基酸残基组成。两个基因均由5个外显子和4个内含子组成,但两者成熟肽编码区在外显子上的分布有明显不同。与其他脊椎动物比较,GHRH同源性为74%—100%,而GHRH-LP同源性为41%—96%,两者间具有一定的相似性(37%—63%)。GHRH基因仅在前脑和延脑中表达,而GHRH-LP基因在中枢神经系统及外周组织中均有表达;在胚胎发育过程中GHRH在神经胚后期检测到表达,其表达水平在仔鱼出膜1d后显著提高,而GHRH-LP在囊胚期及后续发育过程中均检测到表达。基因结构、序列同源性及表达谱研究均表明,GHRH和GHRH-LP存在显著差异,应为具有不同功能的两个基因。

Growth hormone releasing hormone（GHRH） is an important factor involved in growth hormone synthesis and secretion.In mammals,the GHRH and GHRH-LP are encoded by two separate genes.However,in non-mammalian vertebrates,it is believed that the GHRH-like peptide is the counterpart of GHRH in mammals.A real GHRH gene was discovered in goldfish and zebrafish recently.Its gene structure and expression distribution in teleost need to be further studied.In order to distinguish the characteristic of GHRH and GHRH-LP genes and further understand the GH secre-tion net in teleost,the cDNA sequences and genome gene structure of these two genes were determined in this study,and the gene expression pattern of GHRH and GHRH-like peptide in central nervous system,peripheral tissues and em-bryogenesis were compared respectively in largemouth bass（Micropterus salmoide）.The results showed that both GHRH and GHRH-LP genes were consisted of 5 exons and 4 introns,and the exon-intron arrangement of genes were apparently different from each other,but showed conserved pattern with other vertebrates,respectively.The GHRH mature peptide was 27 amino acid length,which shared highly homologous with other vertebrates（74%-100%）;while GHRH-LP mature peptide was 44 amino acid length and showed mediate conservation with other vertebrate（41%-96%）.The tissue distribution study showed that the GHRH mRNA expression was restricted in brain and medulla,and that GHRH-LP was widely distributed in central nerves and peripheral tissues.The GHRH mRNA was first detected at neural stage during embryogenesis period,and maintained at a high level during larval developing;GHRH-LP mRNA was first detected in blastula stage,and retain a considerable expression level at all the next stage examined.The results demonstrate that the GHRH and GHRH-LP are encoded by two different genes in largemouth bass.