Binding of the Ah receptor to receptor binding factors in chromatin.
Dioxin induces biological responses through interaction with a specific intracellular receptor, the Ah receptor, and the subsequent interaction of the Ah receptor with chromatin. We report the binding of the Ah receptor, partially purified from rabbit liver, to receptor binding factors in chromatin. Rabbit liver chromatin proteins (CP) were isolated by adsorption of chromatin to hydroxylapatite followed by sequential extraction with 1-8 M GdnHCl. To assay for receptor binding a portion of each CP fraction was reconstituted to rabbit double-stranded DNA using a reverse gradient dialysis of 7.5 to 0 M GdnHCl. These reconstituted nucleoacidic proteins were then examined for binding to [3H]-2,3,7,8-tetrachlorodibenzo-p-dioxin ([3H]TCDD)-receptor complexes by the streptomycin filter assay. Prior to the binding assay, [3H]TCDD-receptor complexes were partially purified by step elution from DEAE-cellulose columns. CP fractions 2, 5, and 7 were found to bind to the Ah receptor with high affinity. Scatchard analysis yielded Kd values in the nanomolar range. Competition with 2-fold excess unlabeled TCDD-receptor complexes was demonstrated, and binding was reduced markedly when the receptor was prepared in the presence of 10 mM molybdate. Such chromatin receptor binding factors (RBFs) may participate in the interaction of receptor with specific DNA sequences resulting in modulation of specific gene expression.
Department of Pharmacological and Physiological Science, St. Louis University School of Medicine, MO 63104.
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D000818:Animals; D001665:Binding Sites; D002843:Chromatin / Q000378:metabolism*; D008099:Liver / Q000378:metabolism; D008982:Molybdenum / Q000494:pharmacology; D000072317:Polychlorinated Dibenzodioxins / Q000378:metabolism*; D011817:Rabbits; D018336:Receptors, Aryl Hydrocarbon; D011955:Receptors, Drug / Q000187:drug effects / Q000378:metabolism*
D002843:Chromatin; D000072317:Polychlorinated Dibenzodioxins; D018336:Receptors, Aryl Hydrocarbon; D011955:Receptors, Drug; C044659:molybdate; D008982:Molybdenum