Mechanisms underlying lncRNA SAF-mediated survival and persistence of HIV-1 infected macrophages

lncRNA SAF介导的HIV-1感染巨噬细胞存活和持久性的机制

• 批准号: 10374126
• 负责人: Saikat Boliar
• 金额: $ 19.63万
• 依托单位: CORNELL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-03-17 至 2024-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10374126
• 关键词: Alveolar Macrophages; Antisense RNA; Apoptosis; Apoptotic; Biological; Biology; CASP3 gene; CD4 Positive T Lymphocytes; CD95 Antigens; Caspase; Cell Death; Cell Line; Cell Maintenance; Cell Survival; Cells; Cessation of life; Characteristics; Code; Comprehension; DNA; Data; Defective Viruses; Disease Progression; Event; Genetic; Genetic Transcription; Goals; HIV-1; Hela Cells; High-Throughput RNA Sequencing; Human; In Vitro; Infection; Integration Host Factors; Knowledge; Lung; Mediating; Mediator of activation protein; Modality; Molecular; Molecular Target; Myelogenous; Myeloid Cells; Nonlytic; Pathogenesis; Pathway interactions; Population; Proteins; Proteomics; RNA; Receptor Signaling; Regulation; Reporting; Reverse Transcription; Role; Small Interfering RNA; T-Lymphocyte; Therapeutic; Tissues; Untranslated RNA; Up-Regulation; Viral; Viral Load result; Viral Pathogenesis; Viral reservoir; Viremia; Virus; Virus Inhibitors; Virus Replication; antiretroviral therapy; base; comparative; experimental study; improved; in vivo; latent infection; loss of function; macrophage; monocyte; mortality; mutant; novel; novel therapeutics; overexpression; prevent; receptor expression; screening; success; therapeutic candidate; therapeutic development; tool; transcriptome sequencing

Project Summary / Abstract Combination anti-retroviral therapy is highly effective in blocking HIV-1 replication and thereby new infections but it cannot eliminate reservoir cells infected prior to the treatment initiation. Most studies of HIV-1 persistence have focused on latently infected CD4+ T cells. However, tissue-resident macrophages which are self-renewable, long-lived myeloid cells, have recently been shown to sustain prolonged viremia in absence of T lymphocytes. The unique ability of macrophages to support HIV-1 replication without succumbing to virus- induced cell death makes them ideal for viral persistence. Understanding the role of viral and host factors that enable macrophages to evade cell death is needed to develop effective strategies for eliminating these persistently infected myeloid cells. By screening a panel of 90 long non-coding RNAs (lncRNA), we have recently discovered the novel role of the lncRNA SAF (FAS-AS1) in maintenance of cell survival of HIV-1 infected macrophages. This lncRNA is significantly up-regulated in HIV-1 infected human monocyte-derived macrophages (MDM) in vitro and lung alveolar macrophages (AM) in vivo. More importantly, siRNA-mediated inhibition of SAF leads to activation of apoptotic caspases in HIV-1 infected macrophages selectively, while leaving the virus-exposed yet uninfected bystander cells unaffected. This highly specific modulation of the lncRNA SAF results in a marked reduction in viral burden in the macrophage culture, emphasizing the therapeutic potential of targeting this lncRNA. Our central hypothesis is that active HIV-1 infection induces SAF up-regulation in macrophages to regulate cell survival/death pathway(s) that promote viral persistence. We propose an in-depth analysis of both the upstream regulators of SAF transcription and the downstream effectors for its function. Our specific aims are to: (1) define the viral and cellular regulators for induction of lncRNA SAF in HIV-1 infected macrophages; and (2) determine the molecular mechanisms of lncRNA SAF mediated protection of HIV-1 infected macrophages from cell death by identifying the DNA/RNA and/or protein targets of this lncRNA. We will use a panel of specific replication-stage-defective viruses in combination with state-of-the-art genetic and proteomic tools (DNA/RNA-Seq, RNA antisense purification and LC-MS) to achieve our goals. The results from this study will improve our basic scientific knowledge of the biology of a lncRNA in HIV-1 pathogenesis and persistence in macrophages. It will also provide additional modalities for targeting SAF by identifying the regulators and effectors of this lncRNA. Understanding the mechanisms underpinning SAF-mediated cell survival and viral persistence in HIV-1 infected macrophages will aid greatly in optimally exploiting the novel therapeutic potential of this lncRNA.

项目摘要/摘要 联合抗逆转录病毒疗法在阻止HIV-1复制方面非常有效，从而获得了新的 感染，但它不能消除在治疗开始之前感染的储藏细胞。大多数关于HIV-1的研究 持久性主要集中在潜伏感染的CD4+T细胞上。然而，组织驻留的巨噬细胞 最近有研究表明，具有自我更新能力、寿命长的髓系细胞在缺乏 T淋巴细胞。巨噬细胞支持HIV-1复制而不屈服于病毒的独特能力- 诱导的细胞死亡使它们成为病毒持续存在的理想选择。了解病毒和宿主因素在 需要使巨噬细胞逃脱细胞死亡，才能制定有效的策略来消除这些 持续感染的髓系细胞。 通过筛选90个长的非编码RNA(LncRNA)，我们最近发现了这种新的作用 LncRNA SAF(Fas-AS1)在维持HIV-1感染巨噬细胞存活中的作用。这个lncRNA是 HIV-1感染的人单核细胞来源的巨噬细胞(MDM)在体外和肺中的显著上调 体内的肺泡巨噬细胞(AM)。更重要的是，siRNA介导的SAF抑制导致激活 选择性地感染HIV-1的巨噬细胞中的凋亡半胱氨酸酶，同时使暴露于病毒的巨噬细胞仍未被感染 旁观者细胞未受影响。这种对lncRNA SAF的高度特异性调制导致显著减少了 巨噬细胞培养中的病毒负荷，强调靶向这种lncRNA的治疗潜力。我们的 中心假说是活动性HIV-1感染诱导巨噬细胞SAF上调以调节细胞 促进病毒持续存在的生存/死亡途径(S)。我们建议对上游的两个 SAF转录调控因子及其功能的下游效应因子。 我们的具体目标是：(1)确定在HIV-1中诱导lncRNA SAF的病毒和细胞调节因子 感染巨噬细胞；以及(2)确定lncRNA SAF介导的保护巨噬细胞的分子机制 通过鉴定HIV-1的DNA/RNA和/或蛋白质靶标，从细胞死亡中感染HIV-1的巨噬细胞。 我们将使用一组特定的复制阶段缺陷病毒与最先进的基因 以及蛋白质组学工具(DNA/RNA-Seq、RNA反义纯化和LC-MS)来实现我们的目标。结果是 这项研究将提高我们对HIV-1发病机制中lncRNA生物学的基本科学知识 巨噬细胞中的持久性。它还将提供针对苏丹武装部队的其他方式，方法是确定 这个lncRNA的调节者和效应者。了解SAF介导的细胞存活的基础机制 而病毒在感染HIV-1的巨噬细胞中的持久性将极大地帮助优化利用这种新的治疗方法 这个lncRNA的潜力。