The role of XPC protein deficiency in tobacco smoke-induced DNA hypermethylation of tumor suppressor genes

Abstract

DNA hypermethylation of tumor suppressor genes has been frequently observed in cancer patients, and therefore, may provide a valuable biomarker for cancer prevention and treatment. DNA hypermethylation may also provide an important mechanism in cancer progression. Lung cancer is strongly associated with exposure to environmental carcinogens, especially tobacco smoke. DNA damage generated by tobacco smoke is believed to play an important role in lung cancer development. XPC is a DNA damage recognition protein required for DNA repair and other DNA damage responses and attenuated XPC protein levels have been detected in many lung cancer patients. We studied the role of XPC protein deficiency in tobacco smoke-caused DNA hypermethylation of important tumor suppressor genes. Using both normal human fibroblasts (NF) and XPC-deficient hu man fibroblasts (XPC), our DNA methylation studies demonstrated that the XPC deficiency caused elevated levels of DNA hypermethylation in both Brca1 and Mlh1 tumor suppressor genes following exposure to tobacco smoke condensate (TSC). The results of our ChIP studies revealed that the XPC deficiency led to an increased binding of DNA methyltransferase 3A (DNMT3A) at the promoter region CpG island-containing sequences of these genes under the TSC treatment; however, this increase was partially diminished with prior treatment with caffeine. The results of our immuno-precipitation (IP) studies demonstrated a protein-protein interaction of the ATR with DNMT3A. Our western blots revealed that the XPC deficiency caused an increase in TSC-induced ATR phosphorylation at serine 428, an indicator of ATR activation. All these results suggest that XPC deficiency causes an accelerated DNA hypermethylation in important tumor suppressor genes under tobacco smoke exposure and activation of the ATR signaling pathway is involved in this DNA hypermethylation process.