DUT通过靶向RIG-I正调控RLR介导的抗病毒信号通路

DUT positively regulates RLR-mediated antiviral signaling pathway by targeting RIG-I

目的探究DUT(deoxyuridine triphosphate nucleotidohydrolase, dUTPase)对RIG-I(retinoic acid-inducible gene I)介导的RLR(RIG-I-like receptors)抗病毒先天免疫的研究。方法采用免疫共沉淀的方法检测DUT与RIG-I之间的相互作用以及DUT对RIG-I介导的泛素化修饰的影响;应用非变性聚丙烯酰胺凝胶电泳实验验证DUT对RIG-I或仙台病毒(Sendai virus,SeV)诱导的IRF3二聚化的作用;利用双荧光素酶报告基因实验检测RIG-I或SeV介导的DUT对干扰素启动子IFN-β的激活效果;通过实时荧光定量PCR实验检测DUT对SeV诱导的IFN-β转录的影响。结果 DUT与RIG-I相互作用,过表达DUT促进RIG-I或SeV诱导的IRF3的二聚化水平以及IFN-β启动子的活性。此外,过表达DUT也加强了SeV诱导的IFN-β启动子的转录水平,研究结果还表明DUT促进RIG-I泛素化修饰及RIG-I K63泛素化修饰,并且通过与RIG-I之间的相互作用增强RNF135、MEX3C、TRIM4介导的RIG-I K63泛素化修饰。结论 DUT是RIG-I介导的针对RNA病毒天然免疫应答的正调节因子。

The purpose of this study is to investigate the effect of deoxyuridine triphosphate nucleotidohydrolase(DUT) on RIG-I-mediated RLR antiviral innate immunity. Immunoprecipitation was used to detect the interaction between DUT and RIG-I, and the effect of DUT on ubiquitination of RIG-I. The effect of DUT on RIG-I/SeV-induced IRF3 dimerization was verified by nature PAGE. The effect of DUT on SeV-induced IFN-βtranscription was detected by qPCR. Data showed that DUT interacted with RIG-I;DUT overexpression promoted the dimerization of IRF3 induced by RIG-I/SeV and the activity of IFN-β promoter. In addition, DUT enhanced the transcription of IFN-β induced by SeV. Furthermore, DUT promoted ubiquitination of RIG-I and RIG-I K63,enhanced RNF135/MEX3C/TRIM4-mediated RIG-I K63 ubiquitination through interaction with RIG-I. Thus, DUT is a positive regulator for RIG-I-mediated innate immune response to RNA virus.