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Aberrant activation of Wnt/β-catenin axis occurs in several gastrointestinal malignancies due to inactivating mutations of adenomatous polyposis coli (in colorectal cancer) or activating mutations of β-catenin itself [in hepatocellular carcinoma (HCC)]. These lead to β-catenin stabilization, increase in β-catenin/T-cell factor (TCF)-mediated transcriptional activation, and target gene expression, many of which are involved in tumor progression. While studying pharmaceutical agents that can target β-catenin in cancer cells, we observed that the plant compound berberine (BBR), a potent activator of AMP-activated protein kinase (AMPK), can reduce β-catenin expression and downstream signaling in HCC cells in a dose-dependent manner. More in-depth analyses to understand the mechanism revealed that BBR-induced reduction of β-catenin occurs independently of AMPK activation and does not involve transcriptional or post-translational mechanisms. Pretreatment with protein synthesis inhibitor cycloheximide antagonized BBR-induced β-catenin reduction, suggesting that BBR affects β-catenin translation. BBR treatment also antagonized mammalian target of rapamycin (mTOR) activity and was associated with increased recruitment of eukaryotic translation initiation factor 4E-binding protein (4E-BP) 1 in the translational complex, which was revealed by 7-methyl-cap-binding assays, suggesting inhibition of cap-dependent translation. Interestingly, knocking down 4E-BP1 and 4E-BP2 significantly attenuated BBR-induced reduction of β-catenin levels and expression of its downstream target genes. Moreover, cells with 4E-BP knockdown were resistant to BBR-induced cell death and were resensitized to BBR after pharmacological inhibition of β-catenin. Our findings indicate that BBR antagonizes β-catenin pathway by inhibiting β-catenin translation and mTOR activity and thereby reduces HCC cell survival. These also suggest that BBR could be used for targeting HCCs that express mutated/activated β-catenin variants that are currently undruggable. SIGNIFICANCE STATEMENT: β-catenin signaling is aberrantly activated in different gastrointestinal cancers, including hepatocellular carcinoma, which is currently undruggable. In this study we describe a novel mechanism of targeting β-catenin translation via utilizing a plant compound, berberine. Our findings provide a new avenue of targeting β-catenin axis in cancer, which can be utilized toward the designing of effective therapeutic strategies to combat β-catenin-dependent cancers.