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Type: Journal article
Title: Amino acid substitutions in the aryl hydrocarbon receptor ligand binding domain reveal YH439 as an atypical AhR activator
Author: Whelan, F.
Hao, N.
Furness, S.
Whitelaw, M.
Chapman-Smith, A.
Citation: Molecular Pharmacology, 2010; 77(6):1037-1046
Publisher: Amer Soc Pharmacology Experimental Therapeutics
Issue Date: 2010
ISSN: 0026-895X
Statement of
Fiona Whelan, Nan Hao, Sebastian G. B. Furness, Murray L. Whitelaw and Anne Chapman-Smith
Abstract: The aryl hydrocarbon receptor (AhR) is traditionally defined as a transcription factor activated by exogenous polyaromatic and halogenated aromatic hydrocarbon (PAH/HAH) ligands. Active AhR induces genes involved in xenobiotic metabolism, including cytochrome P4501A1, which function to metabolize activating ligands. However, recent studies implicate AhR in biological events that are apparently unrelated to the xenobiotic response, implying that endogenous activation mechanisms exist. Three AhR genes in zebrafish (Danio rerio) encode proteins that demonstrate differential activation in response to PAH/HAHs, with the nonresponsive drAhR1a having some sequence divergence from the PAH/HAH-responsive AhRs in the ligand binding domain (LBD). We used these differences to guide the mutagenesis of mouse AhR (mAhR), aiming to generate variants that functionally discriminate between activation mechanisms. We found substitution of histidine 285 in the LBD with tyrosine gave a receptor that could be activated by isopropyl-2-(1,3-dithietane-2-ylidene)-2-[N-(4-methylthiazol-2-yl)carbamoyl]acetate (YH439), a potential AhR ligand chemically distinct from classic PAH/HAH-type ligands, but prevented activation by both exogenous PAH/HAH ligands and the endogenous activation mimics of suspension culture and application of shear-stressed serum. The differential response of H285Y mAhR to YH439 suggests that this activator has a novel mode of interaction that tolerates tyrosine at position 285 in the LBD and is distinct from the binding mode of the well characterized PAH/HAH ligands. In support of this, the PAH-type antagonist 3′,4′-dimethoxyflavone blocked mAhR activation by 2,3,7,8-tetrachlorodibenzo-p-dioxin but not YH439. Furthermore, the strict correlation between response to exogenous PAH/HAH ligands and mimics of endogenous activation suggests that a PAH-type ligand may underpin endogenous mechanisms of activation.
Keywords: Cell Line; Animals; Zebrafish; Humans; Receptors, Aryl Hydrocarbon; DNA Primers; Electrophoretic Mobility Shift Assay; Amino Acid Substitution; Polymerase Chain Reaction; Mutagenesis; Binding Sites; Amino Acid Sequence; Base Sequence; Sequence Homology, Amino Acid; Molecular Sequence Data
Rights: Copyright © 2010 by the American Society for Pharmacology and Experimental Therapeutics
RMID: 0020097629
DOI: 10.1124/mol.109.062927
Appears in Collections:Molecular and Biomedical Science publications

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