LEDGF/p7 and HIV Integration

LEDGF/p7 和 HIV 整合

• 批准号: 8128056
• 负责人: Eric M. Poeschla
• 金额: $ 3.35万
• 依托单位: MAYO CLINIC ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-05-01 至 2013-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8128056
• 关键词: Address; Basic Science; Biochemical; Biological; Biology; Catalysis; Cell Nucleus; Cells; Chemistry; Chimeric Proteins; Chromatin; Complement; Complex; Data; Employee Strikes; Environment; Evolution; Genetic; Genome; Genomics; HIV; HIV-1; Insertional Mutagenesis; Knock-out; Knowledge; Laboratories; Lentivirus Vector; Life Cycle Stages; Maps; Methods; Modeling; Molecular; Nucleic Acids; Pattern; Property; Proteins; Proteomics; RNA Interference; Reaction; Risk; Role; Site; Subfamily lentivirinae; Techniques; Tertiary Protein Structure; Testing; Therapeutic; Transcription Coactivator; Viral; Virus; base; cofactor; domain mapping; experience; genome-wide; interest; new therapeutic target; pressure; protein structure; public health relevance; research study; transcriptional coactivator p75

DESCRIPTION (provided by applicant): While the essential nucleic acid chemistry of the integration reaction is reasonably understood, how integration of the viral pre-integration complex actually happens in the complex environment of the cell nucleus remains mysterious in many respects. In particular, the integration field is intensely interested in how cellular molecules participate either positively or negatively in the reaction between the pre-integration complex and chromatin, and also in how they determine where and when the reaction occurs in the host cell genome. The transcriptional coactivator LEDGF/p75 has now been securely implicated as an integration cofactor specific for lentiviruses. A basic molecular tethering model has been put forth in which LEDGF/p75 tethers IN to chromatin, key participating protein domains have been identified, and precise structural information exists for the LEDGF/p75 domain that interacts with IN. Hypothesized roles of LEDGF/p75 include acting as a cofactor for catalysis steps, protecting IN and the pre- integration complex in which it resides from degradation, and guiding or tethering the pre-integration complex to chromatin. The latter mechanism could be crucial for the virus to engage chromatin and integrate per se, and it also may determine the distinctive genomic distributions now evident for HIV integration sites. Understanding the importance of LEDGF/p75 to HIV biology may therefore have long-term implications for understanding latency and may provide a new therapeutic target. It could even pave the way for targeting of lentiviral vectors to reduce insertional mutagenesis risks. In this project, we will use effective RNAi knockdown methods, genetic knockout cells, virological analyses, domain mapping and heterologous tethering techniques, genome- wide integration site analyses, and cell biological and proteomic methods to both further understand and exploit the viral biology of LEDGF/p75. PUBLIC HEALTH RELEVANCE This project will elucidate the significance of the transcriptional cofactor LEDGF/p75 in the HIV-1 life cycle, using virological, biochemical, genomic and chimeric protein approaches.

描述（由申请人提供）：虽然整合反应的基本核酸化学已经被合理地理解，但病毒预整合复合物的整合如何在细胞核的复杂环境中实际发生，在许多方面仍然是神秘的。特别是，整合领域对细胞分子如何积极或消极地参与预整合复合体和染色质之间的反应，以及它们如何决定反应在宿主细胞基因组中发生的位置和时间非常感兴趣。转录辅激活因子LEDGF/p75现已被确定为慢病毒特异性的整合辅因子。提出了LEDGF/p75将in与染色质拴住的基本分子拴住模型，确定了关键参与蛋白结构域，并确定了与in相互作用的LEDGF/p75结构域的精确结构信息。假设的LEDGF/p75的作用包括作为催化步骤的辅助因子，保护IN及其所在的预整合复合体免受降解，并引导或将预整合复合体连接到染色质上。后一种机制可能对病毒与染色质结合和自身整合至关重要，它也可能决定HIV整合位点的独特基因组分布。因此，了解LEDGF/p75对HIV生物学的重要性可能对理解潜伏期具有长期意义，并可能提供新的治疗靶点。它甚至可以为慢病毒载体的靶向性铺平道路，以减少插入突变的风险。在这个项目中，我们将使用有效的RNAi敲除方法，基因敲除细胞，病毒学分析，区域定位和异种系聚技术，全基因组整合位点分析，以及细胞生物学和蛋白质组学方法来进一步了解和利用LEDGF/p75的病毒生物学。本项目将利用病毒学、生化、基因组学和嵌合蛋白等方法阐明转录辅助因子LEDGF/p75在HIV-1生命周期中的意义。