ANT2 shRNA downregulates miR-19a and miR-96 through the PI3K/Akt pathway and suppresses tumor growth in hepatocellular carcinoma cells

MicroRNAs (miRNAs) are negative regulators of gene expression, and miRNA deregulation is found in various tumors. We previously reported that suppression of adenine nucleotide translocase 2 (ANT2) by short hairpin RNA (shRNA) inhibits hepatocellular carcinoma (HCC) development by rescuing miR-636 expression. However, the tumor-suppressive mechanisms of ANT2 shRNA are still poorly understood in HCC. Here, we hypothesized that miRNAs that are specifically downregulated by ANT2 shRNA might function as oncomiRs, and we investigated the roles of ANT2 shRNA-regulated miRNAs in the pathogenesis of HCC. Our data show that miR-19a and miR-96, whose expression is regulated by ANT2 suppression, were markedly upregulated in HCC cell lines and clinical samples. Ectopic expression of miR-19a and miR-96 dramatically induced the proliferation and colony formation of hepatoma cells in vitro, whereas inhibition of miR-19a and miR-96 reduced these effects. To investigate the in vivo function, we implanted miR-96-overexpressing HepG2 cells in a xenograft model and demonstrated that the increase in miR-96 promoted tumor growth. We also found that miR-19a and miR-96 inhibited expression of tissue inhibitor of metalloproteinase-2. Taken together, our results suggest that ANT2-regulated miR-19a and miR-96 play an important role in promoting the proliferation of human HCC cells, and the knockdown of ANT2 directly downregulates miR-19a and miR-96, ultimately resulting in the suppression of tumor growth.