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J. Biol. Chem., Vol. 280, Issue 39, 33346-33356, September 30, 2005

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Redefining the Role of the Endogenous XAP2 and C-terminal hsp70-interacting Protein on the Endogenous Ah Receptors Expressed in Mouse and Rat Cell Lines^*

From the Department of Biology, University of South Florida, Tampa, Florida, 33620

Studies using transient expression systems have implicated the XAP2 protein in the control of aryl hydrocarbon receptor (AHR) stability and subcellular location. Thus, studies were performed in cell lines that expressed endogenous rat or mouse Ah^b-1 (C57BL/6) or Ah^b-2 (C3H) AHRs with similar levels of endogenous XAP2. Unliganded rat and mouse Ah^b-2 receptor complexes associated with reduced levels of XAP2 and exhibited dynamic nucleocytoplasmic shuttling in comparison with Ah^b-1 receptors. Rat and mouse Ah^b-2 receptors also exhibited a greater magnitude of ligand-induced degradation than Ah^b-1 receptors. Small interfering RNA reduction of endogenous XAP2 by >80% had minimal impact on the level of Ah^b-2 receptors but resulted in a 25–30% reduction of Ah^b-1 receptors. XAP2 reduction resulted in increased susceptibility of the Ah^b-1 receptor to ligand-induced degradation yet produced higher levels of endogenous CYP1A1 induction. Stable expression of the Ah^b-2 receptor in the C57BL/6 background resulted in a protein with reduced association with XAP2, dynamic nucleocytoplasmic shuttling, and increased levels of ligand-induced degradation. Small interfering RNA reduction of endogenous XAP2 in a C-terminal hsp70-interacting protein knockout mouse cell line, exhibited a 25–30% reduction in the level of endogenous Ah^b-1 AHR and showed high levels of ligand-induced degradation. Thus, endogenous XAP2 exerts a negative function on a small fraction of the endogenous Ah^b-1 receptor complex but appears to have a minimal impact on endogenous rat or Ah^b-2 receptors. This implies that the analysis of the AHR-mediated signaling via rat and mouse Ah^b-2 receptors may better represent the physiology of this signal transduction pathway.

Received for publication, June 17, 2005 , and in revised form, July 29, 2005.

^* This work was supported by National Institutes of Health Grant ES10991 (to R. S. P.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

¹ To whom correspondence should be addressed: University of South Florida, Dept. of Biology, BSF 110, 4202 E Fowler Ave., Tampa, FL 33620. Tel.: 813-974-1596; Fax: 813-974-3263; E-mail: pollenz{at}chuma1.cas.usf.edu.

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