3D Genome Reorganization and Epigenome Dynamics of Clonal Hematopoiesis

项目总结 克隆性造血(CH)通常随着年龄的增长而发生，并与死亡率增加相关，这是一种更高的风险 白血病和心血管疾病。随着美国老年人人口的增加，有一个紧迫的问题 未得到满足的需求以缓解CH.年龄相关性CH是指具有体细胞的造血细胞的异常扩张 突变，主要发生在编码DNA甲基转移酶DNMT3A和去甲基酶TET2的基因上。这些 ‘ch突变’改变了造血干细胞(HSC)的表观基因组(即DNA甲基化和染色质 可访问性)在促进者和增强者。为了更好地了解这些变化如何影响基因调控 为了赋予HSC的子集竞争优势，并为CH的发展做出贡献，我们提出了多个 考虑表观基因组变化对物理相互作用影响的组学方法 增强子和启动子，它们是激活转录的关键。通过绘制基因组的3D染色质 相互作用，我们将确定特定的增强子-启动子对控制关键的HSC细胞功能，如 自我更新和对炎症的反应。尽管已知造血干细胞的表观基因组组织 随着年龄的增长而变化，目前尚不清楚衰老和CH突变是如何共同重塑‘3D表观基因组’(3D 染色质组织和表观基因组)，并对CH.我们的中心假设是3D 表观基因组随着年龄的增长而重塑，并影响对HSC功能至关重要的转录程序。因此，要确定 延长健康寿命、延缓或预防CH相关疾病发生的治疗靶点 美国老龄化人口中的疾病，这项提议试图系统地揭示3D表观基因组 在CH中的细胞竞争的配置，并优先在早期衰老出现的候选基因靶点。 利用CH和人类原代细胞的小鼠模型，我们将通过批量和单细胞多细胞培养来验证这一假说。 正常和突变的组学图谱(包括rna-seq、染色质可及性和染色质相互作用) 以系统地确定其在衰老早期的3D表观基因组重塑。在目标1中，我们将确定 3D表观基因组组织是否与小鼠和人类HSCs的衰老转录程序有关 鉴定和整合年轻的HSC特有的3D基因组、表观基因组和转录组特征， 两个物种的中老年个体并验证顶级候选基因在HSC衰老中的作用 活着。在目标2中，我们将定义与年龄相关的三维表观基因组重塑在CH小鼠模型和 验证顶级候选基因在体内克隆扩增突变的HSCs中的作用。老化的签名 中年出现可能是老年造血干细胞功能改变的最初原因。因此，这项研究将 揭示治疗干预的主要和可修改的目标。在目标3中，我们将确定 老年HSC对表观遗传治疗和抗炎药物的CH相关染色质重组 活着。我们预计，这项研究的结果将加速制定早期干预措施，以 在老年人中抢占CH相关疾病的先机。

PROJECT SUMMARY Clonal hematopoiesis (CH) commonly occurs with aging and is associated with increased mortality, a higher risk of leukemia, and cardiovascular disease. As the population of older adults expands in the US, there is an urgent unmet need to mitigate CH. Age-related CH is the abnormal expansion of hematopoietic cells that have somatic mutations, mostly in genes encoding the DNA methyltransferase DNMT3A and the demethylase TET2. These 'CH-mutations' change the hematopoietic stem cell (HSC) epigenome (i.e., DNA methylation and chromatin accessibility) at promoters and enhancers. To better understand how these changes impact gene regulation to confer a competitive advantage to a subset of HSCs and contribute to CH development, we propose a multi- omics approach that considers the effects of epigenomic changes on the physical interactions between enhancers and promoters, which are critical to activating transcription. By mapping the genome's 3D chromatin interactions, we will identify the specific enhancer-promoter pairs that control key HSC cellular functions such as self-renewal and response to inflammation. Although it is known that the epigenome organization of HSCs changes with age, it remains unclear how aging and CH-mutations jointly remodel the '3D epigenome' (3D chromatin organization and epigenome) of HSCs and contribute to CH. Our central hypothesis is that the 3D epigenome is remodeled with age and impacts transcriptional programs critical for HSC function. Thus, to identify therapeutic targets for elongating healthspan and delaying or preventing the occurrence of CH-associated disease in the aging US population, this proposal seeks to systematically uncover the 3D epigenomic configurations of cellular competition in CH and to prioritize candidate gene targets that emerge in early aging. Using mouse models of CH and human primary cells, we will test this hypothesis via bulk and single-cell multi- omics mapping (including RNA-seq, chromatin accessibility, and chromatin interactions) of normal and mutant HSCs to systematically determine their 3D epigenome remodeling during early aging. In Aim 1, we will determine whether 3D epigenome organization is linked to an aging transcriptional program in mouse and human HSCs by identifying and integrating HSC-specific 3D genomic, epigenomic, and transcriptomic signatures from young, middle-age, and old individuals of both species and validate the roles of top candidate genes in HSC aging in vivo. In Aim 2, we will define age-dependent 3D epigenome remodeling in HSCs of CH mouse models and validate the roles of top candidate genes in clonal expansion of mutant HSCs in vivo. Aging signatures that emerge at middle age will likely be the initial cause of functional changes in aged HSCs. Thus, the study will reveal primary and modifiable targets for therapeutic intervention. In Aim 3, we will determine the response of CH-associated chromatin reorganization in aged HSCs to epigenetic therapy and anti-inflammatory drugs in vivo. We anticipate that the findings from this study will accelerate the development of early interventions to preempt CH-associated disease in aging individuals.