Molecular Interactions of HIV-1 with the Nuclear Pore Complex

HIV-1 与核孔复合物的分子相互作用

• 批准号: 10241258
• 负责人: Gregory B Melikian
• 金额: $ 136.92万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-01 至 2023-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10241258
• 关键词: Address; Adopted; Antiviral Therapy; Architecture; Binding; Biological; Biological Assay; Biology; Biotinylation; Caliber; Capsid; Capsid Proteins; Cell Nucleus; Cells; Chemicals; Complex; Computer Models; Cone; Cryo-electron tomography; Cryoelectron Microscopy; DNA; Data; Development; Dimerization; Docking; Electron Microscopy; Engineering; Event; Evolution; Factor V; Fluorescence; Fullerenes; Future; Gap Junctions; Gene Expression; Genome; HIV; HIV-1; Image; Infection; Integration Host Factors; Interphase Cell; Ions; Karyopherins; Knowledge; Label; Length; Light; Literature; Mediating; Methods; Molecular; Molecular Structure; Monitor; Nuclear; Nuclear Envelope; Nuclear Import; Nuclear Pore; Nuclear Pore Complex; Nuclear Pore Complex Proteins; Pathway interactions; Penetration; Peroxidases; Phytic Acid; Process; Proteins; Proteomics; Protocols documentation; Reporting; Research; Reverse Transcription; Structure; System; Techniques; Viral; Viral Genes; Virus; Virus Integration; Virus Replication; Visualization; analog; ascorbate; base; crosslink; design; experimental study; imaging approach; innovation; insight; integration site; knock-down; macromolecule; multidisciplinary; new therapeutic target; novel; novel strategies; novel virus; nucleocytoplasmic transport; particle; passive transport; pleiotropism; prevent; viral DNA

HIV-1 enters the nucleus of non-dividing cells where the reverse transcribed viral DNA is integrated into the host genome. Whereas the nuclear pore complex (NPC) prevents passive transport of large macromolecules, HIV-1 has evolved effective strategies to penetrate this barrier. The HIV-1 nuclear import is a poorly understood process that involves complex interactions with the nuclear import machinery, including several nucleoporins, transportin-3, and CPSF6, all of which bind the viral capsid core, which comprises hundreds of copies of the capsid protein (CA). Pleiotropic effects caused by nucleoporin knockdown and the ability of HIV-1 to use alternative import pathways have impeded the mechanistic studies of nuclear import and frustrated efforts to identify the full array of host factors involved in this process. Our single particle tracking experiments revealed that HIV-1 infection progresses through CA-dependent docking at the nuclear membrane, followed by uncoating (loss of CA) and CA-dependent nuclear transport to the sites of integration. However, very little is known regarding the molecular details and dynamics of virus-NPC interactions, including the structural changes in the architecture of both HIV-1 and the NPC in the course of nuclear import. There is thus an unmet need for structural and functional studies on the molecular mechanisms of HIV-1 nuclear import in the context of productive infection. We propose to combine cutting-edge approaches developed by our highly collaborative team to delineate the molecular interactions and structural changes in the virus and NPC during the nuclear import. Specifically, we will: (1) identify the host factors involved in HIV-1 import using novel proteomics and chemical cross-linking approaches; (2) obtain cryo-electron tomography (cryo-ET) and cross-linking mass-spec structures of the HIV-1 core/NPC complexes by capturing the incoming virus through our new on-demand pore clogging assay; and (3) develop correlative fluorescence/cryo-ET imaging pipeline to structurally characterize the intermediates of HIV-1 nuclear import. Knowledge of molecular interactions during the HIV-1 nuclear import should help identify novel therapeutic targets to block infection, provide a framework for studies of the nuclear import of other viruses and further fundamental understanding of the nuclear pore function.

HIV-1进入非分裂细胞的细胞核，那里有逆转录的病毒DNA