作 者：
孙犁;卢霞;孔杰;孟宪红;闫允君;李波波;仲伟鹏;陈宝龙;曹宝祥;罗坤;栾生
单 位：
上海海洋大学水产与生命学院;中国水产科学研究院黄海水产研究所农业农村部海洋渔业可持续发展重点实验室;青岛海洋科学与技术国家实验室海洋渔业科学与食物产出过程功能实验室
关键词：
中国明对虾;竞争行为;钙调神经磷酸酶;基因克隆;表达分析
摘 要：
中国明对虾在养殖中表现出较强的竞争行为,对其生长性能产生了较大的影响,然而,到目前为止其发生的分子机制尚不清楚。钙调神经磷酸酶(calcineurin,CN)是高度保守的Ca~(2+)/钙调蛋白(calmodulin, CaM)依赖性丝氨酸/苏氨酸磷酸酶,由催化亚基(CNA)和调节亚基(CN-B)组成,是参与许多重要生理过程的多功能蛋白质。CNB在Ca~(2+)/CaM的介导下主要在动物的中枢神经系统发挥重要作用。另外,前期通过比较转录组分析筛选出的与中国明对虾竞争行为相关的候选基因中包括CNB基因。为了进一步明确CNB在中国明对虾竞争过程中的作用,实验通过RACE技术克隆了中国明对虾CNB基因(FcCN-B)的全长cDNA序列,并利用Real-time PCR技术分析了其在高竞争能力组(HCG)和低竞争能力组(LCG)组间不同组织(神经节、心脏、胃、肝胰腺和肠)中的表达情况。结果显示,FcCN-B的cDNA全长序列为2 867 bp,包括95 bp的5′非编码区(UTR),540 bp的开放阅读框(ORF),和2 232 bp的3′UTR,其中ORF中具有4个保守的EF-hand Ca~(2+)结合结构域。蛋白质同源性分析显示,FcCN-B的氨基酸序列与其他物种具有较高的同源性(78.8%～93.8%),其中最高的是中华绒螯蟹(93.8%)和黑腹果蝇(90.5%);系统进化关系分析显示,脊椎动物和无脊椎动物分别独立为一支,且中国明对虾与中华绒螯蟹单独聚为一支,之后与黑腹果蝇聚类关系最近,提示FcCN-B在中国明对虾中可能具有与其在中华绒螯蟹和果蝇中相类似的功能。Real-time PCR定量结果显示,FcCN-B在HCG组的神经节中的表达极显著高于LCG组,而其在HCG组的心脏中的表达极显著低于LCG组。研究结果表明,calcineurin B基因在中国明对虾的竞争行为中可能发挥一定的作用,将为解析中国明对虾竞争行为的分子机制奠定重要的基础。
译 名：
Identification and characterization of the calcineurin B in Fenneropenaeus chinensis: its implications in competitive ability
作 者：
SUN Li;LU Xia;KONG Jie;MENG Xianhong;YAN Yunjun;LI Bobo;ZHONG Weipeng;CHEN Baolong;CAO Baoxiang;LUO Kun;LUAN Sheng;College of Fisheries and Life Science, Shanghai Ocean University;Key Laboratory for Sustainable Utilization of Marine Fisheries Resources, Ministry of Agriculture and Rural Affairs,Yellow Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences;Laboratory for Marine Fisheries Science and Food Production Processes, Qingdao National Laboratory for Marine Science and Technology;
关键词：
Fenneropenaeus chinensis;;competitive behavior;;calcineurin B;;gene cloning;;expression analysis
摘 要：
Competitive behavior exists widely in animal and plant groups. When groups or individuals have the same needs for the limited resources, competition will occur in intra-species or inter-species. The existence of competitive behavior can significantly affect the growth performance of individuals. There is strong competitive behavior in the breeding of Fenneropenaeus chinensis, and we have proved that the competitive behavior among individuals of F. chinensis has a significant effect on individual growth traits, and this effect can be inherited.However, little information was available for the molecular mechanism of competition behavior in shrimp.Candidate genes involved in competitive behavior of F. chinensis have been identified by comparative transcriptome analysis in our previous study, among which there is a calcineurin(CN) B gene. CN is a highly conserved Ca~(2+)/calmodulin(CaM)-dependent serine/threonine phosphatase, which is composes of a catalytic subunit(CNA) and a regulatory subunit(CNB). Previous studies reported that CN-B played an important role in the central nervous system mediated by Ca~(2+)/CaM. In order to further prove the role of CN-B gene in the competitive behavior, in the present study, the full-length cDNA of CN-B of F. chinensis(FcCN-B) was cloned by RACE technology, and its expression in different tissues(ganglion, heart, stomach, hepatopancreas, and intestine)between high competitive group(HCG) and low competitive group(LCG) was analyzed by Real-time PCR. The results show that the full-length cDNA of FcCN-B is 2 867 bp, containing 95 bp of 5′ untranslated regions(UTR), 540 bp of open reading frame(ORF), and 2 232 bp of 3′ UTR. There are four conserved EFhand Ca~(2+) binding domains in the ORF of FcCN-B. Protein homology analysis indicated that the amino acid sequence of FcCN-B has high homology with other species(78.8%-93.8%), among which FcCN-B has the highest homology with Eriocheir sinensis(93.8%) and then with Drosophila. melanogaster(90.5%). The phylogenetic relationship analysis showed that vertebrates and invertebrates were independently clustered into two branches.Furthermore, F. chinensis was clustered with E. sinensis into an independent branch, and then it was clustered to D. melanogaster, which suggesting that FcCN-B might have similar functions as it did in D. melanogaster. The results of Real-time PCR showed that the expression of FcCN-B in the ganglion was significantly higher in HCG than in LCG, but its expression in heart was significantly lower in HCG than in LCG. This study preliminarily proved that the calcineurin B may play a certain role in the competition behavior of F. chinensis, and will lay an important foundation for investigating the molecular mechanism of the competition behavior of F. chinensis.
