PITX2C overexpression increased the stemness-related characteristics of HCC in in vitro and in vivo functional assays, while mechanistic studies revealed that PITX2C rather than PITX2A/B could activate TGF-β signaling by regulating the transcription of RALYL. Furthermore, PITX2C was found to regulate and cooperate with key factors related to hepatic differentiation and HCC tumorigenicity. The gene discussed is TGFB1; the disease is hepatocellular carcinoma.