Analysis of Coronavirus-Host Cell Interactions

描述（由申请人提供）：病毒基因组的翻译在冠状病毒（CoV）感染后立即开始，产生两个大的多聚蛋白，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参与病毒RNA的合成，有些还具有其他生物学功能。包括SARS-CoV（SCoV）、蝙蝠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CoV 229 E在内的alphaCoV的nsp 1蛋白抑制宿主基因表达。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，CoV的nsp 1蛋白很可能在CoV的致病中起关键作用。与这一观点一致，MHV nsp 1是一个主要的毒力因子，而SCoV nsp 1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Nsp 1蛋白具有共同的生物学功能，抑制宿主基因表达，但使用不同的策略来发挥这一功能。SCoV nsp 1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 nsp 1与40 S核糖体亚基结合以抑制mRNA翻译，并且还诱导模板依赖性核酸内切mRNA切割。与SCoV nsp 1相反，TGEV nsp 1采用不同的策略来抑制mRNA翻译，因为它不能结合40 S核糖体亚基或促进宿主mRNA降解。本申请旨在通过使用SCoV和TGEV nsp 1蛋白作为模型系统来描绘CoV nsp 1诱导的基因表达抑制的新机制。我们将揭示SCoV nsp 1诱导的模板依赖性mRNA切割的机制。我们还将阐明SCoV和TGEV nsp 1诱导的翻译抑制的不同机制，并揭示在nsp 1诱导的翻译抑制条件下，允许在SCoV感染的细胞中稳健的病毒基因表达的策略。这些研究将为理解CoV对宿主基因表达的调控奠定基础，在分子水平上扩展我们对CoV致病性的认识，并可能确定真核基因表达调控的新机制。 公共卫生相关性：拟议的研究将描述冠状病毒nsp 1诱导的基因表达抑制的新机制。这些研究将为理解冠状病毒对宿主基因表达的调节提供基础，扩大我们对冠状病毒致病性的认识，并可能确定真核基因表达调控的新机制。

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. PUBLIC HEALTH RELEVANCE: The proposed studies will delineate the novel mechanisms of coronavirus nsp1-induced inhibition of gene expression. These studies will provide a foundation for understanding the modulation of host gene expression by coronaviruses, expand our knowledge of coronavirus pathogenicity, and potentially identify a novel mechanism for the regulation of eukaryotic gene expression.