NANOSCALE ARCHITECTURE OF ESCRT MACHINERY IN HIV RELEASE

HIV 释放中 ESCRT 机器的纳米级架构

• 批准号: 8993494
• 负责人: Phyllis I Hanson
• 金额: $ 7.63万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-05-01 至 2017-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8993494
• 关键词: ATP phosphohydrolase; Acquired Immunodeficiency Syndrome; Address; Affect; Architecture; Binding; Biochemical; Biogenesis; Caliber; Cell membrane; Cell physiology; Cells; Cellular Membrane; Complex; Cytokinesis; Electron Microscopy; Equine Infectious Anemia Virus; Feline Immunodeficiency Virus; Filament; Gagging; HIV; HIV-1; Health; Image; Integration Host Factors; Knowledge; Label; Libraries; Life; Mediating; Membrane; Membrane Proteins; Modeling; Molecular; Multivesicular Body; Mutagenesis; Pharmaceutical Preparations; Proteins; Public Health; Recruitment Activity; Recycling; Resolution; Retroviridae; Role; Site; Small Interfering RNA; Sorting - Cell Movement; Spottings; Staging; Structure; System; Techniques; Testing; Tissues; Uncertainty; Viral; Viral Genome; Virion; Virus; Virus Assembly; Virus Diseases; Virus Replication; Virus-like particle; Work; base; cellular imaging; electron tomography; gag Gene Products; genetic manipulation; improved; insight; light microscopy; nanoscale; novel; novel strategies; particle; prevent; protein transport; public health relevance; reconstitution; research study; trafficking

 DESCRIPTION (provided by applicant): HIV and AIDS remains a persistent problem in the US and around the world. One reason for the lack of better drugs to prevent HIV infection is insufficient understanding of how the virus is released from infected host cells. HIV and other retroviruses depend on host cell factors known as endosomal sorting complex required for trafficking (ESCRT) machinery for the critical step of membrane scission necessary for budding and release. The virally encoded Gag protein assembles on plasma membranes and produces the necessary curvature to package the viral genome. Gag also has specific motifs to bind to ESCRT-I and/or Alix, which can recruit ESCRT-III proteins. ESCRT-III proteins assemble into filaments on membranes to facilitate scission and release virus. However, the molecular mechanisms responsible for ESCRT-III recruitment and activation to release assembled virions remain unclear. In this proposal, we will use deep-etch electron microscopy (EM) in combination with correlative light microscopy to study the relationship between HIV Gag and the cellular ESCRT machinery. Deep-etch EM is a powerful technique for visualizing cellular membranes and membrane surface proteins and can provide nm resolution views of the protein machinery involved in viral particle assembly. We will apply this to (i) define the molecular architecture of connections between HIV Gag and ESCRTs stabilized by the absence of Vps4 and (ii) together with correlative light microscopy extend these studies to examine transient ESCRT structures present during normal viral particle assembly. Results from these experiments will move understanding of HIV and other retrovirus budding forward, paving the way for developing new strategies to intervene in HIV infection.

 描述（由适用提供）：艾滋病毒和艾滋病在美国和世界各地仍然是一个持久的问题。缺乏更好的药物来预防HIV感染的原因之一是对感染宿主细胞释放病毒的理解不足。 HIV和其他逆转录病毒依赖于宿主细胞因子被称为内体分选复合物（ESCRT）机械所需的膜科学的关键步骤所需的膜科学步骤。病毒编码的GAG蛋白聚集在质膜上，并产生必要的曲率包装病毒基因组。 GAG还具有与ESCRT-I和/或ALIX结合的特定基序，这些基序可以募集ESCRT-III蛋白。 ESCRT-III蛋白在膜上聚集成细丝，以促进细胞和释放病毒。但是，负责ESCRT-III募集的分子机制和释放组装病毒的激活尚不清楚。在此提案中，我们将使用深度蚀刻电子显微镜（EM）与正确的光学显微镜结合使用，以研究HIV GAG与细胞ESCRT机械之间的关系。深蚀刻是一种可视化细胞膜和膜表面蛋白质的强大技术，可以提供与病毒颗粒组装有关的蛋白质机械的NM分辨率视图。我们将将其应用于（i）定义的分子结构 HIV GAG与ESCRT之间的连接稳定在不存在VPS4和（II）的情况下，以及矫正光学显微镜扩展了这些研究，以检查正常病毒颗粒组装过程中存在的瞬时ESCRT结构。这些实验的结果将使人们对艾滋病毒和其他逆转录病毒的前进的了解，为开发新策略介绍以干预HIV感染的道路。