Understanding the regulation of Nrf2 by NF-κB. (A) Schematic presentation of the Nrf2 promoter constructs that were created for this study, being either wild-type [a], κB2 site–deleted [b], or κB1 site–deleted [c] constructs. (B) THP-1 cells were transiently transfected with 0.5 μg of each promoter construct shown in panel A and pRL-CMV for normalization of transfection efficiency. Cell extracts were harvested, and luciferase assays were performed. Values are the means ± SD, n = 4. *P < .01 of deleted κB against untreated Nrf2 wild-type control. TNF (10 mg/mL) treatment acted as a positive control to activate NF-κB and BAY 11-7082 (10μM) to inhibit NF-κB. #P < .01 of TNF treated or BAY 11-7082 treated against untreated controls. (C) ChIP analysis of the Nrf2 promoter using antibodies against p50 and p65. Normal rabbit IgG was used as a control. THP-1 cells were left untreated or treated with BAY 11-7082 for 8 hours. Real-time PCR was performed in triplicate on immunoprecipitated DNA and input DNA. Data presented as percent of input. *P < .05 between the different treatment groups. (D) ChIP-seq data from the ENCODE Consortium demonstrates NF-κB subunit p65 binding to the Nrf2 promoter. NF-κB binding occurs in exon1 of Nrf2 and overlaps with sites of RNA pol II binding. Black bars denote positive signal above background.