PT - JOURNAL ARTICLE AU - Fiona Whelan AU - Nan Hao AU - Sebastian G Furness AU - Murray L Whitelaw AU - Anne Chapman-Smith TI - Amino acid substitutions in the Aryl Hydrocarbon Receptor (AhR) ligand binding domain reveal YH439 as an atypical AhR activator AID - 10.1124/mol.109.062927 DP - 2010 Mar 15 TA - Molecular Pharmacology PG - mol.109.062927 4099 - http://molpharm.aspetjournals.org/content/early/2010/03/15/mol.109.062927.short 4100 - http://molpharm.aspetjournals.org/content/early/2010/03/15/mol.109.062927.full AB - The Aryl hydrocarbon Receptor (AhR) is classically 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 metabolise activating ligands. However, recent studies implicate AhR in biological events which are apparently unrelated to the xenobiotic response, implying endogenous activation mechanisms exist. Three AhR genes in zebrafish (Danio rario) encode proteins which demonstrate differential xenobiotic activation potential in response to PAH/HAHs, with the non-responsive drAhR1a having some sequence divergence from the PAH/HAH-responsive AhRs in the ligand binding domain (LBD). We used these differences to guide mutagenesis of mouse AhR (mAhR), aiming to generate variants which functionally discriminate between activation mechanisms. We found substitution of histidine 285 in the LBD with tyrosine gave a receptor that could be activated by 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 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-characterised PAH/HAH ligands. In support of this, the PAH-type antagonist 3',4'-Dimethoxyflavone blocked mAhR activation by TCDD but not YH439. Further, the strict correlation between response to exogenous PAH/HAH ligands and mimics of endogenous activation suggests a PAH -type ligand may underpin endogenous mechanisms of activation.The American Society for Pharmacology and Experimental Therapeutics