Characterization of the Ah Receptor-associated Protein, ARA9

Abstract

The unliganded aryl hydrocarbon receptor (AHR) is found in a complex with other proteins including the 90-kDa heat shock protein (Hsp90) and a 37-kDa protein we refer to as ARA9. We found that the three tetratricopeptide repeats found in the COOH terminus of ARA9 are necessary and sufficient for interaction with the AHR complex. Conversely, the AHR's "repressor"/Hsp90 binding domain is required for interaction with ARA9. Because ARA9 closely resembles the 52-kDa FK506-binding protein (FKBP52), found in the unliganded glucocorticoid receptor (GR) complex, we compared the binding specificities of ARA9 and FKBP52 for AHR and GR. In co-immunoprecipitation experiments, ARA9 specifically associated with AHR-Hsp90 complex but not with GRHsp90 complexes. In addition, ARA9 showed a greater capacity than FKBP52 to associate with AHR-Hsp90 complexes. The biological importance of this interaction was suggested by the observation that in a yeast expression system ARA9 expression enhanced the response of AHR to the agonist ␤-napthoflavone, decreasing the EC 50 by greater than 5-fold and increasing the maximal response 2.5-fold. In contrast, co-expression of FKBP52 had no effect on AHR signaling. In addition, although ARA9 contains a domain similar to that found in other FK506-binding proteins, ARA9 binding to 3 H-FK506 could not be detected. Finally, we have characterized the developmental and expression pattern of ARA9 in the developing mouse embryo and mapped the ARA9 locus to human chromosome 11q13.3.

Epstein–Barr virus encoded nuclear protein EBNA-3 binds XAP-2, a protein associated with Hepatitis B virus X antigen

“…The three tetratricopeptide repeats found in the Cterminus of p38 are necessary and su cient for interaction with the AHR complex (Carver et al, 1998). The p38 subunit is identical with Xap-2, a cellular protein that also binds the N-terminus of Hepatitis B virus X antigen (Meyer et al, 1998).…”

The Aryl Hydrocarbon Receptor Complex and the Control of Gene Expression

“…177,179 In a yeast model system utilizing a β-galactosidase reporter gene to monitor AhR activity, the ED 50 of the dose-response curve is decreased by approximately fivefold when XAP2 is expressed along with a LEXA-AhR fusion protein. 177 When utilizing endogenous AhR levels in mouse hepatoma cells, however, XAP2 appears to cause a more modest (~ twofold) increase in ligandinduced AhR-driven luciferase activity, while the basal activity remains essentially unaltered. 175 XAP2-mediated enhancement of AhR activity is generally attributed to the elevated cytosolic AhR levels achieved in the presence of high amounts of XAP2, thereby increasing the available ligand-binding sites in a cell.…”

“…Its carboxy-terminal-most TPR displays homology to the TPRs in FKBP52, and is important in mediating the direct interaction between XAP2-AhR and XAP2-HSP90. 81,177 In addition to the TPR motif in XAP2, amino acid residues in the extreme carboxy-terminus of XAP2 are also needed to mediate its interaction with the AhR. Alanine substitution in any of the last four amino acids in XAP2 results in the complete loss of interaction with the AhR, while HSP90 binding is still observed even when the last five amino acids are completely deleted.…”

Phosphodiesterases link the aryl hydrocarbon receptor complex to cyclic nucleotide signaling

“…Another component of the AHR complex is XAP2 (also known as ARA9 or AIP), a 38-kDa protein that was initially identified as a protein binding to the hepatitis B virus X protein [41] and in parallel also as binding partner of AHR [42][43][44]. XAP2 shares sequence identity with FKBP52, an immunophilin of the family of FK506 binding proteins and an established component of the glucocorticoid receptor complex.…”

“…XAP2 shares sequence identity with FKBP52, an immunophilin of the family of FK506 binding proteins and an established component of the glucocorticoid receptor complex. Although XAP2 contains an immunophilin homology domain within its N-terminal region, no binding to immunosuppressant drugs was found [42]. The C-terminus of XAP2 contains multiple tetratricopeptide repeat (TPR) motifs that mediate protein-protein interactions [45].…”