Mechanisms that couple irregular development of fetal melanoblasts to premature exhaustion of adult melanocyte stem cells

项目摘要 在成年期维持组织功能，从而抑制组织变性和疾病， 取决于干细胞数量的维持。成体干细胞是表观遗传编程的， 在胚胎中开始的一系列发育决定的顶点。子宫内环境暴露 胚胎可以影响成年和晚年疾病，可能部分通过影响干细胞发育， 传递到成人干细胞的维持和功能。然而， 胚胎发育和干细胞功能和表型的长期维持在成年人中很差， 定义了我们将采用谱系特异性组蛋白伴侣基因失活来了解 成黑素细胞（Mb）胚胎发育完整性对维持成年黑素细胞（Mc）干的影响 细胞（McSC）。 组蛋白伴侣HIRA将组蛋白变体H3.3沉积到活性基因、启动子和增强子中。 通过体外和体内研究以及野生型小鼠胚胎成黑素细胞（Mbs）的单细胞RNA-seq， 型小鼠和胚胎Mb中缺乏HIRA表达的小鼠，我们已经发现HIRA在 维持PAX 3/SOX 10-MITF Mb特化途径。在Mb中HIRA的失活消耗了 早期/中期胚胎中的Mbs。然而，这种胚胎缺陷在出生后得到了挽救， 正常数量的黑色素细胞，只有一个非常微妙的色素缺陷。然而，在新生儿 在小鼠中，Hira敲除（KO）黑素细胞表现出更高频率的端粒相关DNA损伤灶， 表明Hira敲除McSC和/或黑素细胞具有分子损伤，即使在新生小鼠中也是如此。 事实上，来自新生Hira KO小鼠的成黑素细胞和黑素细胞对促增殖因子的反应很差。 在体外和体内激发，这些小鼠显示出明显加速的McSC和黑素细胞消耗， 在成年期头发变白的速度急剧加快。基于这些广泛的初步数据，我们将 研究HIRA在黑素细胞谱系分化和发育中的作用，并研究HIRA在黑素细胞谱系分化和发育中的作用。 胚胎发育异常与成年期和衰老期间成体干细胞耗竭之间的联系。 PAX 3/SOX 10-MITF信号通路的失调导致发育障碍， 黑素瘤这些研究确定了HIRA在PAX 3/SOX 10-MITF轴中的作用，可以促进治疗。 干预措施，以打击这些发展和肿瘤疾病。此外，完成这些具体 目的是解决组织特异性干细胞胚胎发育的完整性如何影响维持 这些干细胞的数量。

PROJECT SUMMARY Maintenance of tissue function during adulthood, and hence suppression of tissue degeneration and disease, depends on maintenance of stem cell populations. Adult stem cells are epigenetically programmed and a culmination of a series of developmental decisions initiated in the embryo. In utero environmental exposures on the embryo can influence adult and late-life disease, likely in part via effects on stem cell development that are transmitted to maintenance and function of stem cells in the adult. However, the molecular links between embryonic development and long-term maintenance of stem cell function and phenotype in adults are poorly defined. We will employ lineage-specific genetic inactivation of a histone chaperone to understand how embryonic developmental integrity of melanoblasts (Mb) impacts on maintenance adult melanocyte (Mc) stem cells (McSC). The histone chaperone HIRA deposits histone variant H3.3 into active genes, promoters and enhancers. Through in vitro and in vivo studies and single cell RNA-seq of mouse embryo melanoblasts (Mbs) from wild type mice and mice lacking expression of HIRA in embryonic Mbs, we have uncovered a role for HIRA in sustaining the PAX3/SOX10-MITF Mb specification pathway. Inactivation of HIRA in Mbs depletes the number of Mbs in early/mid stage embryos. However, this embryonic defect is rescued by birth and young mice exhibit normal numbers of melanocytic cells, and only a very subtle pigmentation defect. Nevertheless, in new-born mice, Hira knock out (KO) melanocytic cells exhibit a higher frequency of telomere-associated DNA damage foci, indicating that Hira knock out McSC and/or melanocytes harbor molecular damage, even in new-born mice. Indeed, melanoblasts and melanocytes from new-born Hira KO mice respond poorly to pro-proliferative challenge in vitro and in vivo, and these mice show marked accelerated McSC and melanocyte depletion and dramatically accelerated hair greying during adulthood. Building on these extensive preliminary data, we will investigate the role of HIRA in differentiation and development of the melanocytic lineage, and investigate the links between abnormal embryonic development and adult stem cell depletion during adulthood and aging. Dysregulation of the PAX3/SOX10-MITF signaling pathway contributes to developmental disorders and melanoma. These studies to define HIRA's role in the PAX3/SOX10-MITF axis can promote therapeutic interventions to combat these developmental and neoplastic disorders. Moreover, completion of these Specific Aims will address how the integrity of embryonic development of tissue specific stem cells impacts maintenance of those stem cells during adulthood.