胚胎发育相关基因可参与肿瘤干细胞的调控。前期研究发现胰岛发育相关基因Rfx6与肝前体细胞标记物的表达相关；肝癌组织中Rfx6高表达提示肝癌分化差及不良预后；Rfx6过表达可激活ERK蛋白并上调β-catenin表达，促进肝癌细胞增殖、迁移、侵袭及化疗抵抗。因此，我们推测Rfx6可能与肝癌干细胞特征相关。本研究拟：⑴通过Rfx6启动子报告系统、小鼠肝原位移植瘤模型、干细胞球形成等实验，探讨Rfx6与肝癌干细胞干性特征的关系；⑵构建Rfx6表达上调/下调的肝癌细胞模型，进一步探讨Rfx6对肝癌干细胞干性特征的调控作用；⑶结合ERK抑制剂，了解Rfx6-ERK-β-catenin通路对维持肝癌干细胞干性特征的作用，通过ChIP-seq等筛选Rfx6调控的下游基因，探讨Rfx6调控肝癌干细胞干性特征的机制。本课题旨在更深入了解肝癌干细胞的特征，为寻找潜在分子靶点提供新的依据。

Embryonic development associated-genes have recently been gained wide attention due to involving in the regulation of cancer stem cells (CSC). Regulatory factor x6 (Rfx6) was identified as a new member of Rfx family, and regulated the differentiation of pancreatic islet cell. Our previous study showed that Rfx6 mRNA expression was positively correlated with hepatic precursor cell markers, and higher Rfx6 mRNA expression was associated with poor differentiation of tumors and worse prognosis in hepatocellular carcinoma (HCC) patients after partial hepatectomy. In addition, overexpression Rfx6 led to active ERK, increase expression of β-catenin, and promote proliferation, migration, invasion and chemoresistance in HCC cell lines. Therefore, we hypothesized that Rfx6 may involve in the maintaining HCC stemness and the Rfx6-ERK-β-catenin pathway may play a role in regulating the self-renew in HCC cell. To verify it, we will first use promoter reporter system, animal model, and sphere formation assay and other experiments to illustrate the correlation of Rfx6 with HCC stemness in Rfx6pos/Rfx6neg HCC cells. Viral-mediated overexpression/reduction of Rfx6 expression cell lines will be constructed to explore the role of Rfx6 in regulation of HCC stemness. Furthermore, ERK inhibitor will also be used to investigate the role of Rfx6-ERK-β-catenin pathway in maintaining stemness of HCC, and ChIP-seq will be employed to identify genes potentially regulated by Rfx6. Our study will help to unveil the role and mechanisms of Rfx6 involved in HCC stemness. In addition, this study will provide new insight for target therapy of HCC.