Bach1 competes with Nrf2 leading to negative regulation of the antioxidant response element (ARE)-mediated NAD(P)H:quinone oxidoreductase 1 gene expression and induction in response to antioxidants.
The antioxidant response element (ARE) and Nrf2 are known to regulate the expression and coordinated induction of genes encoding detoxifying enzymes including NAD(P)H:quinone oxidoreductase1 (NQO1) in response to antioxidants. In this report, we demonstrate that overexpression of the transcription factor Bach1 in Hep-G2 cells negatively regulated NQO1 gene expression and induction in response to antioxidant t-BHQ. Bandshift and supershift assays revealed that Bach1 binds to the ARE as a heterodimer with small Maf proteins but not as a homodimer or heterodimer with Nrf2. The transfection and ChIP assays revealed that Bach1 and Nrf2 competed with each other to regulate ARE-mediated gene expression. Heme, a negative regulator of Bach1 relieved the Bach1 repression of NQO1 gene expression in transfected cells. The transcription of Bach1 and Nrf2 did not change in response to t-BHQ. Immunofluorescence assays and Western blot analysis revealed that both Bach1 and Nrf2 localized in the cytoplasm and nucleus of the untreated cells. The treatment of cells with t-BHQ resulted in the nuclear accumulation of both Bach1 and Nrf2. Interestingly, the t-BHQ-induced nuclear accumulation of Bach1 was significantly delayed over that of Nrf2. These results led to the conclusion that a balance of Nrf2 versus Bach1 inside the nucleus influences up- or down-regulation of ARE-mediated gene expression. The results further suggest that antioxidant-induced delayed accumulation of Bach1 contributes to the down-regulation of ARE-regulated genes, presumably to reduce the antioxidant enzymes to normal levels.
Dhakshinamoorthy
Saravanakumar
S
Department of Pharmacology, Baylor College of Medicine, Houston, Texas 77030, USA.
Jain
Abhinav K
AK
Bloom
David A
DA
Jaiswal
Anil K
AK
D000975:Antioxidants / Q000378:metabolism*; D001483:Base Sequence; D050976:Basic-Leucine Zipper Transcription Factors; D001667:Binding, Competitive; D015152:Blotting, Northern; D015153:Blotting, Western; D002460:Cell Line; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism; D047369:Chromatin Immunoprecipitation; D003593:Cytoplasm / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D019281:Dimerization; D015536:Down-Regulation; D051856:Fanconi Anemia Complementation Group Proteins; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D006418:Heme / Q000737:chemistry; D006801:Humans; D007700:Kinetics; D008156:Luciferases / Q000378:metabolism; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D016660:NAD(P)H Dehydrogenase (Quinone) / Q000378:metabolism*; D051267:NF-E2-Related Factor 2; D010957:Plasmids / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D014176:Protein Biosynthesis; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D051257:Proto-Oncogene Proteins c-maf; D012333:RNA, Messenger / Q000378:metabolism; D020218:Response Elements; D013347:Subcellular Fractions; D013997:Time Factors; D015534:Trans-Activators / Q000378:metabolism*; D014157:Transcription Factors / Q000378:metabolism*; D014158:Transcription, Genetic; D014162:Transfection; D015854:Up-Regulation
D000975:Antioxidants; C102108:BACH1 protein, human; D050976:Basic-Leucine Zipper Transcription Factors; D004268:DNA-Binding Proteins; D051856:Fanconi Anemia Complementation Group Proteins; C495607:MAF protein, human; D051267:NF-E2-Related Factor 2; C495635:NFE2L2 protein, human; D011518:Proto-Oncogene Proteins; D051257:Proto-Oncogene Proteins c-maf; D012333:RNA, Messenger; D015534:Trans-Activators; D014157:Transcription Factors; D006418:Heme; D008156:Luciferases; D016660:NAD(P)H Dehydrogenase (Quinone); C447479:NQO1 protein, human