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（由申请人提供）：艾滋病毒和艾滋病仍然是美国和世界各地的一个持续问题。缺乏更好的药物来预防艾滋病毒感染的一个原因是对病毒如何从受感染的宿主细胞中释放的了解不足。HIV和其他逆转录病毒依赖于宿主细胞因子，称为运输所需的内体分选复合物（ESCRT）机制，用于出芽和释放所需的膜断裂的关键步骤。病毒编码的Gag蛋白在质膜上组装，并产生包装病毒基因组所需的曲率。Gag还具有与ESCRT-1和/或阿利克斯结合的特异性基序，其可以募集ESCRT-III蛋白。ESCRT-III蛋白在膜上组装成细丝，以促进切割和释放病毒。然而，负责ESCRT-III募集和激活以释放组装的病毒体的分子机制仍不清楚。在这个建议中，我们将使用深蚀刻电子显微镜（EM）结合相关的光学显微镜来研究HIV Gag和细胞ESCRT机制之间的关系。深蚀刻EM是一种用于可视化细胞膜和膜表面蛋白的强大技术，并且可以提供参与病毒颗粒组装的蛋白质机制的nm分辨率视图。我们将应用这一点（i）定义分子结构 HIV Gag和ESCRT之间的连接稳定的Vps 4和（ii）与相关的光学显微镜一起扩展这些研究，以检查正常病毒颗粒组装过程中存在的瞬时ESCRT结构。这些实验的结果将推动对艾滋病毒和其他逆转录病毒萌芽的理解，为开发干预艾滋病毒感染的新策略铺平道路。