HIV-1 Intasome Assembly and Function

HIV-1 整合体的组装和功能

基本信息

批准号：
10794478
负责人：
KRISTINE E YODER
金额：
$ 15.73万
依托单位：
OHIO STATE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-12-15 至 2023-08-24
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10794478
关键词：
Acquired Immunodeficiency Syndrome Address Affect Architecture Binding Biological Biology Biophysics Capsid Cell Nucleus Chimera organism Chromatin Chromosomes Clinical Combination Drug Therapy Complex DNA DNA Repair Disease Drug resistance Exhibits Family member Genome Genomics Goals HIV HIV Genome HIV-1 HIV-1 integrase Histones Human Human Genome Imaging technology In Vitro Individual Infection Integrase Integration Host Factors Kinetics Knowledge Length Lentivirus Lesion Long Terminal Repeats Mechanics Mouse Mammary Tumor Virus Mutation Nucleosomes Outcome Pathogenicity Patients Pattern Peptides Positioning Attribute Post-Translational Protein Processing Process Productivity Proteins Reporting Retroviridae Retroviridae Infections Retrovirology Reverse Transcription Role Side Site Spumavirus Structure Time Treatment Protocols Viral Viral Reverse Transcription Virion Virus animation biophysical chemistry cellular transduction cofactor design dimer feature detection flexibility genomic RNA imaging platform in vitro activity in vivo innovation integration site leukemia meter molecular imaging new therapeutic target pandemic disease prototype single molecule stoichiometry therapeutic target transcriptional coactivator p75 viral DNA viral resistance

项目摘要

HIV-1 INTASOME ASSEMBLY AND FUNCTIONS PROJECT SUMMARY / ABSTRACT New HIV-1 retrovirus infections continue to drive a worldwide pandemic. Combined drug therapies have helped to blunt the clinical outcomes that afflict HIV-1 infected individuals. However, drug-resistance virus mutations that challenge these treatment regimens persist, making identification of new drug targets of crucial importance. HIV-1 integration into the human genome is essential for a productive infection. Integration is catalyzed by the retrovirus encoded integrase (IN), which forms a multimeric complex with the long terminal repeat (LTR) ends of the reverse-transcribed viral cDNA (termed an intasome and/or pre-integration complex). Structural comparisons show that all seven retrovirus genera maintain a conserved intasome core (CIC) configuration, which precisely positions the LTR-ends for catalytic strand-transfer into a genomic target site during integration. Different retrovirus family members expand the size of the CIC by appending additional IN subunits. For example, the prototype foamy virus (PFV) intasome assembles as a simple tetramer while the mouse mammary tumor virus (MMTV) forms an octamer by attaching IN dimers to either side of the CIC. IN octamer, decamer, dodecamer (12-mer) and hexadecamer (16-mer) intasomes have been reported for HIV-1. The contributions of IN-multimer architecture to HIV-1 biology and biophysical chemistry is unknown. The assembly processes that ultimately result in an HIV-1 intasome remain enigmatic. Accumulating evidence suggests that the viral capsid containing the HIV-1 genome is imported into the nucleus where assembly of the intasome occurs in concert with capsid disassembly. Integration into the genomic chromatin then occurs 1-2 µm from the capsid disassembly site. HIV-1 integration is facilitated by host factors that include LEDGF/p75. We and others have found that LEDGF/p75 is required for efficient HIV-1 intasome assembly in vitro. These observations underpin several key unanswered questions: What are the progressions that result in an assembled HIV-1 intasome? What is the role of LEDGF/p75 in intasome assembly? What is the impact of IN- multimer architecture on intasome stability and genomic target site selection in vitro and in vivo? We propose to utilize innovative real-time single molecule imaging and analysis to understand the contributions of IN-multimer architecture on HIV-1 mechanics with the following Specific Aims: 1.) determine the IN-subunit assembly progressions that control multimeric HIV-1 intasome architecture, 2.) determine the role of HIV-1 intasome architecture on the dynamic interactions with defined chromatin target DNA in vitro, and 3.) determine the role of HIV-1 intasome architecture on targeting host chromatin features in cellulo. These studies are designed to interrogate the animated processes that support HIV-1 intasome architecture with the goal of identifying additional retroviral progressions that may be exploited as therapeutic targets.

HIV-1 内体组装和功能项目概要/摘要新的 HIV-1 逆转录病毒感染继续推动全球大流行，联合药物疗法对此有所帮助。削弱HIV-1感染者的临床结果，然而，耐药病毒突变。这些挑战仍然存在，因此确定新的药物靶点至关重要。 HIV-1 整合到人类基因组中对于有效感染至关重要。整合是由 HIV-1 催化的。逆转录病毒编码的整合酶（IN），与长末端重复（LTR）末端形成多聚体复合物逆转录病毒 cDNA（称为整合体和/或预整合复合体）。比较表明所有七个逆转录病毒属都保持保守的嵌体核心（CIC）构型，它精确定位 LTR 末端，以便在整合过程中将催化链转移到基因组靶位点。不同的逆转录病毒家族成员通过附加额外的 IN 亚基来扩大 CIC 的大小。例如，原型泡沫病毒（PFV）嵌体组装成一个简单的四聚体，而小鼠乳腺病毒则组装成一个简单的四聚体。肿瘤病毒 (MMTV) 通过将 IN 二聚体连接到 IN 八聚体、十聚体、据报道，十二聚体（12-mer）和十六聚体（16-mer）嵌体对 HIV-1 的贡献。 IN-多聚体结构对 HIV-1 生物学和生物物理化学的影响尚不清楚。最终产生 HIV-1 嵌体的组装过程仍然是个谜。表明含有 HIV-1 基因组的病毒衣壳被输入细胞核，在那里组装整合体与衣壳解体同时发生，然后整合到基因组染色质中 1-2 µm。包括 LEDGF/p75 在内的宿主因子促进了 HIV-1 的整合。等人发现 LEDGF/p75 是 HIV-1 嵌体体外高效组装所必需的。观察结果支持了几个关键的未解答的问题：是什么进展导致了 LEDGF/p75在intasome组装中的作用是什么？多聚体结构对体内外嵌体稳定性和基因组靶位点选择的影响？我们建议利用创新的实时单分子成像和分析来了解其贡献 HIV-1 机制上 IN 多聚体结构的研究，具体目标如下：1.) 确定 IN 亚基控制多聚体 HIV-1 嵌体结构的组装进程，2.) 决定 HIV-1 的作用体外与特定染色质靶 DNA 动态相互作用的内体结构，以及 3.) 确定 HIV-1 嵌体结构在细胞中靶向宿主染色质特征中的作用。这些研究旨在探究支持 HIV-1 嵌体结构的动画过程目的是确定可用作治疗靶点的其他逆转录病毒进展。