冷诱导RNA结合蛋白通过激活NF-κB信号通路影响H1N1甲型流感病毒的复制  

Activation of Cold-inducible RNA-binding Protein by H1N1 Influenza Virus Contributes to Viral Replication Via Activating NF-κB Pathway

作　　者：聂培婷[1] 汤承[1] 岳华[1] 

机构地区：[1]西南民族大学生命科学与技术学院,成都610041

出　　处：《畜牧兽医学报》2016年第10期2108-2114,共7页ACTA VETERINARIA ET ZOOTECHNICA SINICA

基　　金：国家自然科学基金项目(31172307);四川省教育厅创新团队项目(13TD0057)

摘　　要：冷诱导RNA结合蛋白(CIRP)是NF-κB和ERK信号通路的上游调控因子,而这两条信号通路是甲型流感病毒复制和机体起始免疫所必需的,为探讨CIRP对H1N1甲型流感病毒复制的影响及可能的分子机制,构建了CIRP过表达BHK-21细胞系(Cirp+BHK-21),用Western blot检测NF-кB和ERK1/2的磷酸化水平,研究CIRP对NF-кB和ERK1/2的调节作用;用Real-time RT-PCR检测H1N1甲型流感病毒感染后Cirp+BHK-21和对照细胞中病毒拷贝数的动态变化,以及在特异性阻断剂PDTC阻断NF-кB通路的Cirp+BHK-21细胞中病毒拷贝数的动态变化。Western blot检测结果显示:过表达CIRP显著促进了BHK-21细胞中NF-κB的磷酸化水平(P<0.05),而对ERK1/2的磷酸化水平无显著影响;病毒定量检测结果显示:过表达CIRP能显著促进H1N1甲型流感病毒的增殖,感染后3、9、15、21h病毒在Cirp+BHK-21细胞中的拷贝数分别为对照组的111%、103%、167%和235%(P<0.05);阻断NF-κB信号通路后病毒的拷贝数显著下降,在感染后3、9、15、21h分别为未阻断组的98%、42%、19%(P<0.05)和7%(P<0.05)。从本研究结果可见,CIRP可通过活化NF-κB信号通路促进H1N1甲型流感病毒的复制。The cold-inducible RNA-binding protein（CIRP）is an upstream regulator of the NF-κB and ERK pathways,which are essential for the replication of the influenza virus and the initial immune response.In order to investigate the effect of CIRP on the replication of H1N1 influenza A virus and its possible molecular mechanism,in this study,CIRP overexpression BHK-21（Cirp＋BHK-21）cells were constructed,and the phosphorylation levels of NF-κB and ERK in Cirp＋BHK-21 cells were detected by Western blot to confirm the effect of CIRP on regulation of NF-κB and ERK1/2;Real-time RT-PCR was used to detect the dynamic changes of virus load in Cirp＋BHK-21 and control cells after infected with influenza A virus,the method was also used to detect the dynamic changes of virus load in Cirp＋BHK-21 cells which were blocked by the NF-κB inhibitor PDTC.The results of Western blot exhibited that overexpressed CIRP could significantly increase the expression of phosphorylation level of NF-κB（P〈0.05）,but had no significant effect on the phosphorylation level of ERK.The results of quantitative detection of virus showed that overexpressed CIRP could significantly enhance the proliferation of influenza A virus,the virusload in the Cirp＋BHK-21 cells were 111%,103%,167% and 235%（P〈0.05）at 3,9,15 and 21h PI,respectively,compared to the control group;Blocking the NF-κB was significantly decreased the virus load in the Cirp＋BHK-21,and the virus load in treatment group were 98%,42%,19%（P〈0.05）,7%（P〈0.05）at 3,9,15 and 21hPI,respectively,compared to the unblock group.Therefore,this study confirmed that overexpressed CIRP could enhance the proliferation of influenza A virus via activation of NF-κB pathway.

关 键 词：冷诱导RNA结合蛋白 H1N1甲型流感病毒 病毒复制 NF-ΚB BHK-21细胞 

分 类 号：S852.23[农业科学—基础兽医学]