猪GRK2蛋白抗口蹄疫病毒作用分析  

Evaluation of the Antiviral Effect of Porcine GRK2 against Foot-and-mouth Disease Virus

作　　者：陈文哲 张向乐[2] 顾峰幸 赵振翔 李康丽 薛钊宁 郑海学[2] 张小丽[1] 朱紫祥[2] CHEN Wenzhe;ZHANG Xiangle;GU Fengxing;ZHAO Zhenxiang;LI Kangli;XUE Zhaoning;ZHENG Haixue;ZHANG Xiaoli;ZHU Zixiang(College of Veterinary Medicine,Gansu Agricultural University,Lanzhou 730070,China;State key Laboratory of Veterinary Etiological Biology,College of Veterinary Medicine,Lanzhou University,Lanzhou Veterinary Research Institute,Chinese Academy of Agricultural Sciences,Lanzhou 730046,China)

机构地区：[1]甘肃农业大学动物医学院,兰州730070 [2]中国农业科学院兰州兽医研究所,兰州大学动物医学与生物安全学院,家畜疫病病原生物学国家重点实验室,兰州730046

出　　处：《畜牧兽医学报》2023年第10期4350-4361,共12页ACTA VETERINARIA ET ZOOTECHNICA SINICA

基　　金：国家重点研发计划项目(2021YFD1800300);甘肃省重大专项(21ZD3NA001,20ZD7NA006,19ZDNA001);“十四五”广东省农业科技创新十大主攻方向“揭榜挂帅”项目(2022SDZG02);生猪产业体系(CARS-35);绒毛羊产业体系(CARS-39-13);中国农业科学院基本科研业务费(1610312022006)。

摘　　要：为研究G蛋白偶联受体激酶2(G protein-coupled receptor kinase 2,GRK2)对口蹄疫病毒(foot-and-mouth disease virus,FMDV)复制的影响,从猪的PK-15细胞中提取RNA,通过聚合酶链式反应(polymerase chain reaction,PCR)扩增出猪GRK2的完整CDS序列,并构建了猪GRK2真核表达质粒。Western blot和间接免疫荧光试验表明GRK2可表达并定位于细胞质中。FMDV感染PK-15细胞的GRK2转录水平和蛋白水平呈先上升后下降趋势;过表达GRK2可显著抑制FMDV的复制,下调表达GRK2可显著促进FMDV的复制,表明GRK2在FMDV感染过程中具有抗病毒作用。进一步研究发现FMDV 3C^(pro)蛋白可与GRK2互作并降解GRK2,促进FMDV的复制。研究结果为进一步分析宿主蛋白GRK2的抗FMDV的分子机制奠定了基础。To investigate the effect of G protein-coupled receptor kinase 2(GRK2)on the replication of foot-and-mouth disease virus(FMDV),the porcine cellular RNA from PK-15 cells were extracted and the complete CDS region of GRK2 was obtained by Polymerase Chain Reaction(PCR)methods.Subsequently,the eukaryotic expressing plasmid was constructed.The Western blot and indirect immunofluorescence experiments showed that GRK2 was expressed in the transfected cells and located in the cytoplasm.The infection assay suggested that the transcription level and protein level of GRK2 firstly increased and then decreased in FMDV-infected PK-15 cells.Overexpression assay indicated that GRK2 significantly inhibited FMDV replication,and siRNA assay showed that downregulation of GRK2 promoted FMDV replication,revealing the antiviral function of GRK2 against FMDV.Further studies showed that FMDV 3C^(pro)protein interacted with GRK2 and degraded GRK2 to promote FMDV replication.In conclusion,this study laid a foundation for analysis of the antiviral mechanism of host protein GRK2 against FMDV.

关 键 词：口蹄疫病毒 G蛋白偶联受体激酶2 抗病毒作用 

分 类 号：S852.659.6[农业科学—基础兽医学]