Regulation of rat glutathione S-transferase Ya subunit gene expression. DNA-protein interaction at the antioxidant responsive element.
We have characterized the interaction of the antioxidant responsive element (ARE) in the 5'-flanking region of the rat glutathione S-transferase Ya subunit gene with its trans-acting factor. The ARE core sequence, 5'-ggTGACaaaGC-3', previously identified as the cis-acting element required for activation of the Ya subunit gene by planar aromatic compounds and phenolic antioxidants, is shown to be the high affinity recognition motif for a trans-acting factor(s) as determined by gel mobility shift assays as well as methylation interference and protection studies. The DNA-protein interaction appears to occur in the major groove and involves the GpG dinucleotide preceding and the G residue within the TGAC tetramer on the coding strand of the core sequence. In addition, DNase I protection analysis maps an extended region 5' from the core recognition motif, which was shown previously to be essential for basal activity of the ARE. The trans-acting factor is present in nuclear extracts from untreated and tert-butylhydroquinone-treated cells as determined by photochemical cross-linking experiments. The cross-linked protein appears to be a heterodimer with subunit molecular weights of approximately 28,000 and approximately 45,000.
Nguyen
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Merck Frosst Center for Therapeutic Research, Pointe Claire-Dorval, Quebec, Canada.
Pickett
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CB
D000818:Animals; D001483:Base Sequence; D016172:DNA Fingerprinting; D004273:DNA, Neoplasm / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D015971:Gene Expression Regulation, Enzymologic; D005982:Glutathione Transferase / Q000235:genetics*; D008114:Liver Neoplasms, Experimental / Q000201:enzymology; D008745:Methylation; D008969:Molecular Sequence Data; D009711:Nucleotides / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D013379:Substrate Specificity; D014407:Tumor Cells, Cultured
D004273:DNA, Neoplasm; D004268:DNA-Binding Proteins; D009711:Nucleotides; D005982:Glutathione Transferase