Transcriptional regulatory mechanisms shaping HIV proviral fate

项目摘要 在过去的二十年里，生物医学研究中最令人兴奋的突破之一是发现 抗逆转录病毒疗法（ART），将活跃的艾滋病毒复制抑制到几乎检测不到的水平。尽管如此 尽管ART取得了巨大成就，但它未能治愈艾滋病毒感染，因为潜伏的储存库在含有 静息CD 4淋巴细胞和其他表达低水平病毒产物并可再活化的细胞类型 以响应免疫刺激。因此，这些潜在的储层是功能性固化的关键屏障， 这使得对新方法的需求变得更加迫切。为了实现这一重大目标，我们必须首先确定 新的，关键的宿主因子，在我们利用这些知识进行治疗之前， 临床干预。为此，本提案将研究与以下因素有关的主宿主因子（KAP 1）： 将多种生理免疫信号输入转导成前病毒转录输出，以揭示 分子机制和评估治疗潜力。我们的工作不同于以往的研究，因为 这将有助于解开影响前病毒转录和命运的三个主要特征的作用， （复杂的转录电路结构，整合景观和免疫细胞状态），以帮助设计 消除艾滋病毒的基本方法。我们假设整合景观和免疫细胞 状态通过关键控制器的功能调节感染的结果（活动的、潜伏的、重新激活的） KAP 1等宿主因子。研究这一问题的新技术和实验方法提供了 独特的机会来测试这一假设，并确定潜在的分子机制。我们特别 将研究：1）KAP 1如何与其他宿主辅因子通讯以组装正转录 复合物，以促进响应免疫刺激的潜伏期再激活开关，2）如何整合 景观和细胞状态影响KAP 1介导前病毒转录激活和克隆扩增，3） KAP 1如何识别前病毒染色质作为“支架”来偶联转录起始， 延伸，以及4）KAP 1酶功能如何帮助缓解由新的转录因子引起的抑制。 在潜伏期-再激活转换期间在原病毒处组装的辅阻遏物。知识产生于 本研究将建立干扰各种“可用药袋”功能的基础（例如， 酶和染色质阅读器结构域）用于HIV根除策略。高意义的组合， 在生理相关系统中创新和使用各种实验方法（主要 等待时间和患者样本的模型）使得该提议独一无二。总的来说，这些研究可能有一个 对我们理解HIV潜伏生物学具有革命性的影响，并具有治疗意义。

PROJECT SUMMARY Over the past two decades, one of the most exciting breakthroughs in biomedical research was the discovery of anti-retroviral therapy (ART), which curbs active HIV replication to nearly undetectable levels. Despite this great achievement, ART fails to cure HIV infection because latent reservoirs persist in tissues containing resting CD4 lymphocytes and other cell types that express low levels of viral products and can be reactivated in response to immune stimulation. Thus, these latent reservoirs are a critical barrier for a functional cure, making the need for new approaches even more urgent. To achieve this major goal, we must first identify novel, critical host factors that can be targeted therapeutically before we can leverage this knowledge for clinical intervention. To this end, this proposal will investigate a master host factor (KAP1) implicated in transducing a variety of physiologic immune signal inputs into proviral transcriptional outputs to reveal the molecular mechanisms and evaluate therapeutic potential. Our work is different from previous studies because it will help uncouple the contribution of the three major features influencing proviral transcription and fate (complex transcriptional circuit architecture, integration landscape, and immune cell state) to help devise rationale approaches for HIV eradication. We hypothesize that both the integration landscape and immune cell state regulate the outcome of an infection (active, latent, reactivated) through the function of critical master host factors like KAP1. New technologies and experimental approaches for investigating this problem provide unique opportunities to test this hypothesis and define the underlying molecular mechanisms. Specifically, we will investigate: 1) how KAP1 communicates with other host co-factors to assemble a positive transcription complex to promote the latency-reactivation switch in response to immune stimulation, 2) how the integration landscape and cell state influence KAP1-mediated proviral transcription activation and clonal expansion, 3) how KAP1 recognizes proviral chromatin to function as a ‘scaffold’ to couple transcription initiation and elongation, and 4) how KAP1 enzymatic functions help relieve the repression imparted by novel transcriptional co-repressors assembled at the provirus during the latency-reactivation switch. The knowledge generated in this study will establish the groundwork for interfering with the function of various “druggable pockets” (e.g., enzymatic and chromatin reader domains) for HIV eradication strategies. The combination of high significance, innovation and use of a diverse set of experimental approaches in physiologically relevant systems (primary models of latency and patient samples) make this proposal unique. Collectively, these studies could have a revolutionary impact on our understanding of HIV latency biology and have therapeutic implications.