鲈鱼钙蛋白酶基因cDNA克隆及表达分析

Molecular cloning and tissue distribution of calpain gene from Lateolabrax japonicus

钙蛋白酶超家族(Calpain,CAPN,EC3.4.22.17)是由一类钙依赖性半胱氨酸蛋白酶组成。其在细胞周期、细胞迁移、信号转导以及细胞凋亡中都有着广泛的作用。研究了鲈鱼催化亚基CAPN2的基因结构。鲈鱼(Lateolabrax japonicus)CAPN2 cDNA全长2373 bp,5'非翻译区74 bp、3'非翻译区205 bp、开放阅读框2094 bp,编码一个由697个氨基酸组成的蛋白质,分子量为77.94kDa,等电点为5.02。氨基酸序列分析表明,鲈鱼CAPN2具有较高的保守性,与大西洋鲑(Hippoglossus hippoglossus)、虹鳟(Oncorhynchus mykiss)、斑马鱼(Danio rerio)、斑点叉尾鮰(Ictalurus punctatus)、猪(Sus scrofa)、人(Homo sapiens)、大鼠(Mus musculus)等7个物种的同源性分别为89%、81%、77%、75%、63%、62%、62%。用RT-PCR分析CAPN2基因在10个组织中均有表达,肌、心、肾表达较高,脑、鳃、肠次之,肝、眼、脂肪、脾表达最低。

Calpain （CAPN, EC3.4. 22. 17） is Ca^2＋ -dependent cysteine proteinase. It was implicated in a wide range biology proces- ses including regulation of cell cycle, cell migration, intracellular signaling, and apoptosis. The molecular characterization of the Late- olabrax japonicus catalytic subunits of the m-calpains was described. The cDNA of the CAPN2 in Lateolabrax japonicu~ is 2373 bp in size, with 74 bp 5＇-UTR, 205 bp 3＇-UTR and 2094 bp ORF, encoding a protein of 697 amino acids with a molecular weight of 77.94kDa and pI of 5.02. The sequence alignment analysis indicated that the deduced amino acid sequence of CAPN2 shared high i- dentity with seven other species： Hippoglossus hippoglossus（ 89% ） , Oncorhynchus mykiss（ 81% ）, Danio rerio（ 77% ）, Ictalurus punc- tatus （75 % ） , Sus scrofa （63 % ） , Homo sapiens （62%） and Mus musculus （62%）. Semi-quantitative RT-PCR was used to characterize the expression of CAPN2 in ten tissues including muscle, heart, eye, brain, gill, liver, intestine, kidney, adipose tissue and spleen. The results showed that CAPN2 was mainly expressed in the muscle and kidney.