猪瘟病毒拮抗干扰素诱导蛋白OASL抗病毒作用的分子机制

Interferons (IFNs) and IFN-stimulated genes (ISGs) play key roles in the host antiviral defense. Recently, we have demonstrated that the porcine 2'-5'-oligoadenylate synthase-like protein (pOASL) is an antiviral ISG against classical swine fever virus (CSFV), an economically important pestivirus of pigs. We have also shown that pOASL is a coactivator of the MDA5-mediated IFN signaling pathway to exert anti-CSFV activity. Intriguingly, the expression level and antiviral activity of pOASL increased and then declined following CSFV infection. Thus, we hypothesize that CSFV may antagonize the antiviral effects of pOASL by regulating its expression. In this proposal, we will investigate the effects on the IRF3 or IFN-mediated pOASL-induction pathway after CSFV infection. Secondly, we will screen the CSFV protein(s) affecting the expression of pOASL, and its effects on the mRNA transcription or protein expression of pOASL will be examined by real-time RT-PCR and Western blotting. Finally, we will elucidate the molecular mechanism of the CSFV protein(s) antagonizing the pOASL expression through affecting the stability of IRF3 or regulating the IFN signaling pathway. This study is expected to reveal the molecular mechanism of CSFV antagonizing the antiviral activity of pOASL, which will facilitate the development of anti-CSFV agents based on pOASL and provide new insights for the control of CSFV infections.

在宿主抗病毒应答中，干扰素诱导蛋白发挥着关键作用。此前我们证实，猪源2’-5’寡腺苷酸合成酶样蛋白（pOASL）是一种具有抗猪瘟病毒活性的干扰素诱导蛋白，通过增强MDA5介导的I型干扰素信号通路而发挥抗病毒效应。同时我们发现，随着猪瘟病毒感染时间的延长，pOASL的抗病毒活性随同其表达量呈先上升、后下降的趋势。据此我们推测，猪瘟病毒可能通过调控pOASL的表达以拮抗其抗病毒作用。本项目首先确定猪瘟病毒感染对干扰素和干扰素调控因子3介导的pOASL产生途径的影响；然后，筛选调控pOASL表达的猪瘟病毒蛋白，并通过定量RT-PCR和免疫印迹试验研究其对pOASL的mRNA转录或蛋白表达的影响；最后，根据猪瘟病毒蛋白对干扰素调控因子3的稳定性和干扰素信号通路的调控，阐明其拮抗pOASL表达的机制。本研究可望揭示猪瘟病毒拮抗pOASL抗病毒作用的分子机制，为猪瘟防控和开发抗病毒制剂提供参考。

在宿主抗病毒应答中，干扰素诱导蛋白发挥着关键作用。此前我们证实，猪源2’-5’寡腺苷酸合成酶样蛋白（pOASL）是一种具有抗猪瘟病毒（CSFV）活性的干扰素诱导蛋白。本研究证实了pOASL通过增强MDA5介导的I型干扰素信号通路而发挥抗病毒效应。同时发现，随着CSFV感染时间的延长，pOASL的抗病毒活性随同其表达量呈先上升、后下降的趋势。此外，利用CRISPR/Cas9基因编辑平台成功构建了敲除IRF3和IFNAR2的PK-15细胞系，检测CSFV感染后干扰素和IRF3诱导pOASL产生的途径；最后，我们证实CSFV编码蛋白Npro与pOASL存在相互作用。本研究可望揭示CSFV拮抗pOASL抗病毒作用的分子机制，为猪瘟防控和开发抗病毒制剂提供参考。