Regulation of HIV Transcription Elongation

描述（由申请人提供）：由于潜伏感染细胞的稀少和难以接近，对建立和维持潜伏期的分子事件的研究受到阻碍。HIV前病毒在转录水平受到调控，并涉及转录起始、染色质组织和延伸的变化。特别地，转录延伸已被证明是HIV表达的限制步骤，并且克服该阻断是病毒因子达特的主要活性。我们假设，调节转录延长和过早转录终止的细胞因子对HIV转录有直接影响，包括消除HIV转录和建立潜伏期。初步数据显示，Pol II持续合成能力是在没有达特的情况下HIV转录的检查点，并且两个因子NELF和Pcf 11负调节HIV转录延伸。使用各种生化方法，包括染色质免疫沉淀，Pol II足迹和在体内和体外转录系统，我们建议确定的生化机制和表征的细胞因子，负调控HIV转录延长。我们希望这些研究将为HIV潜伏期的建立、维持和逆转提供新的见解。了解HIV转录延长的调节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：成功地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库，以及确定这些人群是否可以清除艾滋病毒。该提案侧重于抑制HIV转录的生化机制，以深入了解调节HIV潜伏期的因素，长期目标是确定控制不同细胞中HIV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.