Cloning and expression analysis of E3 ubiquitin ligase Nrdpl from grass carp ctenopharyngodon idella | 草鱼E_3泛素连接酶Nrdp1基因cDNA克隆和表达分析

Chinese. 神经调节素受体降解蛋白1（neuregulin receptor degradation protein-1，Nrdpl）是一种新的E3泛素连接酶，可调节细胞增殖和凋亡等。研究采用RACE方法从草鱼（Ctenopharyngodon idella）肝脏中克隆了Nrdpl的全长cDNA序列；采用半定量RT-PCR方法分析该基因在草鱼不同组织中表达情况。结果表明：该序列全长cDNA大小为1 865bp（GenBank登录号：JX864048），其中开放阅读框为954bp，编码318个氨基酸，预测分子量大小为36.09ku，pH为7.0时理论等电点为5.83；5’和3’非翻译区的长度分别为291bp和620bp，在3’非翻译区发现2个mRNA不稳定信号（ATTTA）和1个多聚腺苷酸加尾信号（ATTAAA）。该蛋白序列没有信号肽和跨膜结构，其二级结构主要以α-螺旋为主，不含β-折叠。氨基酸序列保守性分析表明，草鱼Nrdp1与斑马鱼、虹鳟及人的同源性分别为98%、94%和89%；该基因在心脏和脑中的表达量较多，在肌肉和血液中表达很少。

English. Nrdpl (neuregulin receptor degradation protein-1) is a newly defined E3 ubiquitin ligase and regulates the cell proliferation and apoptosis. In the present study, the cDNA sequence of grass carp Ctenopharyngodon idella Nrdpl (gcNrdpl) consists of 1865 bp with a 291 bp 5’ untranslated region (UTR) and a 620 bp 3’ UTR, with an open reading frame of 954 bp encoding the 318 amino acids which is predicted to have no signal peptide and transmembrane helices. The deduced amino acid sequence of gcNrdpl showed 98%, 94% and 89% of identity to zebra fish Danio rerio, rainbow trout Oncorhynchus mykiss and human Homo sapiens Nrdpl, respectively. The domain search revealed that gcNrdpl contains a RING domain (at amino acids 18 - 56) and Pfam domain (at amino acids 137 – 315). Phylogenetic analysis demonstrated that gcNrdpl is clustered in a same clade with zebra fish and rainbow trout Nrdpl. RT-PCR demonstrated that the mRNA is constitutively expressed in all the examined organs examined of healthy fish, with the higher level in heart and brain, and with the lower level in muscle and blood. Therefore, this study would be significant to the discussion on genetic structure and function of Nrdpl from the fish.