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.

低肾单位禀赋导致高血压、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- 低肾单位禀赋个体的基础替代疗法关键取决于科学和 促进天然或多能细胞衍生的肾单位祖细胞的有效增殖的技术进展 细胞（NPC）。在小鼠中的遗传和功能分析表明，NPC居住在帽间充质中， 在成熟期间的“年龄”，即，年轻的E13 NPC在利基市场中自我更新，而老的P0- P2 NPC具有较短的寿命，因为它们以较高的速率退出生态位并分化为新生的细胞。 肾单位NPC衰老的生物学基础还不清楚。利基的变化 微环境不足以解释老NPC比P2-NPC更大的分化倾向 即使在最佳生长因子培养基中也不能维持它们的祖细胞状态。我们的初步研究使用 开放（可接近）染色质的全基因组定位确定了内在的年龄相关染色质状态 年轻人和老年人的NPC。这一令人兴奋的发现促使我们假设老NPC是 表观遗传学上的“启动”分化，这有助于他们有限的寿命。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， 年轻人和老年人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中的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标记、转录谱和转录因子结合起来 占用率我们还将确定体外扩增是否会引发染色质重塑， 通过比较天然和扩增的年轻和老年NPC的表观基因组图谱，在特定 目的2，我们将测试多梳抑制复合物2（PRC 2）抑制重构的假设。 在年轻的NPC分化基因增强。使用基因靶向和表观基因组分析，我们将 证明PRC 2在控制Cdkn 2a/p16和Wnt 4的增强子的接近中的关键作用。我们将 利用CRISPR dCas 9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PRC 2突变型NPC恢复活力。成功实现这些目标， 一般来说，这将促进肾脏发育的领域，但它将特别有利于肾单位的努力， 再生了解肾单位祖细胞成熟过程中的增强子景观， 目前还不存在，可以潜在地开辟一条发展靶向表观遗传治疗的道路， 使老化的NPC变干或恢复活力，并改进现有的NPC传播协议。