Epigenetic regulation of reactive oxygen species-induced tumor progression in hepatocellular carcinoma

Abstract

Oxidative stress is a consequence of an imbalance of antioxidants and reactive species. The most common form of reactive species is derived from oxygen, called reactive oxygen species (ROS). ROS involve in pathogenesis of several diseases including cancers. Cancer genesis and progression are contributed through both genetic and epigenetic mechanisms. Histone modification, a post-translational modification at histone tails, is one of the epigenetic mechanisms known to participate in carcinogenesis and tumor progression. Although the ROS-induced histone modification alteration has been demonstrated in some cancers, the change in histone methylation and gene expression by ROS in hepatocellular carcinoma (HCC) is scarcely reported. This study aimed to investigate the effect of ROS on tumor progression via histone modification in HCC cell lines. Expression of inactive chromatin (H4K20me3, H3K9me3) and active chromatin (H3K4me3) marks and their clinical significance were also investigated. The result showed that ROS promoted epithelial-mesenchymal transition (EMT) in HCC cells, indicated by reduced expression of E-cadherin, and enhanced expression of α-SMA and SNAIL. Cell migration, invasion, and colony formation were higher in HCC cells treated with ROS than the untreated controls. Co-treatment with antioxidant attenuated oxidative stress, inhibited EMT and decreased tumor progressivity in HCC cells exposed to H2O2. H4K20me3, H3K9me3, H3K4me3 and histone methyltransferases (SUV420H2, SUV39H1 and SMYD3) was upregulated in H2O2-treated HCC cells compared with the untreated controls. Chromatin immunoprecipitation-sequencing demonstrated that alteration of histone methylation (H4K20me3 and H3K4me3) by ROS was associated with upregulation of genes involved in DNA repair pathway (MRE11, BRCA1, MMS22L and RBBP8,) telomere maintenance (DCLRE1B, TERF1 and TERF2), and EMT pathway (SOS1 and RHOA) in HCC cells. The transcript expression of MRE11, BRCA1, MMS22L, RBBP8, DCLRE1B, TERF1 and TERF2 in HCC cells were increased following the H2O2 treatment. RBBP8 was selected to validate in the human HCC tissues, and it was overexpressed in the HCC tissues compared with the non-cancerous liver tissues. Expression of H4K20me3 was also higher in HCC tissues than the non-cancerous tissues. Elevated expression of H4K20me3 was associated with tumor recurrence and poor survival in HCC patients. In conclusion, ROS-induced oxidative stress promoted HCC progression through chromatin remodeling that subsequently upregulated genes related to EMT and DNA repairing pathways. H4K20me3 was upregulated in the HCC tissues, and its overexpression was clinically associated with the poor prognosis. The findings suggested that histone methylation, specifically H4K20me3, might be a promising marker for HCC prognosis, and it could be a target for development of HCC therapeutic agent in the future.