Mutant p53 rejuvenates aged stem cells through modulating epigenetic regulators

项目总结/摘要 老年人面临着血液疾病的重大负担，包括贫血、克隆性贫血、贫血 造血、骨髓增生异常综合征（MDS）和急性髓性白血病（AML）。MDS是一组 老年人的血液疾病，在HSC中开始并经常进展为AML。虽然肿瘤中的突变 在老年健康个体和MDS患者的血细胞中发现了抑制性p53，突变型p53的作用可能是由于它们的基因突变而引起的。 p53在HSC衰老和MDS发病机制中的作用尚不清楚。我们最近发现，一些增益- 功能（GOF）突变的p53蛋白通过一种新的机制增强了HSC的自我更新潜力 包括破坏表观遗传途径。我们的目的是描述GOF突变型p53在 HSC老化和MDS的发病机制，并确定用于治疗MDS的新的治疗靶点。我们 假设在老化HSC中获得特异性GOF p53突变驱动了前- 具有增强的自我更新能力的白血病造血干细胞，从而允许克隆进化， 随后获得导致MDS干细胞形成的突变和/或表观遗传变化。 我们推测，药理学抑制途径的合成致死p53突变将消除药物- 耐药MDS干细胞和改善治疗结果。在这项研究中，我们将利用 生物化学、遗传学、分子学和药理学方法以及人类的脊椎动物模型 研究干细胞衰老和MDS发展的表观遗传调控。我们会委聘 无偏见的全基因组方法，包括RNA-seq，ChIP-seq和DNA甲基化测定，以确定 GOF突变型p53靶向并破译GOF突变型p53如何调节衰老HSC中的基因表达。在 此外，我们将描述p53 R248 W/+ Asxl1 +/-小鼠中MDS干细胞的特征，并阐明其机制， 突变型p53增强MDS干细胞自我更新。此外，我们将确定基因和途径， 使用CRISPR-Cas9筛选在人MDS细胞中对突变型p53的合成致死性。我们的成果将填补 关于白血病前造血干细胞和MDS干细胞的起源和发育的重大差距 细胞我们预期这些研究将描述特定突变p53蛋白在老化HSC中的作用， 白血病前HSC和MDS干细胞，并可能在人MDS细胞中鉴定p53 这些突变可能具有消除耐药MDS干细胞的治疗潜力。这些结果 可能会为衰老、MDS和癌症研究开辟新的途径。

PROJECT SUMMARY/ABSTRACT Elderly individuals are faced with a significant burden of hematologic diseases, including anemia, clonal hematopoiesis, myelodysplastic syndromes (MDS), and acute myeloid leukemia (AML). MDS are a group of blood diseases of the elderly that initiate in a HSC and frequently progress to AML. While mutations in tumor suppressor p53 are found in the blood cells of aged healthy individuals and in MDS patients, the role of mutant p53 in HSC aging and pathogenesis of MDS are largely unknown. We recently discovered that some gain-of- function (GOF) mutant p53 proteins enhanced the self-renewal potential of HSCs by a novel mechanism involving disruption of epigenetic pathways. Our objective here is to characterize the role of GOF mutant p53 in HSC aging and pathogenesis of MDS and identify novel therapeutic targets for the treatment of MDS. We hypothesize that acquisition of specific GOF p53 mutations in aged HSCs drives the development of pre- leukemic hematopoietic stem cells with enhanced self-renewal capability, thereby allowing clonal evolution and subsequent acquisition of mutations and/or epigenetic changes that lead to the formation of MDS stem cells. We speculate that pharmacological inhibition of pathways synthetic lethal to p53 mutations will eliminate drug- resistant MDS stem cells and improve treatment outcome. In this proposed research, we will utilize biochemical, genetic, molecular, and pharmacological approaches as well as vertebrate models of human MDS to investigate the epigenetic regulation of stem cell aging and MDS development. We will employ unbiased genome-wide approaches, including RNA-seq, ChIP-seq, and DNA methylation assays, to identify GOF mutant p53 targets and decipher how GOF mutant p53 regulates gene expression in aged HSCs. In addition, we will characterize MDS stem cells in p53R248W/+Asxl1+/- mice and elucidate the mechanism by which mutant p53 enhances MDS stem cell self-renewal. Further, we will identify genes and pathways that are synthetic lethal to mutant p53 in human MDS cells using CRISPR-Cas9 screening. Our results will fill a significant gap regarding the origin and development of pre-leukemic hematopoietic stem cells and MDS stem cells. We anticipate that these studies will delineate the effects of specific mutant p53 proteins in aged HSCs, pre-leukemic HSCs, and MDS stem cells, and will likely identify novel targets in human MDS cells with p53 mutations that may have therapeutic potential for eliminating drug-resistant MDS stem cells. These outcomes would likely open new avenues for aging, MDS, and cancer research.