MBD1 is an Epigenetic Regulator of KEAP1 in Pancreatic Cancer

Background: MBD1 (Methyl-CpG Binding Domain Protein 1) is highly expressed in pancreatic cancer. Nrf2 (NF-E2 p45-related factor 2) and the ‘antioxidant response element’ (ARE)-driven genes that NRF2 controls are frequently upregulated in pancreatic cancer and correlate with poor survival. Keap1 (Kelch-like ECH-associated protein 1) is a dominant negative regulator of NRF2 and is reported to be epigenetically regulated by promoter methylation. However, the role of MBD1 with antioxidant response and its association with KEAP1 has never been reported before and remains unclear. Objective: We investigated the role of MBD1 in antioxidant response and its regulatory function in KEAP1 transcription in pancreatic cancer cells. Method: MBD1 was silenced to examine its role in antioxidant response. To explore the underlying mechanism, transcriptional and protein levels of KEAP1 was examined. The correlation between MBD1 and KEAP1 was confirmed in pancreatic cancer tissue samples by using immunohistochemistry (IHC). Dualluciferase reporter assay and Chromatin immunoprecipitation (ChIP) were used to elucidate he mechanism of MBD1 in KEAP1 transcriptional control. Moreover, co-immunoprecipitation (CoIP) assay was performed to uncover the regulatory role of MBD1 in KEAP1 transcription through its association with c-myc. Results: MBD1 silencing decreased antioxidant response and the related ARE target genes through epigenetic regulation of KEAP1. MBD1 negatively correlated with KEAP1 in pancreatic cancer tissue samples. Moreover, c-myc was a MBD1 interaction partner in KEAP1 epigenetic regulation. Conclusion: MBD1 can induce antioxidant response in pancreatic cancer through down-regulation of KEAP1. c-myc plays a key role in MBD1 mediated epigenetic silencing of KEAP1.