家鸡G蛋白偶联受体85基因的结构、序列与组织表达分析

G蛋白偶联受体(GPCR)超家族是细胞膜上广泛存在的一类受体,是细胞跨膜信号转导的一类重要受体分子,参与许多生理过程调节。它们中仍有很多至今尚未找到内源性配体,这类受体被称为孤儿型受体。G蛋白偶联受体85(GPR85)是GPCR超家族中孤儿型受体的一员。目前,在非哺乳类脊椎动物中,针对GPR85的研究极少。本研究以家鸡Gallus gallus domesticus为模型,通过反转录PCR和RACE-PCR等方法从脑中克隆到GPR85基因的cDNA全长序列,揭示其基因结构,并用实时荧光定量PCR(qPCR)方法探究了该基因在家鸡各组织中的表达情况。结果显示:家鸡GPR85基因位于1号染色体上,由2个外显子组成,其编码区位于第2个外显子上,长为1 113 bp,可编码1个370个氨基酸的7次跨膜受体蛋白。家鸡GPR85与其他脊椎动物(人Homo sapiens、小鼠Mus musculus、大鼠Rattus norvegicus、热带爪蟾Xenopus tropicalis和斑马鱼Danio rerio)的GPR85具有高度的氨基酸序列一致性(>93%)。qPCR分析发现,GPR85基因mRNA在家鸡全脑、垂体、肾上腺、精巢中有较高表达,而在所检测的其他外周组织中表达极低。本研究首次揭示了家鸡GPR85基因的结构与表达特征,为后续探究GPR85基因在家鸡等非哺乳类中的生理功能奠定基础。

Gene Structure,Sequence and Tissue Expression of Chicken G Protein-Coupled Receptor 85 Gene

G protein-coupled receptor(GPCR)is a kind of membrane receptor superfamily.It is an important receptor on cell transmembrane to transmit signals and regulates many important physiological processes.Because the endogenous ligands for some GPCRs have not been identified,these GPCRs are still classified as orphan receptor.G protein-coupled receptor 85(GPR85)is a member of orphan receptor in GPCR superfamily.Up till the present,the information regarding on the structure and expression of GPR85 gene in non-mammalian vertebrates is limited.In this study,by using RT-PCR and RACE PCR,we cloned the full cDNA sequence of GPR85 gene from chicken brain and revealed its gene structure.Furthermore,by using quantitative real-time PCR(qPCR),we also examined the mRNA expression of GPR85 gene in various chicken tissues.The results showed that the chicken GPR85 gene locates on chromosome 1 and consists of 2 exons.The coding region is located on exon 2.Chicken GPR85 gene has an open reading frame of 1 113 bp and encodes a seven-transmembrane receptor.Chicken GPR85 shows extremely high amino acid sequence identity(>93%)with that of Homo sapiens,Mus musculus,Rattus norvegicus,Xenopus tropicalis and Danio rerio.The result of qPCR revealed that chicken GPR85 gene is highly expressed in the brain region,anterior pituitary,adrenal glands and testis,but weakly expressed in other remaining tissues examined.To our knowledge,this study represents the first report concerning the gene structure and expression of GPR85 in chicken and lays the foundation for further exploring the physiological roles of GPR85 in chickens.