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，潜伏感染细胞的储存库形成，在治疗期间持续存在， 这个储库代表了HIV治愈的主要障碍。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，其特征在于染色质结构的明显变化 以及抑制HIV基因表达的组蛋白修饰，这些修饰可以通过细胞传播， 师.潜伏期的这种遗传特性允许潜伏表型在正在进行的克隆中存活下来。 感染细胞在宿主中的扩张，并维持整个水库。此外，病毒沉默 表观遗传编程与CD 4 T细胞内发生的全局变化有关， 从激活状态到静息状态的转变。完全定义了储层的形成过程 形成和编程延迟将是必不可少的指导新的方法， 防止水库播种或维护。迄今为止，研究成藏过程 在临床上，HIV感染被认为是困难的，如果不是不可能的话，由于先前的 观察结果表明，在急性感染过程中，储库很早就形成了。较新 然而，来自几个小组的信息现在显示，水库的大部分是 接种的病毒在ART开始时正在积极复制，这表明ART 引发触发了储存器的播种。这一观察提出了一个令人兴奋的可能性， 我们可以通过纵向观察CD 4 T来研究水库播种的过程， 在ART后立即从PWH中提取细胞，通过这样做，揭示了 在治疗开始时阻止储液器形成。在本建议中，我们的目标是 全面定义表观遗传和基于染色质的变化，发生在CD 4 T细胞中， 使用来自A5248群组的样品的ART后时期。来自这些样品的CD 4 T细胞 将使用一系列已建立并验证的尖端检测方法进行分析， 我们的实验室这些测定包括组合的单细胞多组学（RNAseq/ATACseq） 多个关键组蛋白修饰的高分辨率全基因组作图， 使用核酸酶（CUT&RUN）在靶下切割和释放。通过这种方法，我们将 实现一个综合的理解如何遗传模式的染色质为基础的变化， CD 4 T细胞由ART启动触发，并产生了对ART的新的和关键的见解。 细胞内条件与体内储库接种一致。通过实现这一目标，我们将 揭示了促进水库播种的机制，并提出了新的方法来阻止 在ART开始时形成储层。