虹鳟补体活化调节因子CD46的分子特征及功能  

THE MOLECULAR CHARACTERISTICS AND REGULATORY FUNCTION OF COMPLEMENT ACTIVATION REGULATOR CD46 IN RAINBOW TROUT

作　　者：梁佳欣 马自有 李文硕 王鹏 田丁 刘安琪 张永安 张旭杰 LIANG Jia-Xin;MA Zi-You;LI Wen-Shuo;WANG Peng;TIAN Ding;LIU An-Qi;ZHANG Yong-An;ZHANG Xu-Jie(State Key Laboratory of Agricultural Microbiology,College of Fisheries,Huazhong Agricultural University,Wuhan 430070,China)

机构地区：[1]华中农业大学水产学院,农业微生物学国家重点实验室,武汉430070

出　　处：《水生生物学报》2023年第8期1269-1277,共9页Acta Hydrobiologica Sinica

基　　金：国家自然科学基金(31602184);国家重点研发计划“蓝色粮仓科技创新”专项(2018YFD0900505)资助。

摘　　要：为进一步研究硬骨鱼类中补体活化调节因子的分子特征和功能,研究克隆了虹鳟(Oncorhynchus mykiss)的CD46基因,对其分子特征进行了系统分析,结果显示:虹鳟CD46基因由10个外显子和9个内含子组成,cDNA序列全长2812 bp,编码317个氨基酸,蛋白序列由1个信号肽、4个SCR结构域、1个跨膜区和1个胞内区组成,预测分子量为33.9 kD。基因组共线性分析显示,虹鳟CD46基因位于16号染色体,其基因座在脊椎动物中具有保守的共线性。组织和白细胞亚群表达分析显示,虹鳟CD46基因在各种组织和白细胞亚群中均有表达。为了进一步阐明虹鳟CD46的免疫功能,研究原核表达纯化了标签蛋白GST和融合蛋白GST-CD46。溶血活性实验表明,与GST相比,GST-CD46能够显著抑制虹鳟血清对兔红细胞的溶血活性,且呈现剂量依赖效应,表明虹鳟CD46是补体活化的调节因子。此外,研究用HEK293T细胞过表达了GFP和GFP-CD46。细胞损伤实验显示,与GFP相比,GFP-CD46能够显著抑制虹鳟血清对HEK293T细胞的损伤,进一步表明虹鳟CD46是补体活化的调节因子,能够保护细胞免受补体系统的损伤。总之,研究不仅增加了对虹鳟CD46的分子特征和功能的认知,还为深入研究此分子的免疫功能及其在抗病免疫中的应用提供了理论基础。To further study the molecular characteristics and functions of complement activation regulators in teleost fish,we cloned the CD46 gene of rainbow trout(Oncorhynchus mykiss)and analyzed its molecular characteristics systematically.The results showed that the CD46 gene of rainbow trout was composed of 10 exons and 9 introns;the cDNA sequence of CD46 was 2812 bp and encoded 317 amino acids;the protein sequence of CD46 was composed of 1 signal peptide,4 SCR domains,1 transmembrane region and 1 intracellular region;and the predicted molecular weight of CD46 was 33.9 kD.Synteny analyses showed that the rainbow trout CD46 gene was located on chromosome 16,and its gene locus had conserved synteny in vertebrates.The expression analyses showed that the rainbow trout CD46 gene was expressed in various tissues and leukocyte subpopulations.To further clarify the immune function of rainbow trout CD46,the GST protein and the fusion protein GST-CD46 were prokaryotically expressed and purified.The experiment of hemolytic activity showed that,compared with GST,GST-CD46 could significantly inhibit the hemolytic activity of rainbow trout serum to rabbit erythrocytes in a dose-dependent manner,indicating that rainbow trout CD46 is a regulator of complement activation.In addition,GFP or GFP-CD46 were overexpressed in HEK293T cells.The experiment of cell damage showed that,compared with GFP,GFP-CD46 could significantly inhibit the damage of rainbow trout serum to HEK293T cells,further indicating that rainbow trout CD46 is a regulator of complement activation and can protect cells from the damage of complement system.In conclusion,this study not only increased the knowledge of the molecular characteristics and regulatory function of rainbow trout CD46,but also provided a theoretical basis for the in-depth study of the immune function of this molecule and its application in disease resistance.

关 键 词：CD46 补体 调节因子 虹鳟 

分 类 号：Q344.1[生物学—遗传学]