Role of Tat and SIRT1 in the T-cell Hyperactivation Syndrome Induced by HIV-1

Tat 和 SIRT1 在 HIV-1 诱导的 T 细胞过度激活综合征中的作用

• 批准号: 8206497
• 负责人: Melanie Maria Ott
• 金额: $ 47.27万
• 依托单位: J. DAVID GLADSTONE INSTITUTES
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-01-01 至 2014-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8206497
• 关键词: AIDS/HIV problem; Acquired Immunodeficiency Syndrome; Address; Aging; Antibodies; Antiviral Agents; Apoptotic; Attenuated; Binding; Blood Circulation; CD28 gene; CD3 Antigens; CD4 Positive T Lymphocytes; CD8B1 gene; Cell Culture Techniques; Cell Death; Cell Line; Cells; Chronic; Consensus; Coupled; Cytomegalovirus; Data; Deacetylase; Deacetylation; Development; Disease; Disease Progression; Engineering; Equilibrium; Flow Cytometry; Gene Expression; Generations; Genes; Green Fluorescent Proteins; HIV; HIV Infections; HIV-1; HLA-DR Antigens; Health; Helper-Inducer T-Lymphocyte; Highly Active Antiretroviral Therapy; Histone Deacetylase; Human; Immune; Immune response; Immune system; Individual; Infection; Infectious Agent; Inflammation; Inflammatory; Interleukin-1; Interleukin-2; Interleukin-6; Interphase Cell; Investigation; Laboratories; Lentivirus Vector; Link; Longevity; Measures; Mediating; Microbe; Modeling; Molecular; Molecular Biology; Molecular Profiling; Nature; Nicotinamide adenine dinucleotide; Outcome; Participant; Patients; Peripheral; Phenotype; Plasma; Play; Population; Process; Production; Proteins; Regulation; Regulatory T-Lymphocyte; Reporting; Research; Role; SIV; Shapes; Signal Transduction; Subfamily lentivirinae; Surface; Symptoms; Syndrome; T-Cell Activation; T-Lymphocyte; TNF gene; Testing; Therapeutic; Trans-Activators; Viral; Virus Diseases; cytokine; design; genetic regulatory protein; genome-wide; immune activation; immune function; insight; interest; lymph nodes; microbial; novel; novel strategies; novel therapeutic intervention; p300/CBP-Associated Factor; p65; programs; receptor expression; research study; response; small hairpin RNA; small molecule; tat Protein; transcription factor

DESCRIPTION (provided by applicant): Symptoms of T-cell hyperactivation shape the course and outcome of HIV-1 infection, but the molecular mechanism(s) underlying this chronic immune activation are not well understood. We have identified a novel mechanism by which the HIV-1 Tat protein leads to chronic immune activation. We find that Tat hyperactivates T cells by blocking the deacetylase activity of SIRT1 (Kwon et al, Cell Host Microbe, 2008). SIRT1 is a nicotinamide adenine dinucleotide (NAD+)-dependent histone deacetylase and an important regulator of the transcription factor NF-B. Tat directly interacts with the deacetylase domain of SIRT1 and blocks the ability of SIRT1 to deacetylate the RelA/p65 subunit of NF-B. Because acetylated p65 is more active as a transcription factor, Tat hyperactivates the expression of NF-B-responsive genes, such as interleukin-2 (IL-2). These results support a model where the normal function of SIRT1 as a negative regulator of T-cell activation is suppressed by Tat during HIV infection. We propose to define the novel role of SIRT1 as a regulator of immune activation in HIV-infected T cells. We have new and unpublished findings that link the SIRT1 deacetylase activity with immune-suppressive functions of regulatory T cells (Tregs). Special focus of this proposal lies therefore on the characterization of the function of Tat and SIRT1 in Tregs and on the identification of novel SIRT1 substrates in effector and regulatory T cells populations. The importance of these studies is highlighted by their possible impact on the development of novel therapeutic interventions. Potent small molecule activators of SIRT1 have recently become available with great promise in diseases associated with aging (Milne et al, Nature, 2007). We will use these activators to test the hypothesis that activating the suppressor function of SIRT1 will counterbalance the immune stimulatory function of Tat and may represent a novel strategy to treat chronic immune activation in HIV-infected patients. Our specific aims are: 1) we will study the role of SIRT1 in HIV-infected T cells. We will infect Jurkat T cell lines in which SIRT1 expression is down regulated via shRNAs with a GFP-tagged infectious clone of HIV-1 and study intracellular IL-2 production in infected (GFP+) and uninfected (GFP-) cells in response to activation with anti-CD3 and CD28 antibodies. We will also infect primary CD4+ T cell cultures with HIV-1 and test whether treatment with SIRT1 activators can reverse hyperstimulation of IL-2 production. 2) We will identify novel targets of the SIRT1 deacetylase activity in infected T cells. We have preliminary evidence that Tat by inhibiting SIRT1 induces hyperacetylation of several SIRT1 targets including proapoptotic factors p53 and Foxo3A. Since both factors are important regulators of T-cell death during HIV infection, we will study how Tat manipulates the transcriptional activities and proapoptotic functions of both factors. We will also perform genome-wide expression profiling and identify gene programs that are modulated by Tat expression or SIRT1 knockdown. 3) We will examine how the SIRT1/Tat interaction influences the development and function of Tregs. We have preliminary results showing that SIRT1 promotes TGF--induced Treg development and is involved in deacetylation of the transcription factor FoxP3. We propose to study how SIRT1 regulates expression and function of FoxP3 in Tregs and will characterize how Tat expression during HIV infection can interfere with this process. These studies will bring novel insight into the molecular biology of HIV-induced immune activation and identify yet undefined mechanisms of HDAC-mediated control of the immune response. PUBLIC HEALTH RELEVANCE: We seek to identify and characterize novel regulatory mechanisms controlling T cell hyperactivation during HIV-1 infection. T cell hyperactivation is a hallmark of pathogenic HIV-1 infection and the strongest predictor of the progression to AIDS in infected individuals. Our proposed studies characterize novel mechanisms by which the HIV-1 Tat protein hyperactivates T cells. These studies are directly relevant to HIV/AIDS and may contribute to the development of novel antiviral drugs that will address public need.

