ARA9 Modifies Agonist Signaling through an Increase in Cytosolic Aryl Hydrocarbon Receptor

John J Lapres,Edward Glover,Elizabeth E Dunham,Maureen K Bunger,Christopher A Bradfield

doi:10.1074/jbc.275.9.6153

John J Lapres, Edward Glover + Show 3 more

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https://doi.org/10.1074/jbc.275.9.6153

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Abstract

The aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor that mediates the effects of agonists like 2,3, 7,8-tetrachlorodibenzo-p-dioxin. In the current model for AHR signaling, the unliganded receptor is found in the cytosol as part of a complex with a dimer of the 90-kDa heat shock protein and an immunophilin-like molecule, ARA9. In yeast, expression of ARA9 results in an increase in the maximal agonist response and a leftward shift in the AHR dose-response curve. To better understand the mechanism by which ARA9 modifies AHR signal transduction, we performed a series of coexpression experiments in yeast and mammalian cells. Our results demonstrate that ARA9's influence on AHR signaling is not due to inhibition of a membrane pump or modification of the receptor's transactivation properties. Using receptor photoaffinity labeling experiments, we were able to show that ARA9 enhances AHR signal transduction by increasing the available AHR binding sites within the cytosolic compartment of the cell. Our evidence suggests that ARA9's effects are related to its role as a cellular chaperone; i.e. we observed that expression of ARA9 increases the fraction of AHR in the cytosol and also stabilized the receptor under heat stress.

Highlights

The AHR1 is a ligand-activated transcription factor that mediates the biological effects of halogenated dioxins and related compounds [1]
Our previous work in yeast has shown that ARA9 is capable of modifying aryl hydrocarbon receptor (AHR) signaling by shifting the ␤NF dose-response curve of AHR to the left and increasing the maximal response [13]
Two other laboratories have shown that at a fixed dose of ligand, ARA9 can increase AHR signaling in mammalian cells [11, 12]

Summary

Introduction

The AHR1 is a ligand-activated transcription factor that mediates the biological effects of halogenated dioxins and related compounds [1]. ARA9 contains three TPRs. TPRs are defined by strings of 34 amino acids that have been shown to play roles in protein-protein interaction [14]. TPRs are defined by strings of 34 amino acids that have been shown to play roles in protein-protein interaction [14] This domain structure is similar to that found in the GR-associated immunophilin, FKBP52, which contains two FKBP domains in its amino terminus and three TPRs in its carboxyl terminus [15, 16]. In addition to their structural similarities, ARA9 and FKBP52 are found associated to the cytosolic complexes of AHR and the GR or progesterone receptor, respectively [9, 11, 12, 17, 18] In both mammalian cells and yeast, the presence of ARA9 enhances AHR signaling [11,12,13]. We employ a Gal4-AHRN⌬166 chimera that allowed us to assess the functional consequences of ARA9 expression on AHR signaling in mammalian cells

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