Cytor lncRNA as a positive regulator of HIV gene expression and viral latency

Cytor lncRNA 作为 HIV 基因表达和病毒潜伏期的正调节因子

• 批准号: 10681321
• 负责人: Koh Fujinaga
• 金额: $ 20.48万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-08-10 至 2025-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10681321
• 关键词: Binding; Biochemical; CD4 Positive T Lymphocytes; Cell Separation; Cell physiology; Cells; Complex; Cytoskeleton; Development; Event; Foundations; Gene Activation; Gene Expression; Gene Expression Regulation; Genes; Genetic Transcription; HIV; HIV Infections; HIV tat Protein; Histones; In Vitro; Integration Host Factors; Knowledge; Light; Mediating; Molecular; Monitor; Patients; Phosphotransferases; Physiological; Play; Positive Transcriptional Elongation Factor B; Proteins; Proteomics; Protocols documentation; RNA; RNA Polymerase II; Regulation; Regulator Genes; Rest; Role; Testing; Therapeutic; Transcription Elongation; Transcriptional Elongation Factors; Untranslated RNA; Viral; Viral reservoir; Virus Latency; Virus Replication; antiretroviral therapy; chromatin isolation by RNA purification sequencing; clinically relevant; clinically significant; cofactor; cyclin T1; differential expression; effective therapy; efficacy testing; experimental study; genetic testing; genome-wide; improved; insight; new therapeutic target; promoter; reactivation from latency; recruit; therapeutic RNA; tool; transcriptome; transcriptome sequencing

ABSTRACT To successfully eliminate the HIV reservoir, it is critical to understand the molecular events that control HIV latency. While the key roles that host protein factors play in the regulation of HIV transcription and viral latency have been extensively studied, our understanding of how the non-coding transcriptome, especially long non- coding RNA (lncRNA), contribute to viral latency control is limited. To better understand the regulatory roles lncRNAs play in the control of HIV transcription and viral latency, we employed RNA-Seq analysis and compared the transcriptome in activated versus resting HIV-infected cells. We identified numerous lncRNAs that are differentially expressed, therefore are candidates for HIV latency regulation. Among them, one lncRNA, Cytoskeleton Regulator (Cytor), activates HIV transcription and viral latency. Our results also indicate that the depletion of Cytor suppresses latent HIV reactivation by reducing the occupancy of RNA Polymerase II (Pol II) and the levels of histone activation markers on the viral promoter. Additional biochemical and proteomic analyses showed that Cytor occupies the HIV promoter and associates with Positive Transcription Elongation Factor b (P- TEFb), which is an essential cellular factor for transcription elongation of HIV and cellular genes. In light of these findings, we hypothesize that by recruiting P-TEFb to the HIV promoter, Cytor activates HIV gene expression. To test this hypothesis, we will determine whether Cytor directly binds P-TEFb and recruits the cellular transcription elongation complex to the HIV promoter. Additional preliminary RNA-Seq analysis indicated that Cytor depletion results in broad changes in the host transcriptome. We will therefore identify downstream targets of Cytor and determine their indirect effects on HIV gene expression. Significantly, our results will be further confirmed in a clinically relevant context, as we will manipulate Cytor expression in CD4+ primary cells and determine the magnitude of Cytor’s therapeutic potential for HIV reactivation and latency reversal or, alternatively, to latency induction. Our study will provide new insights into the regulation of HIV transcription and viral latency by lncRNAs. Its successful completion will lay the groundwork for the development of new RNA-based therapies that will be added to current therapeutic protocols to eliminate HIV infection and the persistent viral reservoir.

抽象的 为了成功消除艾滋病毒储存库，了解控制艾滋病毒的分子事件至关重要 而宿主蛋白因子在 HIV 转录和病毒潜伏期的调节中发挥着关键作用。 经过广泛的研究，我们对非编码转录组，特别是长非编码转录组的理解 编码RNA（lncRNA）对病毒潜伏期控制的贡献有限。 lncRNA 在控制 HIV 转录和病毒潜伏期中发挥作用，我们采用 RNA-Seq 分析并比较 我们鉴定了激活的 HIV 感染细胞与静息的 HIV 感染细胞的转录组。 差异表达，因此是 HIV 潜伏期调控的候选者，其中一种 lncRNA， 细胞骨架调节剂 (Cytor) 激活 HIV 转录和病毒潜伏期。 Cytor 的消耗通过减少 RNA 聚合酶 II (Pol II) 的占用来抑制潜在的 HIV 重新激活 以及病毒启动子上的组蛋白激活标记物的水平。额外的生化和蛋白质组学分析。 表明 Cytor 占据 HIV 启动子并与正转录延伸因子 b (P- TEFb)，它是 HIV 和细胞基因转录延伸的重要细胞因子。 研究结果表明，Cytor 通过将 P-TEFb 募集至 HIV 启动子来激活 HIV 基因表达。 为了检验这一假设，我们将确定 Cytor 是否直接结合 P-TEFb 并招募细胞 HIV启动子的转录延伸复合物的额外初步RNA-Seq分析表明 细胞耗竭会导致宿主转录组发生广泛变化，因此我们将确定下游靶标。 Cytor 并确定它们对 HIV 基因表达的间接影响。值得注意的是，我们的结果将进一步得到证实。 在临床相关背景下得到证实，因为我们将操纵 CD4+ 原代细胞中的 Cytor 表达，并且 确定 Cytor 对 HIV 再激活和潜伏期逆转的治疗潜力的大小，或者， 我们的研究将为 HIV 转录和病毒潜伏期的调节提供新的见解。 它的成功完成将为基于 RNA 的新疗法的开发奠定基础。 这将被添加到当前的治疗方案中，以消除艾滋病毒感染和持久的病毒库。