BackgroundHepatocellular carcinoma (HCC) is a leading cause of cancer-related deaths worldwide. Extensive research is currently directed at identifying novel targets for its diagnosis and treatment. AimsWe investigated the biological functions and clinical significance of mucin-type N-acetylglucosaminyltransferase 3 (GCNT3) in HCC. MethodsVariations in the mRNA expression of GCNT3 were examined in normal and HCC tissues. Cell function assays and animal models characterized the effects of GCNT3 on the proliferation, invasion, and migration abilities of HCC cells. Western blot and immunofluorescence analyses were performed to explore further the specific mechanisms whereby GCNT3 affects HCC progression. ResultsThere is a strong correlation between GCNT3 overexpression and tumor formation and metastasis in vivo and in vitro. GCNT3 acted as a regulator of the synthesis of mucin-type O-glycans by interacting with mucin 13 (MUC13) to regulate its expression levels, activating the GSK3β/β-catenin signaling pathway. The activation of GSK3β/β-catenin signaling by GCNT3 was mitigated by MUC13 knockdown. In clinical HCC specimens, GCNT3 expression was upregulated in HCC tissues compared to non-tumor tissues. Further, there was a significant correlation between high GCNT3 expression and poor patient survival. ConclusionsGCNT3 regulated tumor progression in HCC through the MUC13/GSK3-β/β-catenin signaling pathway.