人胚胎干细胞（Human Embryonic Stem Cells, hESCs）可以分化成各种不同的组织和细胞，但在体外维持其未分化状态的条件比较苛刻。前期我们发现Wnt/β-catenin信号通路的小分子抑制剂IWR1可以有效地促进hESCs的自我更新，而激活此信号通路则会诱导分化。为了解析IWR1促进hESCs自我更新的功能与β-catenin的关系及其下游的分子机制，我们将首先运用基因编辑工具CRISPR/Cas9敲除β-catenin基因，以明确IWR1促进hESCs自我更新是否依赖β-catenin的存在；然后利用转录组测序技术筛选被IWR1影响的下游基因，通过基因表达的上调和干扰技术，最终鉴定出能够介导IWR1调控hESCs自我更新的下游关键因子。本课题国内外尚无报道，有望揭示调控hESCs多能性的新机制，深化目前人们对hESCs自我更新调控网络的认识，利于其基础和应用研究。

Human embryonic stem cells (hESCs) are able to differentiate into any type of human cell and tissue. However, they can not be efficiently maintained in vitro. Recently, we found that inhibition of Wnt/β-catenin signaling can greatly promote hESC self-renewal with small molecular IWR1, whereas activation of this pathway will induce hESC differentiation. In order to investigate the molecular mechanism under the self-renewal-promoting effect of IWR1 and the relationship between IWR1 and β-catenin, we will first generate β-catenin-null hESCs using CRISPR/Cas9 to determine if β-catenin is necessary for IWR1 to maintain hESC identity. Then we will perform RNA sequencing, combined with gene gain- and loss-of-function techniques, to screen and identify the core transcriptional factors regulated by IWR1 implicated in promoting hESC self-renewal. Our project will provide an expanded understanding of the current regulatory network of hESC pluripotency, which will not only lay the foundation for the future applications of hESCs, but will also facilitate the development of new culture conditions for the derivation of ESCs from other species.