Integrative Single-Cell Analysis of Transcriptome, Epigenome, and Lineage in HIV Latency and Activation

HIV 潜伏期和激活过程中转录组、表观基因组和谱系的综合单细胞分析

基本信息

批准号：
10543067
负责人：
Kathleen L. Collins
金额：
$ 69.36万
依托单位：
UNIVERSITY OF MICHIGAN AT ANN ARBOR
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-12-19 至 2024-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10543067
关键词：
ATAC-seq Acute Address Affect Atlases Basic Science Biochemical Bioinformatics Biological Markers Cell Lineage Cell Separation Cell model Cells Chromatin Chromium Clinical Treatment Clonal Expansion Complement Computer Models Computing Methodologies DNA Data Development Disease Foundations Gene Expression Genes Genetic Transcription Genome Goals Granulocyte-Macrophage Colony-Stimulating Factor HIV Histone Deacetylase Inhibitor Human In Vitro Infection Link Macrophage Maps Marker Discovery Methods Modeling Patients Pattern Population Proviruses RNA Reporter Residual state Sampling Shock Site Surveys System TNF gene Time Viral Viral Genome Viral reservoir Virion Virus Activation Virus Integration Virus Latency Work XCL1 gene antiretroviral therapy biomarker identification cell type computer framework env Gene Products epigenome epigenomics hematopoietic differentiation insight integration site latent infection multiple omics novel single cell analysis single-cell RNA sequencing transcriptome transcriptomics

项目摘要

Abstract With the development of triple combination antiretroviral therapy, routine HIV treatment eliminates nearly all actively infected cells. Nevertheless, the small reservoir of latently infected cells, which can remain dormant for long periods of time before becoming active and producing new virus particles, represents a crucial barrier to completely curing the disease. Identifying markers that identify latently infected cells or the biochemical factors that control latency activation could enable the effective use of a “shock and kill” strategy, where specific targeting or activation of latently infected cells eliminates the viral reservoir. Our recent work suggests that the global transcriptomic and epigenomic changes during hematopoietic differentiation affect viral latency and activation. Additionally, we recently found that global inhibition of histone deacetylase activity increases viral activation in these cells, further implicating epigenomic changes in activation. These results raise fundamental questions: What are the markers of latently infected cells? How do the transcriptomic and epigenomic state of a cell affect latency and activation? How does differentiation state relate to viral latency? Here, we leverage our experimental platform for identifying latently and actively infected cells, single cell transcriptome and epigenome sequencing, and our recently developed computational integration methods to investigate these questions. Our interdisciplinary team combines expertise in HIV basic science, HIV clinical treatment, and bioinformatics to develop an experimental and computational framework for integrated gene expression, chromatin accessibility, and lineage into a single picture of viral latency and activation. Specifically, this project will (1) use single-cell RNA-seq and single-cell ATAC-seq to map diversity of infected cells, (2) investigate the relationship between hematopoietic differentiation state and viral activation, (3) determine viral integration sites through single-cell RNA-seq, (4) computationally integrate single cell transcriptome and epigenome profiles, and (5) computationally infer cell lineage relationships among viral genomes and infected cells. To accomplish these goals, we will carry out the following aims: (1) Characterize lineage, transcriptomic and epigenomic diversity of single latently and actively infected primary cells. (2) Investigate latency and activation during in vitro differentiation. (3) Survey single cell diversity of re-activated and in vitro infected cells from cART-suppressed patients. Together, these aims will produce a comprehensive, integrated transcriptomic and epigenomic atlas of the HIV reservoir, identify DNA and RNA biomarkers of latency, and characterize clonal expansion patterns. Our work also develops a broadly applicable experimental and computational framework, laying a foundation for the discovery of novel insights into HIV latency and activation.

抽象的随着三合一组合抗逆转录病毒疗法的发展，常规HIV治疗几乎消除了积极感染的细胞。然而，潜在受感染的细胞的小小容，可能仍处于休眠状态长时间活跃之前并产生新病毒颗粒，代表了至关重要的障碍完全治愈疾病。识别识别潜在感染细胞或生化因素的标记控制潜伏期激活可以有效地使用“震惊和杀戮”策略或受感染细胞的激活消除了病毒储层。我们最近的工作表明全球造血分化过程中的转录组和表观基因组变化会影响病毒潜伏期和激活。此外，我们最近发现，整体抑制组蛋白脱乙酰基酶活性会增加病毒活化这些细胞，激活中的进一步隐式表观基因组变化。这些结果提出了基本问题：潜在感染细胞的标记是什么？细胞的转录组和表观基因组状态如何影响潜伏期和激活？分化状态与病毒潜伏期有何关系？在这里，我们利用我们的实验用于识别潜在和主动感染的细胞，单细胞转录组和表观基因组测序的平台，以及我们最近开发的计算整合方法来研究这些问题。我们的跨学科团队将HIV基础科学，HIV临床治疗和生物信息学方面的专业知识结合在一起开发一个实验和计算框架，用于集成基因表达，染色质可及性，并将谱系纳入病毒潜伏期和激活的单一图片。具体来说，该项目将（1）使用单细胞 RNA-Seq和单细胞ATAC-SEQ以绘制受感染细胞的多样性，（2）研究造血分化状态和病毒激活，（3）通过单细胞确定病毒整合位点 RNA-seq，（4）计算整合的单细胞转录组和表观基因组轮廓，以及（5）计算推断病毒基因组和感染细胞之间的细胞谱系关系。为了实现这些目标，我们将携带以下目的：（1）表征单一的谱系，转录组和表观基因组多样性，积极感染的原代细胞。（2）研究体外分化过程中的潜伏期和激活。（3）调查被抑制的患者重新激活和体外感染细胞的单细胞多样性。在一起，这些 AIMS将产生艾滋病毒水库的全面，综合的转录组和表观基因组学地图，识别延迟的DNA和RNA生物标志物，并表征克隆膨胀模式。我们的工作也发展了广泛适用的实验和计算框架，为发现新颖的基础奠定了基础对艾滋病毒潜伏期和激活的见解。