ARF6 promotes the nuclear translocation of β-catenin to facilitate AFP deficiency-mediated metastasis in CTNNB1

The mortality rate associated with hepatocellular carcinoma (HCC) remains alarmingly high, positioning it as a primary lethal cancer worldwide, primarily driven by its high degree of malignancy, recurrence, and metastasis. The high heterogeneity of HCC necessitates the urgent identification of new therapeutic targets and molecular pathways. Our previous research observed that alpha-fetoprotein (AFP) deficiency led to pro-metastatic roles in HCC with specific genotypes. Here, we further investigated the underlying molecular mechanism. Through proteome microarray screening, we identified that ADP-ribosylation factor 6 (ARF6) was a key AFP-interacting protein and AFP deficiency upregulated ARF6 activity, which subsequently promoted the metastatic capacity of HepG2 cells. Mechanistically, the abnormal activation of ARF6 induced the dissociation of β-catenin from its membrane-bound anchors, specifically E-cadherin. The consequent increase in free cytoplasmic β-catenin facilitates its nuclear translocation, which in turn activates Wnt/β-catenin pathway. Crucially, we discovered that the pro-metastatic function of activated ARF6 and the sensitivity to the ARF6 inhibitor were strictly dependent on the CTNNB1 genotype. In highly invasive MHCC-97H cells, which harbor a CTNNB1 nonsense mutation, ARF6 inhibitor SecinH3 failed to inhibit migration and invasion. However, the overexpression of CTNNB1