IFNs are pleiotropic cytokines that have been shown to be important regulators of cell growth. IFN-alpha has recently been recognized to harbor therapeutic potential in prevention and treatment of hepatocellular carcinoma (HCC). However, HCC cells respond differentially to IFN treatment, the mechanism of which is largely unknown. To address this issue, we analyzed the effect of IFN-alpha on different liver tumor cell lines. We found that growth inhibiting effects of IFN-alpha in hepatoma cells require PML-NB induction and, moreover, tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) expression on the mRNA and protein level. RNAi silencing of PML down-regulates TRAIL expression in hepatoma cells and correspondingly blocks IFN-alpha-induced apoptosis. In addition, PML-deficient primary hepatocytes fail to up-regulate TRAIL upon IFN-alpha-treatment in contrast to their wild-type counterparts. These data identify TRAIL as a novel downstream transcriptional target of PML-mediated apoptosis in hepatomas and suggest that PML and TRAIL play important roles in IFN-regulated apoptosis in HCC. Furthermore, the mechanism is independent of the p53 status of the tumor cells. In summary, our results identify central molecules mediating IFN-alpha induced apoptosis in liver tumors, shed light on the differential response of hepatoma cells to IFN exposure and, thus, may contribute to an efficient application of this substance in the treatment of liver cancer.