Understanding HIV reservoir formation by profiling transcriptomic and epigenetic changes in CD4 T cells following ART initiation

通过分析 ART 启动后 CD4 T 细胞的转录组和表观遗传变化来了解 HIV 储存库的形成

• 批准号: 10759940
• 负责人: Edward P Browne
• 金额: $ 27.21万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-08-09 至 2025-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10759940
• 关键词: ATAC-seq; Biological Assay; CD4 Positive T Lymphocytes; Cell Compartmentation; Cell division; Cells; Chromatin; Chromatin Structure; Clinical; Clonal Expansion; Code; Data; Data Set; Effector Cell; Epigenetic Process; Evolution; Gene Expression; Gene Silencing; Genes; Genetic Transcription; Global Change; Goals; HIV; HIV Genome; HIV Infections; Heritability; Histone Code; Histones; Inflammation; Laboratories; Link; Maintenance; Maps; Memory; Methods; Modification; Outcome; Pathway interactions; Peripheral Blood Mononuclear Cell; Persons; Phenotype; Process; Property; Repression; Resolution; Rest; Sampling; Series; Shapes; Tail; Testing; Time; Viral; Viral Antigens; Viral reservoir; Virus; acute infection; antiretroviral therapy; cohort; epigenetic silencing; genome-wide; genome-wide analysis; heritability pattern; histone modification; in vivo; insight; latent HIV reservoir; multiple omics; novel; novel strategies; novel therapeutics; nuclease; prevent; single-cell RNA sequencing; transcription factor; transcriptome sequencing; transcriptomic profiling; transcriptomics; transmission process

During HIV infection, a reservoir of latently infected cells forms that persists during therapy, and this reservoir represents the major barrier to an HIV cure. We and others have shown that HIV latency is an epigenetic phenomenon, characterized by distinct changes to chromatin structure and histone modifications that repress HIV gene expression, that can be transmitted through cell division. This heritable property of latency allows the latent phenotype to survive ongoing clonal expansion of infected cells in hosts and sustain the overall reservoir. Furthermore, viral silencing and epigenetic programming is linked to global changes that occur within CD4 T cells as they transition from an activated to a resting state. Fully defining the process by which the reservoir is formed and by which latency is programmed will be essential to guiding novel approaches to prevent reservoir seeding or maintenance. To date, studying the process of reservoir formation during clinical HIV infection has been considered difficult, if not impossible, due to prior observations suggesting that the reservoir forms very early during acute infection. Newer information from several groups, however, has now revealed that the majority of the reservoir is seeded by viruses that are actively replicating at the time of ART initiation, suggesting that ART initiation triggers seeding of the reservoir. This observation raises the exciting possibility that we could potentially study the process of reservoir seeding by longitudinal observation of CD4 T cells from PWH in the immediately post ART period and, by doing so, reveal potential ways to block the reservoir from forming at the time of therapy initiation. In this proposal we aim to comprehensively define epigenetic and chromatin-based changes that occur in CD4 T cells in the post ART period using samples derived from the A5248 cohort. CD4 T cells from these samples will be profiled using a series of cutting-edge assays that have been established and validated in our laboratory. These assays include combined single-cell multi-omic (RNAseq/ATACseq) analysis, and high-resolution genome-wide mapping of multiple key histone modifications through Cleavage Under Targets and Release using Nuclease (CUT&RUN). From this approach we will achieve an integrated understanding of how heritable patterns of chromatin-based changes in CD4 T cells are triggered by ART initiation and generate novel and critical insights into the intracellular conditions that coincide with reservoir seeding in vivo. By achieving this goal, we will reveal mechanisms that promote reservoir seeding and suggest novel approaches to block reservoir formation at the time of ART initiation.

在 HIV 感染期间，会形成潜伏感染细胞库，并在治疗期间持续存在，并且 这个水库是艾滋病毒治疗的主要障碍。我们和其他人已经证明，艾滋病毒 潜伏期是一种表观遗传现象，其特征是染色质结构的明显变化 和抑制 HIV 基因表达的组蛋白修饰，可通过细胞传播 分配。这种潜伏期的遗传特性使得潜伏表型能够在持续的克隆过程中存活下来。 受感染细胞在宿主体内扩张并维持总体储存库。此外，病毒沉默 表观遗传编程与 CD4 T 细胞内发生的全局变化有关，因为它们 从激活状态转变为静止状态。完全定义储层的过程 形成和编程延迟对于指导新方法至关重要 防止水库播种或维护。目前，对油藏形成过程的研究 由于先前的研究，在临床艾滋病毒感染期间被认为是困难的，即使不是不可能的 观察结果表明，病毒库在急性感染过程中很早就形成。较新 然而，来自多个团体的信息现已显示，水库的大部分已被 由在 ART 启动时正在积极复制的病毒播种，这表明 ART 启动触发了储层的播种。这一观察提出了令人兴奋的可能性： 我们可以通过 CD4 T 的纵向观察来研究水库播种的过程 ART 后立即从 PWH 中提取细胞，通过这样做，揭示了潜在的方法 在治疗开始时阻止水库形成。在本提案中，我们的目标是 全面定义 CD4 T 细胞中发生的表观遗传和染色质变化 ART 后阶段使用来自 A5248 队列的样本。来自这些样本的 CD4 T 细胞 将使用一系列已建立并验证的尖端分析进行分析 我们的实验室。这些测定包括组合单细胞多组学 (RNAseq/ATACseq) 分析以及多个关键组蛋白修饰的高分辨率全基因组图谱 使用核酸酶在目标下切割并释放（CUT&RUN）。通过这种方法，我们将 全面了解基于染色质的变化的遗传模式如何 CD4 T 细胞由 ART 启动触发，并产生新颖且批判性的见解 与体内储库接种相一致的细胞内条件。通过实现这一目标，我们将 揭示促进水库播种的机制并提出新的阻塞方法 ART 启动时储库的形成。