Resistance to 2,3,7,8-Tetrachlorodibenzo-p-dioxin Toxicity and Abnormal Liver Development in Mice Carrying a Mutation in the Nuclear Localization Sequence of the Aryl Hydrocarbon Receptor

doi:10.1074/jbc.M209594200

Resistance to 2,3,7,8-Tetrachlorodibenzo-p-dioxin Toxicity and Abnormal Liver Development in Mice Carrying a Mutation in the Nuclear Localization Sequence of the Aryl Hydrocarbon Receptor*

From the ^‡McArdle Laboratory for Cancer Research and the ^§Training Program in Environmental Toxicology, University of Wisconsin Medical School, Madison, Wisconsin 53706

Abstract

The Ah receptor (AHR) mediates the metabolic adaptation to a number of planar aromatic chemicals. Essential steps in this adaptive mechanism include AHR binding of ligand in the cytosol, translocation of the receptor to the nucleus, dimerization with the Ah receptor nuclear translocator, and binding of this heterodimeric transcription factor to dioxin-responsive elements (DREs) upstream of promoters that regulate the expression of genes involved in xenobiotic metabolism. The AHR is also involved in other aspects of mammalian biology, such as the toxicity of molecules like 2,3,7,8-tetrachlorodibenzo-p-dioxin as well as regulation of normal liver development. In an effort to test whether these additional AHR-mediated processes require a nuclear event, such as DRE binding, we used homologous recombination to generate mice with a mutation in the AHR nuclear localization/DRE binding domain. These Ahr ^nls mice were found to be resistant to all 2,3,7,8-tetrachlorodibenzo-p-dioxin-induced toxic responses that we examined, including hepatomegaly, thymic involution, and cleft palate formation. Moreover, aberrations in liver development observed in these mice were identical to that observed in mice harboring a null allele at the Ahr locus. Taken in sum, these data support a model where most, if not all, of AHR-regulated biology requires nuclear localization.

Footnotes

↵* This work was supported by National Institutes of Health Grants P01-CA22484, P30-CA07175–CA14520, T32-CA09135, T32-ES07015, and F32-ES05877 and a fellowship from the Mary Engsberg Fund.The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵¶ To whom correspondence should be addressed: McArdle Laboratory for Cancer Research, 1400 University Ave., Madison, WI 53706. Tel.: 608-262-2024; Fax: 608-262-2824; E-mail: bradfield@oncology.wisc.edu.
Published, JBC Papers in Press, March 5, 2003, DOI 10.1074/jbc.M209594200
↵2 M. K. Bunger, S. M. Moran, E. Glover, T. L. Thomae, G. P. Lahvis, B. C. Lin, and C. A. Bradfield, unpublished observation.
Abbreviations:

AHR

aryl hydrocarbon receptor

Ahr^nls

the Ahr nls allele generated

AHR^nls

the AHR^nls protein that arises from the mutant allele

ARNT

aryl hydrocarbon receptor nuclear translocator

BEAR

bacterially expressed aryl hydrocarbon receptor

bHLH

basic helix-loop-helix

βNF

β-naphthoflavone

dpm

disintegrations/min

DRE

dioxin response element

DV

ductous venosus

EROD

ethoxyresorufin-o-de-ethylase

ES

embryonic stem

HSP90

90-kDa heat shock protein

NLS

nuclear localization signal

PAS

Period-ARNT-Singleminded

Rb

retinoblastoma

TCDD

2,3,7,8-tetrachlorodibenzo-p-dioxin

XME

xenobiotic metabolizing enzyme

MOPS

4-morpholinepropanesulfonic acid

[¹²⁵I]Br2DpD

2-azido-3-iodo[¹²⁵I]iodo-7,8-dibromodibenzo-p-dioxin

IVC

inferior vena cava
- Received September 18, 2002.
- Revision received February 27, 2003.
The American Society for Biochemistry and Molecular Biology, Inc.