Chelatable metal ions are not required for aryl hydrocarbon receptor transformation to a DNA binding form: phenanthrolines are possible competitive antagonists of 2,3,7,8-tetrachlorodibenzo-p-dioxin.
The aryl hydrocarbon receptor (AhR) mediates the toxicity of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and related compounds by binding DNA and altering gene transcription. We determined whether AhR transformation to a DNA binding form requires chelatable metal ions. The chelator 1,10-phenanthroline and its nonchelating isomers 1,7- and 4,7-phenanthroline blocked, in a concentration-dependent manner, TCDD-elicited transformation of the AhR in rat hepatic cytosol to a form which bound a dioxin-response element (DRE; upstream of the structural gene for cytochrome P4501A1). This was found to be due to the ability of these compounds to competitively inhibit [3H]TCDD specific binding to the AhR under conditions in vitro. EDTA (20 mM) failed to inhibit DRE binding of the transformed AhR, but pretreatment of cytosol with EDTA prior to transformation inhibited DRE binding up to 60%. However, removal of EDTA from the cytosol by gel filtration prior to incubation with TCDD resulted in the same DRE binding as filtered control cytosol without the added divalent metal ions. Both chelators, oxalic acid and iminodiacetic acid, failed to inhibit DRE binding when added prior to AhR transformation. Together these data indicate that chelatable metal ions are not required for AhR transformation to the DNA binding form.
Mahon
M J
MJ
Department of Biophysics, University of Rochester Medical Center, New York 14642.
Gasiewicz
T A
TA
D000818:Animals; D001483:Base Sequence; D001667:Binding, Competitive; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D003600:Cytosol / Q000378:metabolism; D004247:DNA / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004492:Edetic Acid / Q000494:pharmacology; D007700:Kinetics; D008099:Liver / Q000378:metabolism*; D008670:Metals / Q000494:pharmacology*; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides / Q000378:metabolism*; D010618:Phenanthrolines / Q000494:pharmacology*; D000072317:Polychlorinated Dibenzodioxins / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains; D018336:Receptors, Aryl Hydrocarbon; D011955:Receptors, Drug / Q000187:drug effects / Q000302:isolation & purification / Q000378:metabolism*; D013379:Substrate Specificity; D014158:Transcription, Genetic / Q000187:drug effects
D004268:DNA-Binding Proteins; D008670:Metals; D009838:Oligodeoxyribonucleotides; D010618:Phenanthrolines; D000072317:Polychlorinated Dibenzodioxins; D018336:Receptors, Aryl Hydrocarbon; D011955:Receptors, Drug; D004247:DNA; D004492:Edetic Acid