Abstract
The aryl hydrocarbon receptor (AHR) is a ligand-inducible transcription factor that displays interspecies differences with the human and mouse AHR C-terminal region sequences sharing only 58% amino acid sequence identity. Compared with the mouse AHR (mAHR), the human AHR (hAHR) displays ∼10-fold lower relative affinity for prototypical AHR ligands such as 2,3,7,8-tetrachlorodibenzo-p-dioxin, which has been attributed to the amino acid residue valine 381 (alanine 375 in the mAHR) in the ligand binding domain of the hAHR. We investigated whether the 10-fold difference in ligand-binding affinity between the mAHR and hAHR would be observed with a diverse range of AHR ligands. To test this hypothesis, ligand binding assays were performed using the photo-affinity ligand 2-azido-3-[125I]iodo-7,8-dibromodibenzo-p-dioxin and liver cytosol isolated from hepatocyte-specific transgenic hAHR mice and C57BL/6J mice. It is noteworthy that competitive ligand-binding assays revealed that, compared with the mAHR, the hAHR has a higher relative affinity for certain compounds, including indirubin [(2Z)-2,3-biindole-2,3 (1′H,1′H)-dione and quercetin (2-(3,4dihydroxyphenyl)-3,5,7-trihydroxy-4H-chromen-4-one]. Electrophoretic mobility shift assays revealed that indirubin was more efficient at transforming the hAHR compared with the mAHR. Indirubin was also a more potent inducer of Cyp1a1 expression in transgenic hAHR mouse hepatocytes compared with C57BL/6J mouse hepatocytes. These observations suggest that indirubin is a potent hAHR ligand that is able to selectively bind to and activate the hAHR. These discoveries imply that there may be a significant degree of structural divergence between mAHR and hAHR ligands and highlights the importance of the hAHR transgenic mouse as a model to study the hAHR in vivo.
Footnotes
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This work was supported by National Institutes of Health National Institute of Environmental Health Sciences [Grant ES04869]; the Natioanl Institutes of Health National Center for Research Resources [Grant 1C06-RR14520]; and the Dow Chemical Company.
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ABBREVIATIONS: AHR, aryl hydrocarbon receptor; PAL, 2-azido-3-[125I]iodo-7,8-dibromodibenzo-p-dioxin; EMSA, electrophoretic mobility shift assay; ARNT, aryl hydrocarbon receptor nuclear translocator; DRE, dioxin-responsive element; B[a]P, benzo[a]pyrene; TCDD, 2,3,7,8-tetrachlorodibenzo-p-dioxin; Ttr, transthyretin; PCR, polymerase chain reaction; RT-PCR, reverse transcriptase polymerase chain reaction; bp, base pair; MOPS, 3-(N-morpholino)propanesulfonic acid; PVDF, polyvinylidene difluoride; HEDG, HEPES/EDTA/sodium molybdate/glycerol; DLU, digitized light unit; PAGE, polyacrylamide gel electrophoresis.
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↵ The online version of this article (available at http://molpharm.aspetjournals.org) contains supplemental material.
- Received January 15, 2009.
- Accepted March 19, 2009.
- The American Society for Pharmacology and Experimental Therapeutics
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