Analysis of Coronavirus-Host Cell Interactions

描述(申请人提供)：病毒基因组的翻译在冠状病毒(CoV)感染后立即开始，产生两个大的多蛋白，并加工成15或16个成熟的非结构蛋白。这些蛋白质中的大多数都参与了病毒RNA的合成，有些还具有其他生物学功能。包括SARS冠状病毒(SCOV)、蝙蝠冠状病毒(BAT CoV)和小鼠肝炎病毒(MHV)在内的BetaCov和包括传染性胃肠炎病毒(TGEV)和人类CoV229E在内的AlphaCov的nsp1蛋白抑制宿主基因的表达。由于过去的研究表明，抑制宿主基因表达的病毒蛋白是主要的毒力因子，冠状病毒的nsp1蛋白很可能在冠状病毒的致病过程中发挥关键作用。与这一观点一致的是，MHV NSP1是一个主要的毒力因子，而SCOV NSP1抑制感染细胞中I型干扰素和干扰素刺激基因的产生。不同冠状病毒的Nsp1蛋白具有共同的抑制宿主基因表达的生物学功能，但发挥这一功能的策略不同。SCOV nsp1使用一种新的双管齐下的策略来抑制宿主蛋白质合成/基因表达。SCOV NSP1结合到40S核糖体亚基上，抑制mRNA翻译，并诱导模板依赖的内切核糖核酸。与SCOV nsp1不同，TGEV nsp1采用不同的策略来抑制mRNA翻译，因为它不能结合40S核糖体亚基或促进宿主mRNA的降解。本应用旨在以CoV和TGEV NSP1蛋白为模型系统，阐明CoV nsp1诱导基因表达抑制的新机制。我们将揭示SCOV NSP1诱导的模板依赖的mRNA切割的机制。我们还将阐明SCOV和TGEV NSP1诱导的翻译抑制的不同机制，并揭示在NSP1诱导的翻译抑制条件下允许病毒基因在SCOV感染细胞中强劲表达的策略。这些研究将为理解冠状病毒对宿主基因表达的调控提供基础，在分子水平上扩展我们对冠状病毒致病性的认识，并有可能发现一种新的调控真核基因表达的机制。

DESCRIPTION (provided by applicant): The translation of the viral genome begins immediately after coronavirus (CoV) infection to produce two large polyproteins that are processed into 15 or 16 mature, nonstructural proteins. Most of these proteins are involved in viral RNA synthesis, and some have other biological functions. The nsp1 protein of betaCoVs, which includes SARS-CoV (SCoV), bat CoVs, and mouse hepatitis virus (MHV), and of alphaCoVs, which includes transmissible gastroenteritis virus (TGEV) and human CoV 229E, inhibits host gene expression. As past studies have shown that viral proteins that inhibit host gene expression are major virulence factors, nsp1 proteins of CoVs most probably play a critical role in CoV pathogenesis. Consistent with this notion, MHV nsp1 is a major virulence factor, and SCoV nsp1 inhibits the production of type I interferon and interferon-stimulated genes in infected cells. Nsp1 proteins of different CoVs share a common biological function to inhibit host gene expression but use different strategies to exert this function. SCoV nsp1 uses a novel, two-pronged strategy to inhibit host protein synthesis/gene expression. SCoV nsp1 binds to the 40S ribosomal subunit to inhibit mRNA translation and also induces a template-dependent endonucleolytic mRNA cleavage. In contrast to SCoV nsp1, TGEV nsp1 employs a different strategy to inhibit mRNA translation, as it is unable to bind the 40S ribosomal subunit or promote host mRNA degradation. This application aims to delineate the novel mechanisms of CoV nsp1-induced inhibition of gene expression by using SCoV and TGEV nsp1 proteins as model systems. We will uncover the mechanism of SCoV nsp1-induced template-dependent mRNA cleavage. We will also clarify the different mechanisms of SCoV and TGEV nsp1-induced translation inhibition and reveal the strategy that allows robust viral gene expression in SCoV-infected cells under conditions of nsp1-induced translation inhibition. The proposed studies will provide a foundation for understanding the modulation of host gene expression by CoV, expand our knowledge of CoV pathogenicity at the molecular level, and potentially identify a novel mechanism for the regulation of eukaryotic gene expression.