Transcriptional regulation of NF-E2 p45-related factor (NRF2) expression by the aryl hydrocarbon receptor-xenobiotic response element signaling pathway - Direct cross-talk between phase I and II drug-metabolizing enzymes

The aryl hydrocarbon receptor (AHR) and NF-E2 p45-related factor (NRF2) are two distinct transcription factors involved in the regulation of drug-metabolizing enzymes. Increasing evidence from several studies implies that AHR and NRF2 have direct links, but the molecular mechanism remains unknown. In this work we demonstrate for the first time that Nrf2 gene transcription is directly modulated by AHR activation. DNA sequence analyses of the mouse Nrf2 promoter revealed one xenobiotic response element (XRE)-like element (XREL1) located at -712 and two additional XRE-like elements located at +755 (XREL2) and +850 (XREL3). Functional analysis using luciferase assay showed that XREL1, XREL2, and XREL3 are all inducible by 2,3,7,8-tetrachlorodibenzo-p-dioxin treatment, with XREL2 being the most potent. The functionality of these XRE-like elements was further confirmed by mutagenesis and gel shift experiments. Finally, we used chromatin immuno-precipitation assay to show a direct binding of AHR to the Nrf2 promoter. Cells with silenced AHR expression using siRNA also lost NRF2 mRNA induction by 2,3,7,8-tetrachlorodibenzo-p-dioxin. These new data position NRF2- antioxidant response element downstream in the AHR-XRE pathway. Moreover, direct regulation of NRF2 by AHR contributes to couple phase I and II enzymes into an integrated system facilitating more effective xenobiotic and carcinogen detoxification.