描述（由申请人提供）：T细胞过度激活的症状塑造了HIV-1感染的过程和结果，但是这种慢性免疫激活的分子机制尚不清楚。我们已经确定了一种新型机制，HIV-1 TAT蛋白会导致慢性免疫激活。我们发现TAT通过阻断SIRT1的脱乙酰基酶活性来过度激活T细胞（Kwon等，细胞宿主Microbe，2008）。 SIRT1是烟酰胺腺苷二核苷酸（NAD+） - 依赖性组蛋白脱乙酰基酶，也是转录因子NF-B的重要调节剂。 TAT直接与SIRT1的脱乙酰基酶结构域相互作用，并阻止SIRT1脱乙酰基NF-B的RELA/P65亚基的能力。由于乙酰化p65作为转录因子更为活跃，因此TAT过度激活NF-B反应性基因的表达，例如白介素-2（IL-2）。这些结果支持一个模型，其中SIRT1作为T细胞激活的负调节剂的正常功能在HIV感染过程中被TAT抑制。我们建议定义SIRT1作为HIV感染T细胞中免疫激活的调节剂的新作用。我们有新的和未发表的发现，可以将SIRT1脱乙酰基酶活性与调节性T细胞的免疫抑制功能联系起来。因此，该提案的特殊重点在于TAT和SIRT1在Tregs中的功能以及效应子和调节性T细胞种群中新型SIRT1底物的识别的表征。这些研究的重要性是由于它们对新型治疗干预措施发展的可能影响所强调。 SIRT1的有效的小分子激活剂最近在与衰老有关的疾病方面具有巨大的希望（Milne等，Nature，2007）。我们将使用这些激活剂来检验以下假设：激活SIRT1的抑制剂功能将抵消TAT的免疫刺激功能，并可能代表一种治疗HIV感染患者慢性免疫激活的新策略。我们的具体目的是：1）我们将研究SIRT1在HIV感染的T细胞中的作用。我们将感染Jurkat T细胞系，其中SIRT1表达通过shRNA下降，并通过GFP标记的HIV-1的感染性克隆来调节HIV-1的感染性克隆，并研究受感染（GFP+）的细胞内IL-2产生和未感染的（GFP-）细胞，响应于抗CD3和CD28抗体而响应激活而响应激活。我们还将用HIV-1感染原发性CD4+ T细胞培养物，并测试用SIRT1激活剂处理是否可以逆转IL-2产生的过度刺激。 2）我们将确定感染T细胞中SIRT1脱乙酰基酶活性的新靶标。我们有初步的证据表明，通过抑制SIRT1诱导多个SIRT1靶标的高乙酰化，包括凋亡因子p53和FoxO3a。由于这两个因素都是HIV感染期间T细胞死亡的重要调节剂，因此我们将研究TAT如何操纵这两个因素的转录活性和促凋亡功能。我们还将执行全基因组表达分析，并识别由TAT表达或SIRT1敲低调节的基因程序。 3）我们将研究SIRT1/TAT相互作用如何影响Treg的发展和功能。我们的初步结果表明，SIRT1促进了TGF诱导的Treg发育，并参与转录因子Foxp3的脱乙酰化。我们建议研究SIRT1如何调节FOXP3在Tregs中的表达和功能，并将表征HIV感染期间TAT表达如何干扰这一过程。这些研究将带来对HIV诱导的免疫激活的分子生物学的新见解，并确定HDAC介导的免疫反应控制的但不确定的机制。公共卫生相关性：我们试图识别和表征控制HIV-1感染期间T细胞过度激活的新型调节机制。 T细胞过度激活是致病性HIV-1感染的标志，也是感染个体辅助的最强预测指标。我们提出的研究表征了HIV-1 TAT蛋白过度激活T细胞的新型机制。这些研究与艾滋病毒/艾滋病直接相关，可能有助于开发新的抗病毒药物，以满足公众需求。