GENETIC ANALYSIS OF CORONAVIRUS ASSEMBLY INTERACTIONS

冠状病毒组装相互作用的遗传分析

• 批准号: 7027721
• 负责人: Paul Scott Masters
• 金额: $ 42.73万
• 依托单位: WADSWORTH CENTER
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-06-01 至 2010-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7027721
• 关键词: Coronaviridae; gene expression; genetic recombination; glycoproteins; intermolecular interaction; membrane proteins; murine hepatitis virus; nucleocapsid; phosphorylation; protein protein interaction; protein structure; protein structure function; respiratory virus; site directed mutagenesis; virion; virus RNA; virus assembly; virus envelope; virus genetics; virus protein

DESCRIPTION (provided by applicant): Coronaviruses are a family of enveloped RNA viruses that cause respiratory, enteric, and neurologic diseases in mammalian and avian hosts. In humans, three coronaviruses are responsible for upper respiratory tract infections; a fourth human coronavirus is the recently discovered causative agent of SARS. The genomes of coronaviruses are the largest among all the RNA viruses, which has made their genetic manipulation a formidable problem. Our laboratory developed the earliest reverse genetic system for coronaviruses, called targeted RNA recombination, with the prototype coronavirus mouse hepatitis virus (MHV). This method exploits the high rate of RNA recombination in MHV to transduce site-specific mutations into the viral genome by recombination with synthetic RNA introduced into infected cells. Coupled with a powerful host-range-based selection system, targeted RNA recombination has become a robust and versatile technique that has been used to answer fundamental questions about viral protein structure and function, host species specificity, virion assembly, and the complex mechanism of coronavirus RNA synthesis. The major object of this proposal is to further extend our genetic studies of coronavirus assembly mechanisms. Targeted RNA recombination and complementary biochemical analyses will be used to answer basic questions about the functional roles of MHV structural proteins in viral replication: how the small envelope protein cooperates with the membrane protein to drive formation and budding of the virus envelope; how the spike glycoprotein endodomain specifies its inclusion in virion assembly; and how the nucleocapsid protein binds to genomic RNA and is selected for incorporation into virions. An understanding of the molecular biology of coronaviruses is critical for their control and prophylaxis. The proposed studies will provide fundamental insights into the coronavirus life cycle, potential targets for antiviral chemotherapy, and a possible means to manipulate these infectious agents for vaccine design.

描述（由申请人提供）： 冠状病毒是一个包膜RNA病毒家族，可引起哺乳动物和禽类宿主的呼吸道、肠道和神经系统疾病。 在人类中，三种冠状病毒是上呼吸道感染的原因;第四种人类冠状病毒是最近发现的SARS病原体。 冠状病毒的基因组是所有RNA病毒中最大的，这使得它们的基因操纵成为一个棘手的问题。 我们的实验室开发了最早的冠状病毒反向遗传系统，称为靶向RNA重组，其原型是冠状病毒小鼠肝炎病毒（MHV）。 该方法利用MHV中RNA重组的高速率，通过与引入感染细胞的合成RNA重组，将位点特异性突变插入病毒基因组。 结合强大的基于宿主范围的选择系统，靶向RNA重组已成为一种强大且通用的技术，已用于回答有关病毒蛋白质结构和功能、宿主物种特异性、病毒体组装以及冠状病毒RNA合成的复杂机制的基本问题。 这项建议的主要目的是进一步扩展我们对冠状病毒组装机制的遗传研究。 靶向RNA重组和互补生化分析将用于回答有关MHV结构蛋白在病毒复制中的功能作用的基本问题：小包膜蛋白如何与膜蛋白合作以驱动病毒包膜的形成和出芽;刺突糖蛋白内域如何指定其在病毒体组装中的包含;以及核衣壳蛋白如何与基因组RNA结合并被选择掺入病毒体。 了解冠状病毒的分子生物学对于控制和预防至关重要。拟议的研究将提供对冠状病毒生命周期的基本见解，抗病毒化疗的潜在靶点，以及操纵这些感染因子用于疫苗设计的可能方法。