Studies of HIV and Host Transcriptional Control by Tat

描述(申请人提供)：病毒已经进化出复杂的机制来劫持细胞资源以进行复制和传播。转录调控对HIV来说是必不可少的 转录它的基因组并应对持续感染。HIV的转录程序依赖于Tat，Tat通过与新生病毒RNA(TAR)上的细胞转录延伸因子P-TEFb组装来激活转录起始和延伸之间的转换。TAT从启动子组装的7SK SnRNP复合体中捕获P-TEFb以激活P-TEFb激酶。然而，对P-TEFb从7SK SnRNP释放和与TAR结合时激活P-TEFb的步骤的详细剖析仍未完成。除了控制病毒转录，TAT似乎还调节宿主转录，有利于感染的建立，并影响免疫系统的生物学。然而，TAT的基因靶点和使用的分子机制在很大程度上仍然不清楚。这项研究计划将使用跨学科的方法来阐明TAT从不活跃的储存库中捕获P-TEFb以激活病毒转录的机制，并定义TAT如何利用细胞复合体来施加建立感染所需的广泛的基因表达变化。该项目将在德克萨斯大学西南医学中心进行，该中心拥有最先进的技术，在免疫学和计算生物学等病毒学以外的领域拥有非常丰富的合作者氛围。这项拟议的工作在很大程度上建立在初步数据的基础上，这是一套生化、遗传学和病毒学分析，导致鉴定了TAT向7SK SnRNP招募的n酶，以释放P-TEFb并激活病毒转录。此外，一套基因组方法导致了直接的TAT靶基因的识别，并定义了TAT如何在全基因组范围内改变TAT辅助因子的占有率和染色质状态图。这些初步数据以及一种计算方法揭示了受共同机制调控的基因类别，并揭示了细胞蛋白质复合体和染色质修饰酶是重新编程过程的调节器。本研究项目将：(1)进一步明确TAT从7SK SnRNP释放P-TEFb的机制；(2)阐明TAT如何靶向细胞复合体以重新编程宿主转录；(3)确定选定的TAT靶基因在艾滋病发病机制中的作用。总而言之，这些发现将更好地阐明TAT激活的机制，并揭示TAT如何操纵生物相关的细胞基因来改变T细胞过程。

DESCRIPTION (provided by applicant): Viruses have evolved intricate mechanisms for hijacking cellular resources to replicate and spread. Transcription regulation is essential for HIV to transcribe its genome and for persistent infection. The transcriptional program of HIV relies on Tat, which functions by assembling with the cellular transcription elongation factor P-TEFb on nascent viral RNAs (TAR) to activate the transition between transcription initiation and elongation. Tat captures P-TEFb from the promoter-assembled 7SK snRNP complex to activate the P-TEFb kinase. However, detailed dissection of the steps of P-TEFb release from 7SK snRNP and activation of the P-TEFb kinase when bound to TAR, is still incomplete. In addition to controlling viral transcription, Tat appears to modulate host transcription to favor the establishment of infection and affect the biology of immune system. However, the gene targets of Tat and the molecular mechanisms used remain largely unclear. This Research Plan will use interdisciplinary approaches to elucidate the mechanism by which Tat captures P-TEFb from the inactive reservoir to activate viral transcription as well as define how Tat exploits cellular complexes to impose widespread gene expression changes required for the establishment of infection. The project will be conducted at UT Southwestern Medical Center, which has state-of-the-art technologies and a very rich atmosphere of collaborators in fields other than virology such as immunology and computational biology. The work proposed builds heavily on preliminary data, a set of biochemical, genetic, virology assays that led to the identification of n enzyme recruited by Tat to 7SK snRNP to release P-TEFb and activate viral transcription. In addition, a set of genomic approaches led to the identification of direct Tat target genes, and defined how Tat alters the occupancy of Tat cofactors and the chromatin-state maps in a genome-wide scale. This preliminary data, along with a computational approach, revealed gene classes regulated by common mechanisms and revealed cellular protein complexes and chromatin-modifying enzymes as modulators of the reprogramming process. This research project will: (1) further define the mechanism of P-TEFb release from 7SK snRNP by Tat, (2) delineate how Tat targets cellular complexes to reprogram host transcription, and (3) define the role of selected Tat target genes in the pathogenesis of AIDS. Collectively, these discoveries will better elucidate the mechanism of Tat activation as well as reveal how Tat manipulates biologically relevant cellular genes to alter T cell processes.