Overexpression of DNA damage-induced 45 α gene contributes to esophageal squamous cell cancer by promoter hypomethylation

Background: Environmental factors-induced dysfunction of esophageal squamous epithelium, including genomic DNA impairment and apoptosis, play an important role in the pathogenesis of esophageal squamous cell cancer. DNA damage-induced 45 alpha (GADD45 alpha) has been found promoting DNA repair and removing methylation marker, Therefore, in this study we will investigate whether GADD45a expression is induced and its mechanism in esophageal squamous cell cancer. Methods: Two human esophageal squamous cell lines (ESCC), ECA109 and KYSE510 were cultured in RPMI-1640 medium supplemented with 10% fetal bovine serum (FBS). Lipofectamine 2000 was used to transfect cells. mRNA level of GADD45 alpha was measured by reverse transcription-quantitive PCR (RT-qPCR), protein level of GADD45 alpha was detected by western blot and Immunohistochemistry. Global DNA methylation of tissue sample was measured using the Methylamp Global DNA Methylation Quantification Ultra kit (Epigentek Group) and promoter methylation was measured by bisulfite sequencing. Results: GADD45a mRNA and protein levels were increased significantly in tumor tissue than that in adjacent normal tissue. Hypomethylation of global genomic DNA and GADD45 alpha promoter were found in ESCC. The cell sensitivity to Cisplatin DDP was decreased significantly in Eca109 and Kyse510 cells, in which GADD45 alpha expression was down-regulated by RNA interference (RNAi). In addition, silence of GADD45a expression in ESCC cells inhibited proliferation and promoted apoptosis. Conclusion: Overexpression of GADD45 alpha gene is due to DNA hypomethylation in ESCC. GADD45 alpha may be a protective factor in DDP chemotherapy for esophageal squamous cell carcinoma.