CHEETAH Center for the Structural Biology of HIV Infection, Restriction, and Viral Dynamics

CHEETAH HIV 感染、限制和病毒动力学结构生物学中心

• 批准号: 10663363
• 负责人: Pamela J Bjorkman
• 金额: $ 170.74万
• 依托单位: UNIVERSITY OF UTAH
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-07-11 至 2027-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10663363
• 关键词: 3-Dimensional; Architecture; Binding; Biological Assay; Biology of HIV Infection; Capsid; Cell Communication; Cell Nucleus; Cell membrane; Cell-Free System; Cells; Chromatin; Complementary DNA; Complex; Coupled; DNA; DNA Integration; Development; Docking; Electrons; Exposure to; Fluorescence Microscopy; Foundations; Genome; HIV-1; Host Factor 1 Protein; Image; In Situ; Infection; Integration Host Factors; Knowledge; Learning; Life Cycle Stages; Macaca; Methods; Molecular; Molecular Conformation; Nuclear; Nuclear Import; Nuclear Pore; Nucleosome Core Particle; Pathway interactions; Plastic Embedding; Process; Productivity; Proviruses; RNA; Replication Initiation; Resolution; Reverse Transcription; Role; Series; Stains; Structure; System; Systems Integration; Time; Tissue Sample; Transcription Initiation; Ubiquitin; Viral; Viral Reverse Transcription; Virus; Virus Integration; Visualization; design; dimer; electron tomography; experimental study; genomic RNA; in vivo; inhibitor; novel; receptor; reconstitution; simian human immunodeficiency virus; structural biology; transcriptional coactivator p75; viral DNA; viral RNA; viral genomics

PROJECT SUMMARY To initiate an infection, HIV-1 must enter the cell, reverse transcribe the viral RNA into DNA, enter the nucleus, and integrate into host cell chromatin. Studies in Project 1, Infecting the Cell, will focus on defining these key steps in the first half of the HIV-1 life cycle in molecular and mechanistic detail. A particular theme will be to elucidate the organization, functional roles, and dynamic transformations of the viral core particle. Studies in Aim 1 (Virus Entry) will build on our previous structural studies of different states of HIV-1 Env and imaging of viral entry complexes in situ to provide structural details of the virus-target cell interaction at the time of entry including: 1) the number of Envs and average spacing in a typical interaction with a target cell, 2) whether there is rearrangement of the Env subunits that are not bound to host receptors, and 3) the structure of the hypothesized pre-hairpin intermediate formed between the virus and host cell membranes by Env and host receptor(s) at sufficient resolution to dock coordinates from components of known structure. Studies in Aim 2 (Genome Structure and Reverse Transcription Initiation) will build on our previous structures of HIV-1 replication initiation complexes to reveal: 1) the structural basis for the transition from initiation to elongation, 2) how the dimeric structure of the viral RNA modulates reverse transcription initiation, 3) how the global dynamic architecture of HIV-1 genomic RNA is influenced by the capsid, and 4) how this dynamic architecture influences the process of reverse transcription, including strand jumps. Studies in Aim 3 (Capsid Functions in Infection) will build on our development of a cell-free system that efficiently reconstitutes the coupled processes of viral endogenous reverse transcription and integration to: 1) characterize the viral genomic template at different stages of reverse transcription, 2) reveal how the capsid facilitates reverse transcription, 3) define how the potent new capsid inhibitor, lenacapavir, binds the capsid and inhibits nuclear entry, and 4) reconstitute and image core passage through nuclear pores. Studies in Aim 4 (Core Uncoating and Integration) will build on our cell-free integration assay and our ability to reconstruct viral capsid lattices at high resolution to fill a series of fundamental knowledge gaps, including: 1) the identities and functional roles of host factors that promote uncoating and integration, 2) how the viral capsid “opens” to expose the integrating viral DNA, and 3) the 3D architectures of active, native intasomes.

项目摘要 为了引发感染，HIV-1必须进入细胞，将病毒RNA转换为DNA，进入细胞核，然后 集成到宿主细胞染色质中。项目1的研究感染了细胞，将重点放在第一个中定义这些关键步骤 HIV-1生命周期的一半，分子和机械细节。一个特定的主题将是阐明组织， 功能作用和病毒核心粒子的动态变换。 AIM 1（病毒进入）中的研究将基于我们先前对HIV-1 Env的状态的结构研究和成像的成像 原位病毒进入复合物在进入时提供病毒靶向细胞相互作用的结构细节，包括： 1）在与目标单元的典型互动中的ENV数量和平均间距，2）是否有重排 不属于主机接收器的ENV亚基，以及3）假设的前中级的结构 由Env和宿主受体以足够的分辨率坐在坐标的病毒和宿主细胞膜之间形成 来自已知结构的组成部分。 AIM 2的研究（基因组结构和逆转录开始）将基于我们先前的HIV-1结构 复制倡议络合物要揭示：1）从主动性到伸长的过渡的结构基础，2） 病毒RNA的二聚体结构调节逆转录启动，3）全局动态体系结构如何 HIV-1基因组RNA受CAPSID的影响，4）这种动态结构如何影响反向的过程 转录，包括链跳。 AIM 3中的研究（感染中的Capsid功能）将建立在我们对无细胞系统的开发上，该系统有效地 重构病毒内源性逆转录和整合的耦合过程：1）表征病毒 逆转录不同阶段的基因组模板，2）揭示衣壳栖息地如何逆转录，3） 定义潜在的新的CAPSID抑制剂Lenacapavir如何结合衣壳并抑制核进入，以及4）重新构造和 图像核心密码通过核孔。 AIM 4的研究（核心不涂层和集成）将基于我们的无细胞集成测定法和我们重建的能力 高分辨率的病毒式衣壳晶格以填补一系列基本知识差距，包括：1）身份和身份 宿主因素的功能作用，促进脱落和整合的宿主因素，2）病毒capsid如何揭示病毒式衣壳 整合病毒DNA和3）活性天然静态的3D架构。