猪血凝性脑脊髓炎病毒核衣壳蛋白可抑制Ⅰ型干扰素产生

Nucleocapsid protein of porcine hemagglutinating encephalomyelitis virus inhibits the production of interferon-β

为明确猪血凝性脑脊髓炎病毒(PHEV)合成的病毒蛋白抑制Ⅰ型干扰素(IFN)产生的分子机制,本研究将PHEV接种RAW264.7细胞后利用放线菌酮药物处理,结果显示,随着药物对病毒蛋白合成的抑制,β-干扰素(IFN-β)的生成增多;通过构建不同的病毒蛋白真核表达质粒,转染至HEK293T细胞,经双荧光素酶报告基因系统筛选,发现病毒核衣壳(N)蛋白能抑制IFN-β启动子活性,且证明N蛋白通过干扰素调节因子3(IRF3)或IRF3上游信号分子调控IFN通路;免疫共沉淀分析进一步验证,确定线粒体抗病毒信号蛋白(MAVS)与PHEV N蛋白存在互作,为抑制IFN-β产生的潜在靶标。结果表明,PHEV可通过其自身合成的N蛋白与宿主细胞MAVS相互作用,从而抑制IFN-β产生。本研究对于解析PHEV逃避宿主免疫应答机制具有重要参考意义。

In order to clarify the molecular mechanism of which viral protein synthesized by porcine hemagglutinating encephalomyelitis virus(PHEV)inhibits the production of typeⅠinterferon,RAW264.7cells were inoculated with PHEV and treated with cycloheximide.The results showed that the production of IFN-βincreased with the inhibition of viral protein synthesis by drugs.Different eukaryotic expression plasmids of viral proteins were constructed and transfected into HEK293Tcells respectively.Through the screening of the dual luciferase reporter gene system,it was found that the viral nucleocapsid(N)protein could inhibit the activity of the IFN-βpromoter,indicating that the N protein regulated the IFN pathway through IRF3or IRF3upstream signal molecules.By co-immunoprecipitation analysis,it was further verified and confirmed that the mitochondrial antiviral signal protein(MAVS)interacted with PHEV N protein,which was the potential target for inhibiting IFN-βproduction.In summary,this study confirms that PHEV can interact with MAVS through N protein to inhibit IFN-βproduction.The research results have great significance for analyzing the mechanism of PHEV evading host immune response.