Inhibition of Ah (dioxin) receptor transformation by 9-hydroxy ellipticine. Involvement of protein kinase C?
9-Hydroxy ellipticine (9-OHE), a metabolite of the anti-neoplastic agent ellipticine, is known to bind the aryl hydrocarbon (Ah) receptor in rat lung cytosol and to inhibit aryl hydrocarbon hydroxylase activity (AHH) in rat hepatic microsomes. In this study, the effects of 9-OHE on the transformation of the rat hepatic cytosolic Ah receptor to a form that binds the xenobiotic responsive enhancer element-3 (XRE-3) of the cytochrome P4501A1 gene was investigated. Sucrose density gradient analysis of [3H]-2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) binding in rat hepatic cytosol indicated that 9-OHE inhibited binding of the radiolabeled ligand to the Ah receptor with an IC50 of 90 microM. Gel retardation assays revealed that at low concentrations of 9-OHE the Ah receptor bound to XRE-3, as was the case with the TCDD-liganded receptor. However, in the presence of high concentrations of 9-OHE, the Ah receptor failed to transform to a form that could bind to XRE-3. In vitro studies indicated that incubation of rat hepatic cytosol with TCDD resulted in concentration-dependent increases in levels of protein kinase C (PKC) mediated phosphorylation as compared to vehicle-treated extracts. Furthermore, 9-OHE concentrations that exhibited agonist activity with respect to Ah receptor transformation did not alter PKC phosphorylation in hepatic cytosol, whereas higher concentrations exhibited significant concentration-dependent decrease in PKC-mediated phosphorylation. These results demonstrate that the antagonistic effect of 9-OHE observed at high concentrations is due to inhibition of Ah receptor-XRE complex formation, a phenomenon that correlates with alterations in PKC activity.
Kurl
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Program in Clinical Pharmacology, Brown University School of Medicine, Providence, RI 02912.
DePetrillo
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Olnes
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MJ
D000818:Animals; D001483:Base Sequence; D001667:Binding, Competitive; D016384:Consensus Sequence; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics; D003600:Cytosol / Q000187:drug effects; D004611:Ellipticines / Q000494:pharmacology*; D004742:Enhancer Elements, Genetic; D008099:Liver / Q000187:drug effects* / Q000378:metabolism; D008297:Male; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D010766:Phosphorylation; D000072317:Polychlorinated Dibenzodioxins / Q000494:pharmacology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D018336:Receptors, Aryl Hydrocarbon / Q000037:antagonists & inhibitors / Q000187:drug effects*
D004611:Ellipticines; D009838:Oligodeoxyribonucleotides; D000072317:Polychlorinated Dibenzodioxins; D018336:Receptors, Aryl Hydrocarbon; D003577:Cytochrome P-450 Enzyme System; C008009:9-hydroxyellipticine; D011493:Protein Kinase C