Molecular mech/gene expression reprogramming by HIV TAT

描述（由申请人提供）：本提案的目标是深入了解HIV-1达特如何介导其对宿主细胞基因表达的影响。达特是HIV-1感染过程中最先表达的基因之一，对病毒基因表达和病毒产生至关重要。达特通过作为延伸因子并与TAR（HIV病毒转录物起始处存在的RNA序列）以及宿主细胞因子CDK 9和细胞周期蛋白T1相互作用来增加HIV-1基因表达。达特与这些和其他宿主细胞转录调节因子的相互作用可能会影响宿主细胞基因表达。事实上，有相当多的证据表明，达特可以影响T淋巴细胞，神经元和抗原呈递细胞的生理学。我们实验室最近的研究表明，树突状细胞和巨噬细胞的基因表达程序被病原体暴露所修饰，这些修饰可以为发病机制提供重要线索。我们已经证明，HIV-1对树突状细胞的重编程可以创造有利于病毒传播的条件，并且这种作用是由HIV-1反式激活因子达特介导的。该建议旨在提高我们对达特表达引起HIV-1靶向免疫细胞中宿主细胞基因表达变化的方式的理解。为了实现这一点，该提议的具体目标是1）开发表达多种野生型和突变的达特蛋白的腺病毒和逆转录病毒载体的文库。2)确定达特的两个野生型等位基因对HIV-1感染靶向免疫细胞中宿主细胞基因表达的影响。3)定位负责修饰宿主细胞基因表达程序的达特结构域。4)研究转录因子STAT 1和IRF 7在达特表达过程中的基因组规模定位，因为这些因子被HIV-1感染和达特表达上调。5)研究达特通过基因组规模的定位分析实验检测DNA和RNA结合来重编程宿主细胞基因表达的机制。

DESCRIPTION (provided by applicant): The goal of this proposal is to gain insights into how HIV-1 Tat mediates its effects on host cell gene expression. Tat is among the first genes expressed during HIV-1 infection and is essential for viral gene expression and virus production. Tat increases HIV-1 gene expression by functioning as an elongation factor and interacting with TAR, a RNA sequence present at the beginning of the HIV viral transcripts, and with the host cell factors CDK9 and cyclin T1. The interaction of Tat with these and other host cell transcriptional regulators might be expected to affect host cell gene expression. Indeed, there is considerable evidence that Tat can affect the physiology of T lymphocytes, neurons and antigen presenting cells. Recent studies from our laboratory have shown that the gene expression programs of dendritic cells and macrophages are modified by pathogen exposure and that these modifications can provide important clues to pathogenesis. We have shown that dendritic cell reprogramming by HIV-1 can create conditions that favor virus spread, and that the effect is mediated by the HIV-1 transactivator Tat. This proposal is designed to improve our understanding of the means by which Tat expression causes changes in host cell gene expression in immune cells targeted by HIV-1. To accomplish this, the specific aims of the proposal are 1) To develop a library of adenovirus and retroviral vectors expressing multiple wild type and mutated Tat proteins. 2) To determine the effects of two wild type alleles of Tat on host cell gene expression in immune cells targeted by HIV-1 infection. 3) To map the domain of Tat that is responsible for modifying the host cell gene expression program. 4) To investigate the genome-scale location of the transcription factors STAT1 and IRF7 during Tat expression, as these factors are upregulated by HIV-1 infection and Tat expression. 5) To investigate the mechanism by which Tat reprograms host cell gene expression using genome-scale location analysis experiments designed to detect DNA and RNA binding.