大鳞副泥鳅和泥鳅GNB2L1基因的克隆、表达及系统进化分析

由G蛋白β2亚基类似物1基因(GNB2L1)编码的蛋白激酶C受体(RACK1)是一个高度保守的锚定蛋白,属于WD40结构域蛋白家族成员,在细胞信号转导等生命过程中发挥着重要作用。本文采用RACE技术和基因克隆技术分别对大鳞副泥鳅(Paramisgurnus dabryanus)和泥鳅(Misgurnus anguillicaudatus)精巢组织的GNB2L1基因c DNA序列进行了克隆。序列分析表明,大鳞副泥鳅GNB2L1基因c DNA序列全长1 115 bp,开放阅读框(ORF)长965 bp,编码317个氨基酸;泥鳅GNB2L1基因c DNA序列的开放阅读框长965 bp,编码317个氨基酸;两种泥鳅GNB2L1基因编码的蛋白与其他鱼类的RACK1蛋白的同源性为94%~97%,且不同进化地位物种的GNB2L1基因均由8个外显子和7个内含子组成。以GNB2L1基因为标记基因,构建的鱼类系统发育树显示,大鳞副泥鳅和泥鳅在进化上的亲缘关系最近。RT-PCR结果显示,GNB2L1基因在大鳞副泥鳅成体各组织中均有表达,且在脑组织的表达量高于其他组织。以上结果表明,GNB2L1基因为一个进化保守基因,可能在大鳞副泥鳅的细胞活动中发挥着重要作用。

Gene Structure, Expression and Phylogenetic Analysis of GNB2L1 Gene in Two Chinese Loaches

As a member of the tryptophan-aspartate repeat (WD-repeat) family of proteins, the receptor for activated C kinase 1 (RACK1) is an intracellular adaptor protein that plays an important role in the regulation of various signaling pathways. In this study, the guanine-nucleotide-binding protein beta polypeptide 2-like 1(GNB2L1) gene encoding RACK1 protein was obtained from the gonad of Paramisgurnus dabryanus by rapid amplification of cDNA ends (RACE). The full-length cDNA of the GNB2L1 gene is 1 115 bp and contains an open reading frame of 965 bp, encoding for a putative RACK1 protein with 317 amino acids. The GNB2L1 gene was also isolated from the gonad of Misgurnus anguillicaudatus by gene cloning technique, of which the encoding cDNA sequence is 965 bp long and encodes a putative RACK1 protein with 317 aa. Sequence analysis showed that RACK1 proteins of two loaches display 94%-97% sequence similarities to those of other teleost (Fig.1), and the GNB2L1 genes in vertebrate, including in teleost, have seven introns and eight exons (Fig.2). The phylogenetic tree based on multiple sequence alignment of GNB2L1 genes of two loaches and other closely related teleost revealed that the closest relationship to M. anguillicaudatus and P. dabryanus (Fig.4). Semi-quantative RT-PCR confirmed that GNB2L1 gene is expressed ubiquitously in many P. dabryanus tissues, especially higher in brain (Fig.3). These results implied that GNB2L1 gene is strongly conserved through evolution, and may be essential for cellular functions in P. dabryanus.