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)项目总结/摘要 潜伏的人类免疫缺陷病毒（HIV）在休眠的受感染细胞池中持续存在（称为“病毒库”） 不受联合抗逆转录病毒疗法（cART）影响的患者。难以触及的解剖间隔 治疗药物，因为大脑和肠道是病毒的避难所， 感染.在病毒学抑制的个体中，这些储库中的潜伏前病毒变得随机化 在cART停止后活跃，重新接种感染。HIV间接感染长寿脑细胞 损害神经元持续性脑感染的作用导致一系列称为HIV相关的综合征， 神经认知障碍（HAND）。cART降低了严重HAND的发生率，如HIV相关的 痴呆症;然而，轻度形式的手仍然是普遍的，具有不成比例的发病率， 种族/族裔少数群体和相当大的社会经济影响。受感染者释放病毒性神经毒素 cART药物本身引起的氧化应激是两个重要因素 手里所有上述情况都支持有效抑制艾滋病毒感染的必要性。禁用集成 使用基因编辑的前病毒，虽然有希望，但仍然是一个有风险的主张，并且有几个主要问题， 例如逃逸突变的产生，这必须在将基因编辑转移到临床之前解决。 使用.感染细胞的根除/灭菌可能对CNS敏感/药物敏感患者产生不良后果。 保护环境。通过诱导深潜伏期（“阻断和锁定”）策略使储库失活是 更适合大脑，因为它们不需要激活，杀死和清除受感染的脑细胞 然而，所有这些策略都需要对前病毒活性进行一定程度的控制，从而解释了 对了解病毒潜伏机制有相当大的兴趣。一个反馈机制， 转录的反式激活因子（达特）蛋白的表达调节生产性原病毒转录。达特 低于或高于阈值的细胞水平导致原病毒进入或退出潜伏期。药理学 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 NQO 1稳定病毒所需的达特（蛋白质稳态）的细胞水平。NQO 1还稳定细胞 许多蛋白质的水平对维持细胞健康和科普病毒感染至关重要。我们 目的是研究这一特征不明确的蛋白质抑制机制，以促进其靶向。我们 建议研究：（1）所涉及的物理复合物，它们的缔合热力学和结构， 使用生物物理方法纯化蛋白质;和（2）修饰的NQO 1活性对病毒潜伏期的影响， 细胞外达特或cART对神经元的损伤，以及对氧化和解折叠的脱靶效应， 蛋白质应激反应通过破坏达特蛋白质稳态来控制前病毒转录具有潜在的应用价值。 在病毒的根除或终身灭活中， 储库，以及减少达特在CNS中的有害作用的神经保护疗法。