Epigenetic Control of Nephron Progenitor Cell Lifespan

肾单位祖细胞寿命的表观遗传控制

• 批准号: 9755419
• 负责人: Samir S El-Dahr
• 金额: $ 33.9万
• 依托单位: TULANE UNIVERSITY OF LOUISIANA
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-08-15 至 2021-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9755419
• 关键词: ATAC-seq; Adult; Age; Aging; Biological; Cell Aging; Cells; ChIP-seq; Child; Chromatin; Chronic Kidney Failure; Clustered Regularly Interspaced Short Palindromic Repeats; Complex; Development; Differentiated Gene; End stage renal failure; Endowment; Enhancers; Epigenetic Process; Fostering; Gene Activation; Gene Expression; Gene Targeting; Genes; Genetic; Genetic Transcription; Genome; Growth Factor; Histone Code; Histones; Hypertension; Individual; Intrinsic factor; Kidney; Knowledge; Lead; Longevity; Maps; Mesenchyme; Modeling; Multipotent Stem Cells; Mus; Natural regeneration; Nephrons; Nuclear Pore Complex; Nucleic Acid Regulatory Sequences; Nucleosomes; Polycomb; Protocols documentation; Regulator Genes; Renal Replacement Therapy; Replacement Therapy; Repression; Role; Signal Transduction; Stem cells; Systems Biology; Testing; Tissues; Writing; age related; base; beta catenin; cell age; chromatin remodeling; epigenetic therapy; epigenome; epigenome editing; epigenomics; genome-wide; loss of function; mutant; nephrogenesis; novel strategies; premature; progenitor; promoter; self-renewal; stemness; transcription factor; transcriptome sequencing

Low nephron endowment causes hypertension, chronic kidney disease, and end-stage renal disease requiring renal replacement therapy in children and adults. The premise of multi-potent progenitor cell- based replacement therapy for individuals with low nephron endowment critically depends on scientific and technical advances that foster efficient propagation of native or pluripotent cell-derived nephron progenitor cells (NPC). Genetic and functional analyses in mice indicate that NPC residing in the cap mesenchyme “age” during maturation, i.e., while Young E13 NPC stay in the niche and engage in self-renewal, Old P0- P2 NPC have a shorter life span because they exit the niche at a higher rate and differentiate into nascent nephrons. The biological basis of NPC aging is not well understood. Changes in the niche microenvironment are not sufficient to explain Old NPC's greater propensity to differentiate since P2-NPC cannot sustain their progenitor state even in optimal growth factor media. Our preliminary studies using genome-wide mapping of open (accessible) chromatin identified intrinsic age-associated chromatin state transitions in Young and Old NPC. This exciting finding prompted us to hypothesize that Old NPC are epigenetically “primed” for differentiation, which contributes to their limited life span. Specific Aim 1 will utilize an integrative system biology approach to delineate the active enhancer landscape and transcriptional regulatory network of the Young and Old NPC. We will construct a comprehensive map of the age-dependent chromatin state transitions in freshly isolated Young (E13) and Old (P0-P2) NPC, and integrate this knowledge with enhancer histone signatures, transcriptional profiles, and transcription factor occupancy. We will also determine whether ex vivo expansion triggers remodeling of the chromatin landscape by comparing the epigenomic profiles of native and expanded Young and Old NPC. In Specific Aim 2, we will test the hypothesis that the Polycomb Repressive Complex 2 (PRC2) restrains remodeling of differentiation gene enhancers in the Young NPC. Using gene targeting and epigenome profiling, we will demonstrate the critical role of PRC2 in controlling access to enhancers of Cdkn2a/p16 and Wnt4. We will utilize CRISPR dCas9-targeted epigenome editing to re-write the histone signature of developmental enhancers and rejuvenate the PRC2-mutant NPC. Successful completion of these aims has the potential to advance the field of kidney development in general, but it will especially benefit the efforts of nephron regeneration. Knowledge of the enhancer landscape of nephron progenitors during maturation, which does not currently exist, can potentially open the way to development of targeted epigenetic therapy to maintain the stemness or rejuvenate the aging NPC, and to refine existing NPC propagation protocols.

肾单位禀赋低下导致高血压、慢性肾病和终末期肾病 儿童和成人需要肾脏替代治疗。多能祖细胞的前提—— 对肾单位禀赋低下的个体进行基于替代疗法的治疗关键取决于科学和 促进天然或多能细胞衍生的肾单位祖细胞有效繁殖的技术进步 细胞（NPC）。小鼠遗传和功能分析表明 NPC 存在于帽间充质中 成熟期的“年龄”，即年轻的E13 NPC停留在利基市场并进行自我更新，而老的P0- P2 NPC 的寿命较短，因为它们以较高的速率退出利基并分化为新生 肾单位。鼻咽癌衰老的生物学基础尚不清楚。利基市场的变化 微环境不足以解释老 NPC 自 P2-NPC 以来更倾向于分化 即使在最佳生长因子培养基中也无法维持其祖细胞状态。我们的初步研究使用 开放（可访问）染色质的全基因组图谱确定了内在的与年龄相关的染色质状态 年轻和年长 NPC 的过渡。这一令人兴奋的发现促使我们假设 Old NPC 是 表观遗传为分化“做好了准备”，这导致它们的寿命有限。具体目标 1 将 利用综合系统生物学方法来描绘活性增强子景观和 年轻和年长 NPC 的转录调控网络。我们将构建一个全面的地图 新鲜分离的年轻 (E13) 和老年 (P0-P2) NPC 中年龄依赖性染色质状态转变，以及 将这些知识与增强子组蛋白特征、转录谱和转录因子相结合 占用。我们还将确定离体扩增是否会触发染色质重塑 通过比较本地和扩展的年轻和老年 NPC 的表观基因组图谱。具体来说 目标 2，我们将检验多梳抑制复合物 2 (PRC2) 抑制重塑的假设 Young NPC 中的分化基因增强子。使用基因打靶和表观基因组分析，我们将 证明 PRC2 在控制 Cdkn2a/p16 和 Wnt4 增强子的访问方面的关键作用。我们将 利用 CRISPR dCas9 靶向表观基因组编辑来重写发育的组蛋白特征 增强子并使 PRC2 突变 NPC 恢复活力。成功完成这些目标有潜力 总体上推进肾脏发育领域，尤其有利于肾单位的努力 再生。了解肾单位祖细胞在成熟过程中的增强子景观，这确实 目前尚不存在，有可能为开发靶向表观遗传疗法以维持 干性或使老化的 NPC 恢复活力，并完善现有的 NPC 繁殖方案。