2,3,7,8-tetrachlorodibenzo-p-dioxin-induced degradation of aryl hydrocarbon receptor (AhR) by the ubiquitin-proteasome pathway. Role of the transcription activaton and DNA binding of AhR.
Activation of the aryl hydrocarbon receptor (AhR) by 2,3,7, 8-tetrachlorodibenzo-p-dioxin (TCDD), a potent agonist of AhR, induces a marked reduction in steady state AhR. To analyze the mechanism of regulation of ligand-activated AhR, we examined the biochemical pathway and function of the down-regulation of the receptor by TCDD. Pulse-chase experiments reveal that TCDD shortens the half-life (t1/2) of AhR from 28 to 3 h in mouse hepatoma cells. Inhibitors of the 26 S proteasome, lactacystin and MG132, block the TCDD-induced turnover of AhR. The TCDD-induced degradation of AhR involves ubiquitination of the AhR protein, because (a) TCDD induces formation of high molecular weight, ubiquitinated AhR and (b) degradation of AhR is inhibited in ts20 cells, which bear a temperature-sensitive mutation in the ubiquitin-activating enzyme E1, at a nonpermissive temperature. Inhibition of proteasomal degradation of AhR increases the amount of the nuclear AhR.Arnt complex and "superinduces" the expression of endogenous CYP1A1 gene by TCDD, indicating that the proteasomal degradation of AhR serves as a mechanism for controlling the activity of the activated receptor. We also show that deletion of the transcription activation domain of AhR abolishes the degradation, whereas a mutation in the DNA-binding region of AhR or Arnt reduces the degradation; these data implicate the transcription activation domain and DNA binding in AhR degradation. Our findings provide new insights into the regulation of TCDD-activated AhR through ubiquitin-mediated protein degradation.
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Molecular Toxicology Laboratory, Toxicology and Molecular Biology Branch, National Institute for Occupational Safety and Health, Centers for Disease Control and Prevention, Morgantown, West Virginia 26505, USA. qam1@cdc.gov
Baldwin
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D000818:Animals; D019363:Cytochrome P-450 CYP1A1 / Q000096:biosynthesis; D004268:DNA-Binding Proteins / Q000378:metabolism; D004790:Enzyme Induction; D006207:Half-Life; D008024:Ligands; D051379:Mice; D010447:Peptide Hydrolases / Q000187:drug effects / Q000378:metabolism*; D000072317:Polychlorinated Dibenzodioxins / Q000494:pharmacology*; D011480:Protease Inhibitors / Q000494:pharmacology; D046988:Proteasome Endopeptidase Complex; D011485:Protein Binding; D011499:Protein Processing, Post-Translational; D018336:Receptors, Aryl Hydrocarbon / Q000819:agonists* / Q000378:metabolism*; D014452:Ubiquitins / Q000378:metabolism*
D004268:DNA-Binding Proteins; D008024:Ligands; D000072317:Polychlorinated Dibenzodioxins; D011480:Protease Inhibitors; D018336:Receptors, Aryl Hydrocarbon; D014452:Ubiquitins; D019363:Cytochrome P-450 CYP1A1; D010447:Peptide Hydrolases; D046988:Proteasome Endopeptidase Complex; C061553:ATP dependent 26S protease