内质网应激对人成体干细胞功能的影响及机制研究

Loss of proteostasis is one of the hallmarks of aging. Aging and aging-related diseases are linked to impaired proteostasis. Adult stem cells are vital for body repair and tissue regeneration, and its dysfunction is closely associated with organism aging and related disorders. However, the limited understanding of the regulation of stem cell senescence or dysfunction during aging or in pathologic settings largely hinders the translational application of stem cells. So far, it is still unclear whether and how endoplasmic reticulum stress (ER stress) response could impact on human adult stem cells. Here, we propose to use CRISPR/Cas9-mediated targeted gene editing to knockout the classical ER stress components, and study the functional outcomes on human adult stem cells including neural stem cells and mechenchymal stem cells. By combination of epigenomic, proteomic and genomic analyses, we expect to reveal the ER stress-associated molecular signaling networks in different types of human adult stem cells. The implement of the project will provide theoretical basis to a better understanding of the mechanism of human adult stem cell aging, as well as a platform for seeking novel drug intervenes.

蛋白内稳态失衡是个体衰老的特征之一。内质网应激反应可诱发蛋白稳态失衡，其功能紊乱是引发个体衰老、老年性疾病发生的病因之一。人成体干细胞对于人体自我修复和组织再生至关重要，其功能紊乱与衰老、疾病密不可分，然而，由于人们对病理或衰老状态下干细胞功能调控机理缺乏深入理解限制了干细胞的临床应用。目前，尚不清楚内质网应激反应对人成体干细胞功能及衰老的影响。阐明内质网应激反应对成体干细胞功能的影响将为理解和治疗老年性疾病提供理论依据。本课题拟利用全新CRISPR/Cas9基因组靶向修饰技术构建内质网应激通路基因缺失的人类胚胎干细胞（hESC），以此为“起始材料”进行定向分化，以获得高纯度人成体神经干细胞（hNSC）及间充质干细胞（hMSC），同时结合表观遗传学组、蛋白组学、基因组学等分析多种方法，阐明内质网应激对特定成体干细胞功能及衰老的影响及分子机理。

内质网对细胞内蛋白稳态的维持起到非常重要的作用，当细胞遭遇外界各种刺激时将导致内质网中积累大量未折叠蛋白，从而诱发内质网应激反应。内质网应激失调导致的蛋白质稳态失衡是造成衰老的重要原因，与个体衰老、老年性疾病的发生密切相关。成体干细胞的自我更新与分化被认为是机体组织器官损伤后再修复，保持机体正常功能的重要基础。成体干细胞功能紊乱或衰老被认为是导致个体衰老的最重要因素。ATF6作为内质网应激反应的重要转录因子，它是否参与了干细胞稳态及衰老的调控是重要的科学问题。本课题首次利用CRISPR/Cas9技术、干细胞定向分化等技术获得ATF6基因敲除的人类干细胞模型，首次发现ATF6蛋白缺失导致多个膜细胞器稳态的失衡，最终导致人间充质干细胞加速衰老。同时通过多组学技术首次鉴定了一系列受ATF6调控的新型靶基因，发现FOS作为ATF6的下游靶基因参与膜细胞器稳态的调控。本课题揭示了内质网应激失调导致干细胞衰老的分子机制，也为未来干预衰老提供了重要的理论依据。