Nrf2, a Cap’n’Collar Transcription Factor, Regulates Induction of the Heme Oxygenase-1 Gene

doi:10.1074/jbc.274.37.26071

Nrf2, a Cap’n’Collar Transcription Factor, Regulates Induction of the Heme Oxygenase-1 Gene*

From the ^‡Department of Molecular Genetics, Alton Ochsner Medical Foundation and ^§Department of Biochemistry and Molecular Biology, Louisiana State University Medical Center, New Orleans, Louisiana 70121 and the ^‖Section of Pulmonary and Critical Care Medicine, Yale University School of Medicine, New Haven, Connecticut 06250

Abstract

Stress response elements, which mediate induction of the mouse heme oxygenase-1 (HO-1) gene by several agents, resemble the binding site for the activator protein-1 (Jun/Fos), Maf, and Cap’n’Collar/basic leucine zipper (CNC-bZIP) families of proteins. In L929 fibroblasts, significant activation of an HO-1 enhancer-reporter fusion gene was observed only with the CNC-bZIP class of proteins with Nrf2 exhibiting the highest level oftrans-activation, between 25- and 30-fold. To further examine the role of this factor in HO-1 gene regulation, a dominant-negative mutant, Nrf2M, was generated and conditionally expressed in L929 cells. The mutant protein was detected in cytoplasmic and nuclear fractions but did not affect cell growth. Under conditions of Nrf2M overexpression, HO-1 mRNA accumulation in response to heme, cadmium, zinc, arsenite, andtert-butylhydroquinone was inhibited by 85–95%. In contrast, overexpression of a dominant-negative mutant of c-Jun decreased L929 cell growth but did not inhibit HO-1 gene activation. Nrf2 does not homodimerize, but CNC-bZIP·small Maf protein heterodimers and Nrf2·Jun protein complexes are proposed to function as trans-activators. Co-expression of Jun proteins or p18, however, had no significant affect or inhibited Nrf2-mediated trans-activation. Taken together, these results implicate Nrf2 in the induction of the HO-1 gene but suggest that the Nrf2 partner in this function is a factor other than p18 or Jun proteins.

Footnotes

↵* This work was supported by United States Public Health Service Grants DK-43135 and HL60234.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: Dept. of Molecular Genetics, Alton Ochsner Medical Foundation, 1516 Jefferson Hwy., New Orleans, LA 70121. Tel.: 504-842-3314; Fax: 504-842-3381; E-mail: jalam@ochsner.org.
↵2 J. Alam and D. Stewart, unpublished observations.
↵3 J. Alam, C. Dunne, D. Stewart, C. Touchard, S. Otterbein, and A. M. K. Choi, manuscript in preparation.
Abbreviations:

heme

ferriprotoporphyrin IX

HO-1

heme oxygenase-1

AP-1

activator protein-1

Nrf

NF-E2 related factor

NF-E2

nuclear factor-erythroid 2

CNC-bZIP

Cap’n’Collar/basic leucine zipper

CAT

chloramphenicol acetyltransferase

StRE

stress response element

ARE

anti-oxidant response element

Tc

tetracycline

TBHQ

tert-butylhydroquinone
- Received May 13, 1999.
- Revision received June 18, 1999.
The American Society for Biochemistry and Molecular Biology, Inc.