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: 10548654
• 负责人: Koh Fujinaga
• 金额: $ 17.98万
• 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 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10548654
• 关键词: Binding; Biochemical; CD4 Positive T Lymphocytes; Cell physiology; Cells; Complex; Cytoskeleton; DNA Polymerase II; DNA-Directed RNA Polymerase; 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; Regulation; Regulator Genes; Rest; Role; Testing; Therapeutic; Transcription Elongation; Transcriptional Elongation Factors; Untranslated RNA; Viral; Viral reservoir; Virus Latency; Virus Replication; antiretroviral therapy; base; chromatin isolation by RNA purification sequencing; clinically relevant; clinically significant; cyclin T1; differential expression; effective therapy; efficacy testing; experimental study; genetic testing; genome-wide; improved; insight; new therapeutic target; promoter; reactivation from latency; recruit; 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HIV储藏，了解控制HIV的分子事件至关重要 潜伏期。而宿主蛋白质因子在HIV转录和病毒潜伏期的调节中起着关键作用 我们对非编码转录组（尤其是长期非编码转录组）的理解是广泛研究的 编码RNA（LNCRNA），有限的病毒潜伏期控制受到限制。更好地了解监管角色 LNCRNA在控制HIV转录和病毒潜伏期的控制方面发挥了作用，我们采用了RNA-Seq分析并进行了比较 活化的HIV感染细胞中的转录组。我们确定了许多LNCRNA 因此，以差异表达的是艾滋病毒潜伏期调节的候选者。其中一个lncrna， 细胞骨架调节剂（细胞）激活HIV转录和病毒潜伏期。我们的结果还表明 细胞体的耗竭可通过降低RNA聚合酶II的占用（POL II）来抑制潜在的HIV重新激活 以及病毒启动子上组蛋白激活标记的水平。其他生化和蛋白质组学分析 结果表明，细胞分子占据了HIV启动子，并与阳性转录伸长因子B（p-） TEFB），这是HIV和细胞基因转录伸长的重要细胞因子。鉴于这些 调查结果，我们假设通过将P-TEFB募集到HIV启动子来激活HIV基因表达。 为了检验该假设，我们将确定细胞器是否直接结合P-TEFB并募集细胞 转录伸长复合物向HIV启动子。其他初步的RNA-seq分析表明 细胞部署会导致主机转录组的广泛变化。因此，我们将确定下游目标 细胞分子并确定其对HIV基因表达的间接影响。重要的是，我们的结果将进一步 在临床上相关的环境中确认，因为我们将操纵CD4+主细胞中的细胞表达和 确定细胞对艾滋病毒重新激活的治疗潜力的大小和潜伏期的反转或 诱导潜伏期。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 由lncrnas。它的成功完成将为开发新的基于RNA的疗法奠定基础 这将添加到当前的治疗方案中，以消除HIV感染和持续的病毒参与者。