凡纳滨对虾HMIT基因的克隆与表达分析  

Cloning and expression analysis of HMIT gene in Litopenaeus vannamei

作　　者：刘洪涛 杨明秋 汪坤 何玉贵 LU Hongtao;YANG Mingqiu;WANG Kun;HE Yugui(Haincan Prorincial Key Laboratory of Tropical Mariculural Technologies,Hainan Academy of Ocean and Fisheries Sciences,Haikou 571126,China)

机构地区：[1]海南省热带海水养殖技术重点实验室,海南省海洋与渔业科学院海水渔业研究所,海口571126

出　　处：《海洋渔业》2023年第1期14-25,共12页Marine Fisheries

基　　金：海南省属科研院所技术开发专项项目(2018—2019);财政部和农业农村部:国家现代农业产业技术体系(CARS-48);海南省热带海水养殖技术重点实验室建设专项项目(2021)。

摘　　要：为了解H+/肌醇转运蛋白(H+/myoinositol transporter,HMIT)基因在肌醇跨膜转运中发挥的作用,克隆了凡纳滨对虾(Litopenaeus vannamei)的HMIT基因,命名为Lv-HMIT,并分析了Lv-HMIT的结构和表达特征。Lv-HMIT的开放阅读框为1 650 bp,编码549个氨基酸。物理性质显示,它是一种不稳定的亲水性蛋白质,二级结构与三级结构预测都显示其主要由α螺旋和无规则卷曲为主;跨膜结构分析显示,Lv-HMIT存在11个跨膜螺旋结构;亚细胞定位显示,它分布于质膜、内质网和线粒体。多重比对发现,Lv-HMIT的跨膜结构和糖基化位点都比较保守。系统进化显示,Lv-HMIT与雪蟹(Chionoecetes opilio)的HMIT先聚为一支,再与其他节肢动物聚类。实时荧光定量PCR结果显示,Lv-HMIT mRNA在肝胰腺中表达量最高,其次是鳃,在心、胃、眼柄、肠、肌肉和血细胞等组织中有少量表达。Lv-HMIT mRNA从无节幼体期检测到表达,在变态至蚤状幼体、糠虾幼体和仔虾各阶段时都呈现上调表达,说明它可能与幼体变态发育过程有关。十足目虹彩病毒(decapod iridescent virus 1, DIV1)感染24 h后,Lv-HMIT mRNA在肝胰腺中表达显著下调,而鳃中表达量显著上调,显示它参与了对虾对病毒的免疫响应过程。研究结果可为深入了解凡纳滨对虾HMIT的结构功能及在对虾幼体发育、病毒免疫的调控机制提供基础数据。The H+/myoinositol transporter(HMIT) gene, also known as solute carrier family 2 member 13(SLC2A13) or GLUT13, is the only H+-coupled transporter in the glucose transporter family. However, it does not transport glucose, but H+coupling specifically transmembrane transports inositol. It is observed that the addition of moderate amount of inositol can improve the growth performance, non-specific immune enzyme activity, and antioxidant capacity of Litopenaeus vannamei. Inositol is involved in glucose and lipid metabolism, osmotic pressure regulation, signal transduction, and other physiological processes in living organisms, but its absorption and transport mechanisms are largely unknown. In this study, we obtained the full sequence of HMIT gene from the hepatopancreas of L. vannamei by RACE and performed bioinformatics analysis. We used real-time fluorescence quantitative PCR to determine the distribution of HMIT expression in eight tissues, including blood cells, eye stalk, gills, heart, intestine, stomach, hepatopancreas and muscle. We also determined the expression changes of HMIT gene in each early larval developmental stage of L. vannamei, including nauplius, zoea, mysis, postlarvae, and the expression changes of HMIT in hepatopancreas and gills of L. vannamei injected with 100 μL of decapod iridescent virus 1(DIV1) suspension(the infection dose was about 104 viruses per g of shrimp body weight for 24 h). The results showed that an HMIT gene was cloned from L. vannamei named Lv-HMIT, it was also the first report of HMIT gene in crustaceans. The open reading frame of Lv-HMIT was 1 650 bp in length, encoding 549 amino acids, its’ predicted molecular mass was 58 972.42 Da and the theoretical isoelectric point was 5.57. Analysis of physical properties demonstrated that Lv-HMIT was an unstable hydrophilic protein. A total of 70 phosphorylation sites, 2 O-glycosylation sites, and 1 N-glycosylation site were predicted, some of which might affect the structure of the protein and the realization of functions such as

关 键 词：凡纳滨对虾 HMIT 组织表达 发育表达 十足目虹彩病毒 

分 类 号：Q785[生物学—分子生物学] S917.4[农业科学—水产科学]