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

基本信息

批准号：
10159666
负责人：
Saikat Boliar
金额：
$ 23.55万
依托单位：
CORNELL UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-03-17 至 2023-02-28
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10159666
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

项目摘要

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.

项目概要/摘要联合抗逆转录病毒疗法在阻断 HIV-1 复制方面非常有效，因此成为新的治疗方法但它不能消除治疗开始前感染的储存细胞。大多数 HIV-1 研究持久性主要集中在潜伏感染的 CD4+ T 细胞上。然而，组织驻留巨噬细胞自我更新、寿命长的骨髓细胞最近被证明可以在缺乏病毒的情况下维持长时间的病毒血症 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的作用需要使巨噬细胞逃避细胞死亡，以制定有效的策略来消除这些细胞持续感染的骨髓细胞。通过筛选一组 90 个长非编码 RNA (lncRNA)，我们最近发现了新的作用 lncRNA SAF (FAS-AS1) 在维持 HIV-1 感染巨噬细胞的细胞存活中的作用。这个lncRNA是在体外和肺中 HIV-1 感染的人单核细胞衍生巨噬细胞 (MDM) 中显着上调体内肺泡巨噬细胞（AM）。更重要的是，siRNA介导的SAF抑制导致激活 HIV-1 感染的巨噬细胞中选择性地产生凋亡半胱天冬酶，同时使病毒暴露但未受感染旁观者细胞不受影响。 lncRNA SAF 的这种高度特异性调节导致巨噬细胞培养物中的病毒负荷，强调了针对该 lncRNA 的治疗潜力。我们的中心假设是，活跃的 HIV-1 感染会诱导巨噬细胞中 SAF 上调，从而调节细胞促进病毒持续存在的生存/死亡途径。我们建议对上游进行深入分析 SAF 转录的调节因子及其功能的下游效应子。我们的具体目标是：(1) 定义在 HIV-1 中诱导 lncRNA SAF 的病毒和细胞调节因子受感染的巨噬细胞； (2)确定lncRNA SAF介导的保护的分子机制通过识别该 lncRNA 的 DNA/RNA 和/或蛋白质靶标，HIV-1 感染巨噬细胞免于细胞死亡。我们将使用一组特定的复制阶段缺陷病毒与最先进的遗传技术相结合和蛋白质组学工具（DNA/RNA-Seq、RNA 反义纯化和 LC-MS）来实现我们的目标。结果这项研究将提高我们对 HIV-1 发病机制中 lncRNA 生物学的基础科学知识巨噬细胞中的持久性。它还将通过识别苏丹武装部队的目标，提供额外的模式该 lncRNA 的调节子和效应子。了解 SAF 介导的细胞存活的机制 HIV-1感染的巨噬细胞中病毒的持续存在将极大地有助于最佳地利用新的治疗方法该lncRNA的潜力。