甲型肝炎病毒利用细胞的miR-149-3p拮抗β干扰素产生的分子机制

It has been confirmed that hepatitis A virus (HAV) completely inhibits production of the antiviral interferon β (IFN-β) and establishes persistent infection state when it proliferates in cultured cells in vitro. As we known, miRNAs involve in interactions between host cells and viruses, but whether miRNAs involve in the processes of HAV antagonizing the IFN-β induction is unclear. In the preliminary studies, we have confirmed that HAV antagonized the IFN-β production by up-regulating the expression of the cellular miR-149-3p. Furthermore, target gene analysis showed that the TRAF3 and TRIM14, which are adaptor molecules in the IFN-β induction signaling pathway, are potential targets of miR-149-3p. Therefore, we hypothesized that HAV makes use of cellular miR-149-3p to antagonize the IFN-β production by targeting the TRAF3 and TRIM14. As well as, the potential target TRIM14 in IFN-β antagonism process has not been reported. Here, we intend to reveal the signal transduction pathways and mechanisms of regulation of HAV induced miR-149-3p, and we will also clarify the molecular mechanisms of miR-149-3p antagonising the IFN-β production by RNAi, ChIP etc. technology. The study will reveal a new mechanism that the picornavirus HAV escape from the host antiviral innate immunity by cellular miR-149-3p, and provide new ideas for prevention of the related RNA viruses.

甲肝病毒（HAV）在体外增殖时完全抑制了细胞抗病毒IFN-β产生并建立起持续性感染，已知miRNA介导了病毒与细胞的互作，但miRNA是否介导了HAV对IFN-β诱导的拮抗尚不明确。前期我们已经证实：HAV通过上调细胞的miR-149-3p拮抗IFN-β产生。靶基因分析显示，IFN-β诱生信号通路中的接头分子TRAF3和TRIM14是miR-149-3p的潜在靶点。因此，我们推测：HAV利用细胞的miR-149-3p靶向TRAF3和TRIM14以拮抗IFN-β产生。并且，以TRIM14为靶点的IFN-β拮抗目前未见报道。我们拟用RNAi、ChIP等技术观察HAV诱导miR-149-3p上调的信号转导通路和机制，并观察miR-149-3p拮抗IFN-β产生的分子机制。该研究将揭示一种小RNA病毒HAV利用宿主miR-149-3p逃逸细胞抗病毒固有免疫的新机制，为相关RNA病毒防治提供新思路。

甲型肝炎病毒（HAV）在体外细胞培养系统中形成持续性感染，细胞用于对抗病毒感染的β干扰素被病毒抑制，使得HAV逃逸了β干扰素的抗病毒作用，其确切机制尚未完全阐明。本课题从miRNA切入，从新的视角探讨了HAV拮抗β干扰素产生的分子机制。本课题首先利用RNA-seq和二代测序，发现了HAV感染KMB-17细胞后大量差异表达调控的miRNAs分子，进一步通过生物信息学分析和实验验证相结合，筛选了调控IFN-β表达的miRNA分子miR-149-3p和miR-146a-5p。进一步，在HAV和KMB-17细胞系统中，利用RNAi技术、Real-time PCR、转录谱芯片等方法相结合发现HAV感染诱导细胞miR-149-3p和miR-146a-5p转录上调依赖于部分转录因子的活性改变，同时在分子水平明确了miR-149-3p和miR-146a-5p通过靶向其相应的靶基因TRAF3和TRAF6而拮抗IFN-β的表达，进而调控HAV的基因组水平，维持病毒的持续性感染状态。本研究揭示了一种HAV实现逃逸细胞抗病毒免疫反应的新机制，对认识HAV和宿主细胞的相互作用具有重要意义。