猪流行性腹泻病毒nsp1蛋白对IFN-β激活的JAK/STAT信号通路的影响  

Effect on IFN-β-activated JAK/STAT signaling pathway by nonstructural protein 1 of porcine epidemic diarrhea virus

作　　者：施开创[1] 尹彦文[1] 陆文俊[1] 屈素洁[1] SHI Kai-chuang;YIN Yan-wen;LU Wen-jun;QU Su-jie(Guangxi Center for Animal Disease Control and Prevention,Nanning 530001,China)

机构地区：[1]广西动物疫病预防控制中心,广西南宁530001

出　　处：《中国兽医科学》2021年第5期629-637,共9页Chinese Veterinary Science

基　　金：广西科技重大专项(桂科AA17204057);广西水产畜牧科技项目(桂渔牧科201528017);广西农业科技项目(Z201954,Z202031)。

摘　　要：为探索猪流行性腹泻病毒(PEDV)非结构蛋白nsp1对干扰素(IFN)激活的Janus激酶/信号转导和转录激活因子(JAK/STAT)信号通路的影响,将PEDV nsp1蛋白表达质粒转染HeLa细胞,并用IFN-β处理,应用Western-blot对STAT1、STAT2磷酸化以及早幼粒细胞白血病蛋白(PML)表达水平进行检测,应用IFA试验对STAT1、STAT2核移位以及PML蛋白表达水平进行检测,应用双荧光素酶分析系统对干扰素刺激反应元件(ISRE)-Luc进行检测,应用荧光定量RT-PCR(qRT-PCR)对IFN刺激基因15(ISG15)、ISG56以及PML转录水平进行检测。结果显示,nsp1蛋白过表达细胞内STAT1和STAT2的蛋白表达水平、磷酸化作用以及磷酸化STAT1(pSTAT1)、pSTAT2的核移位不受影响,而ISRE启动子的表达水平受到显著抑制,ISG15、ISG56转录水平显著下调,PML转录水平和蛋白表达水平显著降低。结果表明,PEDV nsp1蛋白可以抑制IFN-β激活的JAK/STAT信号通路。To investigate the effect of nonstructural protein 1(nsp1) of porcine epidemic diarrhea virus(PEDV) on the interferon(IFN)-activated Janus kinase/signal transducer and activator of transcription(JAK/STAT) signaling pathway,PEDV-nsp1 protein expressing plasmid was transfected into He La cells and then treated with IFN-β.The phosphorylation of STAT1 and STAT2 and the expression of promyelocytic leukemia(PML) protein were detected by Western blot;the nuclear translocation of STAT1 and STAT2 and the expression of PML protein were detected by indirect immunofluorescence assay(IFA);the expression of interferon-stimulated response element(ISRE)-Luc was analyzed by the Dual Luciferase Assay System;the expression levels of IFN-stimulated gene 15(ISG15),ISG56 and PML were detected by fluorescence quantitative RT-PCR(q RT-PCR).As a result,the expression and phosphorylation of STAT1 and STAT2 and the nuclear translocation of phosphorylated STAT1(p STAT1) and p STAT2 in cells nsp1 overexpressed were not affected.However,the IFN-β-activated expression of ISRE reporter was significantly inhibited,the transcript levels of ISG15 and ISG56 genes were significantly reduced,and the levels of PML gene transcription and protein expression were manifestly down-regulated.The results indicated that PEDV nsp1 protein can inhibit IFN-β-activated JAK/STAT signaling pathway.

关 键 词：猪流行性腹泻病毒 nsp1蛋白 Janus激酶/信号转导和转录激活因子信号通路 抑制 

分 类 号：S852.659.6[农业科学—基础兽医学]