Although Hepatitis B virus (HBV) X gene mutations are frequently detected in HBV-related human hepatocellular carcinoma (HCC) patients, causative HBx mutations in the development of HCC have not yet been determined. We herein identified C1485T and C1653T mutations in the HBx gene as independent risk of HCC for HBV through the analysis using serum from chronic hepatitis B patients. We generated transgenic mice expressing wild-type (WT-HBxTg) and mutant (C1485T-HBxTg) HBx to assess the carcinogenic potential of mutated HBx. C1485T-HBxTg mice were more susceptible to diethylnitrosamine-induced hepatocarcinogenesis than WT-HBxTg mice and control non-Tg mice. The promotion of hepatocarcinogenesis in C1485T-HBxTg mice was accompanied by the activation of β-catenin and Jun N-terminal kinase (JNK) signaling pathways as well as the production of reactive oxygen species, whereas the activation of nuclear factor-kappa B in the livers of C1485T-HBxTg mice was attenuated. These results demonstrate that the HBx C1485T mutation contributes to human and murine hepatocarcinogenesis.