Sox9 confers stemness properties in hepatocellular carcinoma through Frizzled-7 mediated Wnt/β-catenin signaling

// Carmen Oi-Ning Leung 1 , Wing-Nga Mak 1 , Alan Ka-Lun Kai 1 , Kwan-Shuen Chan 1 , Terence Kin-Wah Lee 1, 2 , Irene Oi-Lin Ng 1, 2 , Regina Cheuk-Lam Lo 1, 2 1 Department of Pathology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Pok Fu Lam, Hong Kong 2 State Key Laboratory for Liver Research, The University of Hong Kong, Pok Fu Lam, Hong Kong Correspondence to: Regina Cheuk-Lam Lo, email: reginalo@pathology.hku.hk Irene Oi-Lin Ng, email: iolng@hku.hk Keywords: Sox9, liver cancer, tumor-initiating cells Received: October 05, 2015      Accepted: March 29, 2016      Published: April 19, 2016 ABSTRACT Sox9, an SRY-related HMG box transcription factor, is a progenitor/precursor cell marker of the liver expressed during embryogenesis and following liver injury. In this study, we investigated the role of Sox9 and its molecular mechanism with reference to stemness properties in hepatocellular carcinoma (HCC). Here, we observed upregulation of Sox9 in human HCC tissues compared with the non-tumorous liver counterparts ( p < 0.001). Upregulation of Sox9 transcript level was associated with poorer tumor cell differentiation ( p = 0.003), venous invasion ( p = 0.026), advanced tumor stage ( p = 0.044) and shorter overall survival ( p = 0.042). Transcript levels of Sox9 and CD24 were positively correlated. Silencing of Sox9 in HCC cells inhibited in vitro cell proliferation and tumorsphere formation, sensitized HCC cells to chemotherapeutic agents, and suppressed in vivo tumorigenicity. In addition, knockdown of Sox9 suppressed HCC cell migration, invasion, and in vivo lung metastasis. Further studies showed that Sox9 endowed stemness features through activation of Wnt/β-catenin signaling, which was confirmed by the partial rescue effect on tumorigenicity and self-renewal upon transfection of active β-catenin in Sox9 knockdown cells. By ChIP and luciferase promoter assays, Frizzled-7 was identified to be the direct transcriptional target of Sox9. In conclusion, Sox9 confers stemness properties of HCC through Frizzled-7 mediated Wnt/β-catenin pathway.