In vivo characterization of RNA virus assembly lines with EM tomography

使用 EM 断层扫描对 RNA 病毒装配线进行体内表征

• 批准号: 8468182
• 负责人: Jason Kenneth Lanman
• 金额: $ 20.42万
• 依托单位: PURDUE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-08-25 至 2014-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8468182
• 关键词: Address; Biochemical Genetics; Capsid; Cells; Complex; Cytoskeleton; Development; Direct Costs; Electron Microscopy; Environment; Excision; Facilities and Administrative Costs; Future; Genome; Goals; Housing; Image; In Situ; Influenza; Instruction; Location; Microscopy; Mitochondria; Molecular; Positioning Attribute; Process; RNA; RNA Viruses; Research Personnel; Resolution; Societies; System; Three-Dimensional Imaging; Time; Tomogram; United States; Viral; Virion; Virus; Virus Assembly; cost; human disease; in vivo; influenzavirus; novel; pandemic disease; reconstruction; tomography; trafficking; viral RNA

Virus assembly occurs in the complex three-dimensional environment of the cell, and new advances in electron microscopy tomography (EMT) now make it possible to construct high-resolution three-dimensional images where the spatial location of components involved in virus assembly can be determined. The simplicity of the Flock House virus (FHV) capsid and the extensive structural, biochemical, and genetic characterization of the virus make it an excellent system for studying in situ virus assembly. The focus of this proposal is to use to EMT to answer key questions about Flock House virus assembly in the cell and then extend these approaches to Influenza virus genome packaging and assembly. Aim 1: Three-dimensional imaging of the Flock House virus factories in situ. Our hypothesis is FHV assembles in viral factories, which are formed from extensively modified mitochondria and are then trafficked on cytoskeleton to viral arrays. The viral factories will be imaged using EMT to understand the virus assembly process around the modified mitochondria and entire infected cells will reconstructed to investigate the extensive morphological changes that occur during the virus lifecycle. Aim 2: EM-tomography analysis of influenza genome packaging. The Influenza virus genome is divided into eight RNA segments, and a long-standing question has been what is the mechanism for packaging all eight segments. Our hypothesis is that each of the eight viral RNAs (vRNA) are specifically packaged, and the removal of one vRNA will produce virus particles containing seven segments instead of eight. Virus particles missing a vRNA will be analyzed with EMT to determine the number of segments and which segments are packaged in each virus particle.

病毒组装发生在复杂的三维细胞环境中，并在这方面取得了新的进展