Aryl hydrocarbon (Ah) receptor levels are selectively modulated by hsp90-associated immunophilin homolog XAP2

The aryl hydrocarbon receptor (AhR) is a ligand-inducible transcription fac tor that mediates biological responses to halogenated aromatic hydrocarbons . The unliganded AhR is a cytoplasmic, tetrameric complex consisting of the AhR ligand-binding subunit, a dimer of hsp90, and the hepatitis B virus X- associated protein 2 (XAP2). The role of XAP2 as a member of the AhR core c omplex is poorly understood. XAP2 shares significant homology with the immu nophilins FKBP12 and FKBP52, including a highly conserved, C-terminal, tetr atricopeptide repeat (TPR) domain. XAP2 forms a complex with hsp90 and the AhR but can also bind to both independently. This binding is mediated by th e conserved TPR domain. Single-point mutations in this region are sufficien t to disrupt the association of XAP2 with both the AhR and hsp90 in cells. Cotransfection of the AhR and XAP2 in COS-1 cells results in increased AhR levels compared with cells transfected with the AhR alone. In contrast, coe xpression of the AhR with the TPR containing proteins FKBP52, protein phosp hatase 5 (PP5), or XAP2 TPR-mutants deficient in binding to the AhR and hsp 90 does not affect AhR levels and coexpression of the AhR with the TPR doma in of PP5 results in AhR down-regulation. These results demonstrate that XA P2 is apparently unique among hsp90-binding proteins in its ability to enha nce AhR levels. A yellow fluorescent protein (YFP)-XAP2-FLAG was constructe d and biochemically characterized, and no loss of function was detected. YF P-XAP2-FLAG was transiently transfected into NIH 3T3 and was found to local ize in both the nucleus and the cytoplasm when visualized by fluorescence m icroscopy. Treatment of Hepa-1 cells with the hsp90-binding benzoquinone an samycin, geldanamycin, and the macrocyclic antifungal compound radicicol re sulted in AhR but not XAP2 or FKBP52 turnover. Taken together, these result s suggest that XAP2/hsp90 and FKBP52/hsp90 complexes are similar yet exhibi t unique functional specificity.