The Ah receptor (AhR) is a ligand-dependent transcription factor known to regulate the toxic and biological effects of a variety of exogenous chemicals, such as the toxic halogenated aromatic hydrocarbons (HAHs) and polycyclic aromatic hydrocarbons (PAHs), and these effects appear to result from AhR-dependent alterations in gene expression. The AhR is also involved in several endogenous developmental and physiological processes, although the responsible endogenous ligand(s) is unknown. While HAHs and PAHs are the prototypical and highest affinity ligands, the AhR can bind and be activated by a diverse range of structurally dissimilar compounds, even though species- and ligand-specific differences in AhR ligand binding specificity and functionality exist. Site-directed mutagenesis and functional analysis studies based on our 3-dimensional (3D) homology model of the AhR ligand binding domain (LBD) performed so far have allowed initial understanding of aspects of the process by which high affinity HAH ligands like 2,3,7,8-tetrachlorodibenzo-p- dioxin (TCDD) can bind to and activate the AhR. However, these same studies suggest that significant differences exist in the amino acid residues to which structurally unrelated AhR ligands specifically interact. We hypothesize that differences in the binding sites and interactions of structurally diverse AhR ligands within the AhR LBD are primarily responsible for the observed ligand promiscuity of the AhR and that these differences could contribute to ligand-specific alterations in AhR conformational states that lead to differences in AhR functionality. To test this hypothesis, we propose to develop a new homology model of the AhR LBD based on recently released X-ray structures of the HIF-2a template complexed with ligands that also bind to the AhR and use this updated model for docking analysis of AhR ligands. Structurally driven site-directed mutagenesis and AhR functional analysis of the interactions of structurally diverse AhR agonists/antagonists with specific amino acids within the AhR LBD, coupled with similar analyses of chimeric mouse AhRs containing the LBD domain of AhRs which do not bind TCDD, will facilitate further identification of residues and structural characteristics of the LBD required for ligand binding. The molecular mechanisms by which binding of structurally diverse ligands within the LBD stimulates transformation of the AhR into its DNA binding form (loss of hsp90 and binding of Arnt) will be examined through analysis of AhR:hsp90 and AhR:Arnt interactions and ligand selective effects on coactivator binding and AhR-dependent gene expression determined. The residues/regions of both proteins involved in complex formation and functional activity will be defined and modeled and ligand-specific alterations in these mechanisms examined. The studies proposed here will provide detailed analysis of the molecular mechanisms by which structurally diverse ligands bind to and activate the AhR. In addition, they will yield insights into the molecular mechanisms of ligand-dependent AhR transformation, the influence of ligand structure on these processes and the diversity of AhR responsiveness.
Little is known about the molecular details and mechanisms by which structurally diverse exogenous and endogenous chemicals can to bind to and activate/inhibit the Ah (dioxin) receptor (AhR), a chemical-responsive cellular protein responsible for mediating the toxic, biological and developmental effects produced by these substances. The work proposed in this application will increase our understanding of the structure of the AhR and the mechanisms by which diverse chemicals can differentially activate or inhibit the AhR or AhR signaling pathways. These studies will not only lead to the identification and development of inhibitors of AhR-dependent toxicity with therapeutic potential (i.e. as anticancer agents and/or anti-toxins), but will provide avenues in which to gain insights into the normal physiological role(s) of this poorly understood receptor.
|Frauenstein, Katrin; Tigges, Julia; Soshilov, Anatoly A et al. (2015) Activation of the aryl hydrocarbon receptor by the widely used Src family kinase inhibitor 4-amino-5-(4-chlorophenyl)-7-(dimethylethyl)pyrazolo[3,4-d]pyrimidine (PP2). Arch Toxicol 89:1329-36|
|Soshilov, Anatoly A; Denison, Michael S (2014) Ligand promiscuity of aryl hydrocarbon receptor agonists and antagonists revealed by site-directed mutagenesis. Mol Cell Biol 34:1707-19|
|DeGroot, Danica E; Hayashi, Ai; Denison, Michael S (2014) Lack of ligand-selective binding of the aryl hydrocarbon receptor to putative DNA binding sites regulating expression of Bax and paraoxonase 1 genes. Arch Biochem Biophys 541:13-20|
|DeGroot, Danica E; Denison, Michael S (2014) Nucleotide specificity of DNA binding of the aryl hydrocarbon receptor:ARNT complex is unaffected by ligand structure. Toxicol Sci 137:102-13|
|Vogel, Christoph F A; Khan, Elaine M; Leung, Patrick S C et al. (2014) Cross-talk between aryl hydrocarbon receptor and the inflammatory response: a role for nuclear factor-*B. J Biol Chem 289:1866-75|
|Hu, Qin; He, Guochun; Zhao, Jing et al. (2013) Ginsenosides are novel naturally-occurring aryl hydrocarbon receptor ligands. PLoS One 8:e66258|
|Fraccalvieri, Domenico; Soshilov, Anatoly A; Karchner, Sibel I et al. (2013) Comparative analysis of homology models of the AH receptor ligand binding domain: verification of structure-function predictions by site-directed mutagenesis of a nonfunctional receptor. Biochemistry 52:714-25|
|Magiatis, Prokopios; Pappas, Periklis; Gaitanis, George et al. (2013) Malassezia yeasts produce a collection of exceptionally potent activators of the Ah (dioxin) receptor detected in diseased human skin. J Invest Dermatol 133:2023-30|
|Zhao, Bin; Bohonowych, Jessica E S; Timme-Laragy, Alicia et al. (2013) Common commercial and consumer products contain activators of the aryl hydrocarbon (dioxin) receptor. PLoS One 8:e56860|
|Pandini, Alessandro; Fraccalvieri, Domenico; Bonati, Laura (2013) Artificial neural networks for efficient clustering of conformational ensembles and their potential for medicinal chemistry. Curr Top Med Chem 13:642-51|
Showing the most recent 10 out of 54 publications