Characterization of the redox control of HIV-Tat proteostasis by cellular NQO1

细胞 NQO1 对 HIV-Tat 蛋白质稳态氧化还原控制的表征

• 批准号: 9695502
• 负责人: Mario A. Bianchet
• 金额: $ 24.56万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-09-15 至 2020-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9695502
• 关键词: AIDS Dementia Complex; Anatomy; Binding; Brain; Calorimetry; Cell model; Cells; Characteristics; Clinical; Complex; Development; Dinucleoside Phosphates; Dose; Down-Regulation; Drug-sensitive; Environment; Enzymes; Equilibrium; Feedback; Flavins; Generations; Genes; Genetic Transcription; Global Change; Goals; HIV; HIV Infections; HIV-associated neurocognitive disorder; Handedness; Heat shock proteins; Histones; Homeostasis; Incidence; Individual; Infection; Intake; Maintenance; Measures; Molecular Conformation; Mutation; NQO1 gene; Neurons; Neurotoxins; Outcome; Oxidation-Reduction; Oxidative Stress; Personal Satisfaction; Pharmaceutical Preparations; Pharmacology; Process; Proteins; Proviruses; Rattus; Research; Source; Sterilization; Stress; Structure; Syndrome; Therapeutic; Therapeutic Intervention; Thermodynamics; Titrations; Toxic effect; Trans-Activators; Transcription Coactivator; Viral; Viral Physiology; Viral Proteins; Viral Regulatory Proteins; Viral reservoir; Virus; Virus Diseases; Virus Inactivation; Virus Latency; Virus Replication; X-Ray Crystallography; Xenobiotics; adverse outcome; antiretroviral therapy; biological adaptation to stress; biophysical techniques; brain cell; cofactor; ethnic minority population; extracellular; genetic regulatory protein; inhibitor/antagonist; interest; multicatalytic endopeptidase complex; neurotoxicity; protein protein interaction; proteostasis; racial and ethnic; response; rev Protein; socioeconomics; tat Protein; uptake; virology

1) Project Summary/Abstract Latent human immunodeficiency virus (HIV) persists in pools of dormant infected cells (called “viral reservoirs”) that are unaffected by combined antiretroviral therapies (cARTs). Anatomic compartments hard to reach with therapeutic drugs as the brain and gut are viral sanctuaries harboring reservoirs or maintaining low-grade infections. In virologically suppressed individuals, latent proviruses in those reservoirs become stochastically active upon the discontinuation of cART, reseeding the infection. HIV infects long-lived brain cells indirectly damaging neurons. The action of a persistent brain infection causes a set of syndromes called HIV associated neurocognitive disorders (HAND). cART decreased the incidence of severe HAND such as HIV-associated dementia; however milder forms of HAND still are prevalent, having a disproportionate incidence on racial/ethnic minorities and a considerable socioeconomic impact. Viral neurotoxins to be released by infected cells despite cART and the oxidative stress caused by the cART drugs themselves are both significant factors in HAND. All the above support the need for efficient suppression of the HIV infection. Disable integrated proviruses using gene editing, although promising, is still a risky proposition and has several major issues, such as the generation of escape mutations, which must be resolved before moving gene-editing to clinical use. Eradication/sterilization of the infected cells could have adverse outcomes in the CNS sensitive/drug- protected environment. The reservoirs inactivation by inducing deep-latency (“block and lock”) strategies are more suitable for the brain because they do not require activate, killing, and clearing the infected brain cells Nevertheless, all of these strategies require a level of control on the provirus activity explaining the considerable interest in understanding viral latency mechanisms. A feedback mechanism driven by the expression of the trans-activator of transcription (Tat) protein regulates productive provirus transcription. Tat cellular levels under or over a threshold result in provirus entry or exit from latency. A pharmacologically exploitable feedback mechanism, involving the viral regulatory protein Rev and a host cytoprotective enzyme NQO1, stabilizes the cellular level of Tat (proteostasis) required for the virus. NQO1 also stabilizes cellular levels of many proteins essential for the cell well-being maintenance and to cope with the viral infection. Our objective is to investigate this poorly characterized proteostasis mechanism to facilitate its targeting. We are proposing to investigate: (1) the physical complexes involved, their association thermodynamics, and structure, using biophysical methods with purified proteins; and (2) the effect of modify NQO1 activity on viral latency, on the damage to neurons produced by extracellular Tat or cART, and off-target effects on oxidative- and unfolde protein stress responses. Controlling proviral transcription by disrupting Tat proteostasis has the potential to be a significantly synergistic part of viral activity manipulation in the eradication or lifelong inactivation of viral reservoirs, as well as of neuroprotective therapies that decrease the detrimental effects of Tat in the CNS.

1）项目摘要/摘要 潜在的人类免疫缺陷病毒（HIV）持续存在于休眠感染细胞的池中（称为“病毒储层”） 不受组合抗逆转录病毒疗法（CART）的影响。解剖区室很难与 作为大脑和肠道的治疗药物是藏有储量或保持低级的病毒庇护所 感染。在病毒学抑制的个体中，这些水库中的潜在病毒随机变为 在停用购物车时活跃起来，恢复感染。 HIV感染长寿命的脑细胞间接感染 破坏性神经元。持续性大脑感染的作用引起一组称为HIV相关的综合征 神经认知障碍（手）。卡车减少了严重的手的事件，例如与HIV相关的 失智;但是，米勒的手形式仍然很普遍，对 种族/族裔少数群体和考虑的社会经济影响。病毒神经毒素将被感染释放 细胞dospite Cart和由CART药物本身引起的氧化应激都是重要的因素 手头。以上所有这些都支持有效抑制HIV感染的需求。禁用集成 使用基因编辑的病理虽然很有希望，但仍然是一个冒险的主张，并且有几个主要问题， 例如，逃生突变的产生，必须在将基因编辑到临床之前就必须解决 使用。受感染细胞的根除/灭菌可能在中枢神经系统敏感/药物中会产生不利的结果 受保护的环境。诱发深层（“块和锁定”）策略灭活的水库灭活是 更适合大脑，因为它们不需要激活，杀死和清除感染的脑细胞 然而，所有这些策略都需要对Provirus活动的一定程度的控制，以解释 对理解病毒潜伏机制的极大兴趣。由 转录（TAT）蛋白的反式激活剂的表达可调节生产性病毒转录。 tat 阈值下方或超出阈值下的蜂窝水平会导致进入或退出潜伏期。一家药品 可利用的反馈机制，涉及病毒调节蛋白Rev和宿主细胞保护酶 NQO1稳定了病毒所需的TAT（蛋白质量）的细胞水平。 NQO1还稳定了细胞 许多蛋白质的水平对于细胞健康维持和应对病毒感染所必需的水平。我们的 目的是研究这种特征性较差的蛋白质病机制以促进其靶向。我们是 提议调查：（1）涉及的物理络合物，它们的关联热力学和结构， 使用纯化蛋白质的生物物理方法； （2）修饰NQO1活性对病毒潜伏期的影响， 细胞外TAT或手推车产生的神经元的损害，以及靶向氧化和展开的靶向影响 蛋白质应激反应。通过破坏TAT蛋白质量来控制临时转录的潜力 病毒活性操纵的显着协同部分在消除或终身失活的病毒中 储层以及神经保护疗法，可减少TAT在中枢神经系统中的有害作用。