Regulation of HIV Transcription Elongation

描述（由申请人提供）：由于潜伏感染细胞的稀缺性和不可接近性，对建立和维持潜伏期的分子事件的研究受到了阻碍。HIV病毒在转录水平上受到调控，涉及转录起始、染色质组织和延伸的变化。特别是，转录延伸已被证明是HIV表达的一个限制步骤，克服这一障碍是病毒因子Tat的主要活性。我们假设调节转录延伸和转录过早终止的细胞因子对HIV转录有直接影响，包括抑制HIV转录和建立潜伏期。初步数据显示，在没有Tat的情况下，Pol II的加工性是HIV转录的一个检查点，NELF和Pcf11两个因子负向调节HIV转录伸长。使用多种生化方法，包括染色质免疫沉淀，Pol II足迹和体内和体外转录系统，我们建议确定生化机制和表征负调控HIV转录伸长的细胞因子。我们期望这些研究将为HIV潜伏期的建立、维持和逆转提供新的见解。了解HIV转录延伸的调控将为控制和清除不同细胞库中的HIV提供新的细胞靶点。公共卫生相关性：成功根除艾滋病毒感染将需要了解如何建立和维持低表达病毒的细胞储存库，以及确定这些人群是否可以清除艾滋病毒。本研究的重点是抑制HIV转录的生化机制，以期深入了解调节HIV潜伏期的因素，并找到控制HIV在不同细胞中表达的新靶点。

DESCRIPTION (provided by applicant): Studies of the molecular events that establish and maintain latency have been hampered by the rarity and inaccessibility of latently infected cells. HIV provirus is regulated at the level of transcription and involves changes in transcription initiation, chromatin organization and elongation. In particular, transcription elongation has been demonstrated to be a limiting step for HIV expression and overcoming this block is the primary activity of the viral factor Tat. We hypothesize that cellular factors that regulate transcription elongation and premature transcription termination have a direct impact on HIV transcription, including extinguishing HIV transcription and establishing latency. Preliminary data show that Pol II processivity is a check point for HIV transcription in the absence of Tat and that two factors, NELF and Pcf11, negatively regulate HIV transcription elongation. Using a variety of biochemical approaches, including chromatin immunoprecipitation, Pol II foot printing and in vivo and in vitro transcription systems, we propose to determine the biochemical mechanisms and characterize the cellular factors that negatively regulate HIV transcription elongation. We expect that these studies will provide new insights into the establishment, maintenance and reversal of HIV latency. Understanding the regulation of HIV transcription elongation will provide novel cellular targets for controlling and purging HIV in different cellular reservoirs. PUBLIC HEALTH RELEVANCE: Successfully eradicating HIV infection will require understanding how cellular reservoirs that poorly express virus are established and maintained as well as determining whether these populations can be purged of HIV. This proposal focuses on the biochemical mechanisms that repress HIV transcription, in order to gain insights into factors that regulate HIV latency with long term goals of identifying novel targets for controlling HIV expression in different cells.