Abstract
Nrf2 regulates expression of genes encoding enzymes with antioxidant (e.g. heme oxygenase-1 (HO-1)) or xenobiotic detoxification (e.g. NAD(P)H:quinone oxidoreductase, glutathione S-transferase) functions via the stress- or antioxidant-response elements (StRE/ARE). Nrf2 heterodimerizes with small Maf proteins, but the role of such dimers in gene induction is controversial, and other partners may exist. By using the yeast two-hybrid assay, we identified activating transcription factor (ATF) 4 as a potential Nrf2-interacting protein. Association between Nrf2 and ATF4 in mammalian cells was confirmed by co-immunoprecipitation and mammalian two-hybrid assays. Furthermore, Nrf2.ATF4 dimers bound to an StRE sequence from the ho-1 gene. CdCl(2), a potent inducer of HO-1, increased expression of ATF4 in mouse hepatoma cells, and detectable induction of ATF4 protein preceded that of HO-1 (30 min versus 2 h). A dominant-negative mutant of ATF4 inhibited basal and CdCl(2)-stimulated expression of a StRE-dependent/luciferase fusion construct (pE1-luc) in hepatoma cells but only basal expression in mammary epithelial MCF-7 cells. A dominant mutant of Nrf2 was equally inhibitory in both cell types in the presence or absence of CdCl(2). These results indicate that ATF4 regulates basal and CdCl(2)-induced expression of the ho-1 gene in a cell-specific manner and possibly in a complex with Nrf2.
Publication types
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Activating Transcription Factor 3
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Activating Transcription Factor 4
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Amino Acid Sequence
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Animals
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Breast
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Cadmium Chloride / pharmacology
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Cell Line
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Cloning, Molecular
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Conserved Sequence
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DNA-Binding Proteins / chemistry
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DNA-Binding Proteins / genetics
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DNA-Binding Proteins / metabolism*
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Dimerization
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Epithelial Cells
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Female
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Gene Expression Regulation / drug effects
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Gene Expression Regulation, Enzymologic*
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Genes, Reporter
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Heme Oxygenase (Decyclizing) / genetics*
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Heme Oxygenase-1
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Humans
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Leucine Zippers
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Liver Neoplasms, Experimental
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Membrane Proteins
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Mice
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Molecular Sequence Data
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NF-E2-Related Factor 2
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Rats
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Recombinant Proteins / chemistry
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Recombinant Proteins / metabolism
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Saccharomyces cerevisiae
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Sequence Alignment
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Sequence Homology, Amino Acid
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Trans-Activators / chemistry
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Trans-Activators / genetics
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Trans-Activators / metabolism*
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Transcription Factors / chemistry
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Transcription Factors / genetics*
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Transcription Factors / metabolism*
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Transcriptional Activation
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Transfection
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Tumor Cells, Cultured
Substances
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ATF4 protein, human
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Activating Transcription Factor 3
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Atf3 protein, rat
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Atf4 protein, rat
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DNA-Binding Proteins
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Membrane Proteins
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NF-E2-Related Factor 2
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NFE2L2 protein, human
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Nfe2l2 protein, mouse
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Nfe2l2 protein, rat
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Recombinant Proteins
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Trans-Activators
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Transcription Factors
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Activating Transcription Factor 4
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HMOX1 protein, human
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Heme Oxygenase (Decyclizing)
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Heme Oxygenase-1
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Hmox1 protein, mouse
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Cadmium Chloride