鲈3种视黄醇类X受体基因cDNA的克隆和组织表达

Molecular cloning,tissue specific expression of three retinoid X receptors of Lateolabrax japonicus

摘要:

视黄醇类X受体(retinoid X receptor,RXR)属于配体激活的核受体家族,在调节细胞内各种信号途径中起重要作用。采用RT-PCR和RACE技术,分离到鲈RXR基因3种亚型RXRα、RXRβ和RXRγ的 cDNA序列。结果表明,得到的RXRα是长度为1 686 bp 3′末端不完整的cDNA序列,其中包括364 bp的5′非翻译区和1 322 bp的编码序列,可编码440个氨基酸。RXRβ cDNA全长为1 741 bp,其中包括150 bp的5′非翻译区,256 bp的3′非翻译区和1 335 bp的开放阅读框,共编码444个氨基酸,理论分子量为49.5 ku。RXRγ cDNA全长为1 847 bp,其中5′非翻译区为88 bp,3′非翻译区为397 bp,开放阅读框为1 362 bp,共编码453个氨基酸,分子量约为49.9 ku。氨基酸序列分析显示,鲈RXRα、RXRβ和RXRγ和其它物种的RXRs结构类似,在DNA结合区有P box和D box结构,配体结合区包括12个α螺旋和2个β折叠结构,H12的C端含有AF-2的激活区。系统进化树分析表明,鲈RXRα、RXRβ和RXRγ分别与其他动物的相应亚型聚类。组织分布特异性研究表明,鲈RXRα在肌肉、心脏、眼、大脑、肠、肾脏、脂肪、脾脏、鳃和肝脏10种组织中均有表达,但表达量较低; RXRβ在这10组织中都有较高表达,但心脏中表达量较低;在所检测的10种组织中, RXRγ都有表达,在肌肉、肠 、肾脏、脂肪和脾脏表达量较高。

Abstract:

The retinoid X receptor(RXR)is a member of nuclear hormone receptor family proteins and plays an important role in the regulation of intracellular receptor signaling pathways.Three retinoid X receptor subtypes(RXRα,RXRβ and RXRγ)cDNAs were obtained from Lateolabrax japonicus by RT-PCR and RACE.The obtained cDNA sequence of RXRα was 1 686 bp in length,containing a 5′-untranslated region(UTR)of 364 bp and an uncompleted CDS of 1 322 bp,and encoded a putative protein of 440 amino acids.The full length cDNA sequence of RXRβ was 1 741 bp in length,containing a 5′ UTR of 150 bp,3′UTR of 256 bp and an ORF of 1 335 bp which encoded a putative protein of 444 amino acids with an estimated molecular weight of 49.5 ku.The RXRγ cDNA was 1 847 bp in length with 88 bp of 5′-UTR,397 bp of 3′-UTR and an ORF of 1 362 bp which encoded a putative protein of 453 amino acids with an estimated molecular weight of 49.9 ku.Like other RXRs,the DNA-binding domain(DBD)of L.japonicus RXRα, RXRβ and RXRγ contain P box and D box and the ligand binding domain(LBD)contains 12 α-helices and 2 β-sheets with a ligand-dependent activation function AF-2 in the COOH terminus of the H12. In the phylogenetic tree, L. japonicus RXRα, RXRβ and RXRγ were clustered with corresponding subtypes respectively.The tissue-specific expression of each RXR gene was explored using RT-PCR.The mRNA of RXRα were expressed in all tissues examined with low level.The mRNA of RXRβ were expressed homogeneously in all tested tissues excepting a low expression in heart. RXRγ mRNA were also expressed in all tissues tested with high levels in muscle, intestine,kidney,fat and spleen.The results lay the foundation for further study of the functions of RXRα, RXRβ and RXRγ.