DNA结合蛋白Arid3b在干细胞多能性建立及定向分化中的功能与机制研究

Pluripotency of stem cell is co-regulated by multiple factors. In our previous studies, we defined a large number of newly found proteins which interact with the core transcription factors (TFs), including Oct4, Sox2 and Nanog (OSN), and we confirmed their key roles in pluripotency regulation. However, the function and mechanism of the DNA binding protein Arid3b, which interacts with all the three TFs (OSN), is still not reported. In our preliminary results, we found that Arid3b could promote the reprogramming efficiency of somatic cells, and also improve the mesendodermal fate decision of embryonic stem cell. In the upstream, Arid3b was regulated by the fate decision key factors of cardiomyocytes and B cell, and might be regulated by multiple RNA binding proteins and microRNAs. Moreover, histone demethylase and histone acetyltransferase were observed in Arid3b interactome. Thus, we hypothesize that Arid3b is regulated by microRNAs and RNA binding proteins, then regulates the establishment of pluripotency, as well as cardiomyocytes and B cell differentiation from ESC through histone modification. In the current project, via epigenetic and bioinformatic methods, we will investigate the detail functions and mechnisms of Arid3b in pluripotency regulation, which will provide scientific evidences and technical support for preclinical study and clinical transformation of pluripotent stem cells.

干细胞多能性受多种因子共同调控，我们前期阐明了核心转录因子Oct4、Sox2或Nanog（OSN）的部分互作蛋白在多能性调控网络中的重要作用。但与OSN共同互作的DNA结合蛋白Arid3b，在干细胞中是否起着核心调控作用及机制仍不清晰。近期我们预实验数据显示①Arid3b促进胚胎干细胞向中胚层分化；②调控体细胞重编程；③受中胚层来源的心肌细胞和B细胞发育决定因子的调控；④可能受多种RNA结合蛋白Ddx5与microRNA共同调控；⑤与组蛋白乙酰转移酶和组蛋白去甲基化酶互作。据此我们提出假设，Arid3b上游受RNA结合蛋白与microRNA调控，并通过组蛋白修饰调控干细胞多能性建立及向心肌和B细胞定向分化。本项目拟采用表观遗传学和生物信息学等方法，深层次阐明Arid3b在多能干细胞建立和定向分化中的功能、调控通路与表观机制，为多能性干细胞的临床转化提供理论依据和方法指导。

干细胞多能性主要受到核心多能性因子OCT4、SOX2、NANOG（OSN）的协同调控。ARID3B是与干细胞同时互作的转录因子，但是目前关于它的研究较少，且主要集中于肿瘤领域。对于其在干细胞中是否起着核心调控作用及机制仍不清晰。通过该项目的研究，我们发现：1）ARID3B对于小鼠胚胎干细胞多能性的维持、建立以及自我更新是必需的2）ARID3B能够通过在细胞核内形成液-液相分离的形式发挥其调控功能3）ARID3B高度富集在小鼠胚胎干细胞命运相关的超级增强子上，并且调控超级增强子上相分离的形成4）ARID3B能够通过在超级增强子上形成相分离，调控核心多能性因子在超级增强子上的富集，从而影响干细胞多能性。通过该研究，我们发现了ARID3B这一转录因子在小鼠胚胎干细胞多能性调控中发挥的重要作用。并且进一步揭示了干细胞命运相关超级增强子上相分离的形成机制以及核心多能性因子在其上的富集机制。将相分离与干细胞命运调控联系起来，是对干细胞多能性调控机制的一个重要补充，为多能性干细胞的临床转化提供了理论依据和方法指导。