Function of DNA methyltransferase 3a in lead (Pb 2+ )‐Induced Cyclooxygenase‐2 gene

Lead ions (Pb 2+ ) are toxic industrial pollutants associated with chronic inflammatory diseases in humans and animals. Previously, we found that Pb 2+ ions induce COX‐ 2 gene expression via the EGF receptor/nuclear factor‐κB signal transduction pathway in epidermoid carcinoma cell line A431. In this study, to see whether Pb 2+ ions affect COX‐2 expression by epigenetic mechanisms, we looked at the mRNAs of DNA methyltransferases (DNMTs) using real‐time PCR of total RNA from these cells. Cells exposed to Pb 2+ had low levels of DNMT3a mRNA, whereas the levels of DNMT1 and DNMT3b mRNAs remained unchanged. Pretreatment of cells with DNMT inhibitor 5‐aza‐2'‐deoxycytidine (5 μM) followed by Pb 2+ (1 μM) significantly increased levels of COX‐2 mRNA compared with cells treated with Pb 2+ alone. Overexpression of tumor suppressor gene Rb correlated with an increase in COX‐2 mRNA and a decrease in DNMT3a mRNA. Conversely, overexpression of transcription factor E2F1 correlated with a decrease in COX‐2 mRNA and an increase in DMNT3a mRNA. Pretreatment with EGFR inhibitors AG1478 and PD153035 significantly limited Pb 2+ ‐induced reduction in DNMT3a mRNA. In addition, gene knockdown of DNMT3a with short hairpin RNA correlated with increased COX‐2 mRNA induced by Pb 2+ . Our findings suggest Pb 2+ ions induce COX‐ 2 expression indirectly by reducing DNMT3a methylation of the COX‐2 promoter via transcription factors Rb and E2F1. © 2014 Wiley Periodicals, Inc. Environ Toxicol 30: 1024–1032, 2015.