Investigating virus-host interactions with prime editing and genetic code expansion

通过初等编辑和遗传密码扩展研究病毒与宿主的相互作用

• 批准号: 10286563
• 负责人: Michael D Vahey
• 金额: $ 23.63万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-05-21 至 2023-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10286563
• 关键词: Amber; Amino Acids; Antiviral Therapy; Biochemical; Biological Assay; Biology; COVID-19 pandemic; CRISPR library; Cell Line; Cells; Characteristics; Chemicals; Code; Data Set; Dependence; Development; Epidemic; Exhibits; Family; Fluorescence; Fruit; Genetic Code; Genome; Genome engineering; Goals; Human; Image; Infection; Influenza; Influenza A virus; Integration Host Factors; Intervention; Knock-in; Label; Laboratories; Libraries; Machine Learning; Measures; Mediating; Medical; Methodology; Methods; Modification; Molecular; Morphogenesis; Morphology; Mutation; Nature; Pathway interactions; Play; Proteins; Research; Resources; Role; Site; Terminator Codon; Therapeutic; Therapeutic Intervention; Transfer RNA; Viral; Viral Proteins; Virion; Virus; Virus Diseases; Virus Replication; airway epithelium; base; combat; genome editing; influenzavirus; insight; mortality; novel; pandemic disease; prevent; protein function; respiratory virus; screening; stable cell line; tool; unnatural amino acids; virus host interaction

PROJECT SUMMARY Host proteins play essential roles in the replication of all viruses. This represents a vulnerability that could potentially be exploited to develop interventions against viruses currently circulating in humans as well as new and emerging viral threats. However, realizing this potential requires mechanistic understanding of how specific host factors function during viral infection. This proposal will develop a versatile screening methodology to investigate the mechanisms through which host proteins enhance virus replication. This will be achieved using recent developments in genome engineering and chemical biology that enable site-specific modification of host factors with unnatural amino acids. By genetically re-coding >100 host proteins implicated in the replication of influenza A virus (IAV), we will investigate (1) the effects of depleting specific host factors on virus replication and the subcellular localization of viral proteins; (2) the effects of viral infection on the expression and subcellular localization of host proteins; and (3) the effects of host factor depletion on the assembly and morphogenesis of new virus particles. If successful, this project will provide insight into the host dependency of IAV and establish a resource for investigating host protein function in the replication of additional, evolutionarily-divergent respiratory viruses.

项目摘要 宿主蛋白在所有病毒的复制中起着重要作用。这代表了一个漏洞， 可能被用来开发针对目前在人类中传播的病毒以及新的 和新出现的病毒威胁。然而，实现这一潜力需要机械地理解如何具体 宿主因子在病毒感染期间起作用。该提案将开发一种通用的筛选方法， 研究宿主蛋白增强病毒复制的机制。这将通过使用 基因组工程和化学生物学的最新进展， 含有非天然氨基酸的因子。通过基因重新编码涉及复制的>100种宿主蛋白质， 甲型流感病毒（IAV），我们将研究（1）消耗特定宿主因子对病毒复制的影响 病毒感染对蛋白表达和亚细胞定位的影响 宿主蛋白的定位;（3）宿主因子缺失对宿主蛋白组装和形态发生的影响。 新的病毒粒子如果成功，该项目将深入了解IAV的宿主依赖性，并建立 一个资源，用于调查宿主蛋白质的功能，在复制额外的，进化上不同的 呼吸道病毒