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[Cancer Research 62, 5196-5203, September 15, 2002]
© 2002 American Association for Cancer Research


Carcinogenesis

Identification of Nrf2-regulated Genes Induced by the Chemopreventive Agent Sulforaphane by Oligonucleotide Microarray1

Rajesh K. Thimmulappa, Kim H. Mai, Sorachai Srisuma, Thomas W. Kensler, Masayuki Yamamoto and Shyam Biswal2

Department of Environmental Health Sciences, Bloomberg School of Public Health, Johns Hopkins University, Baltimore, Maryland 21205-2179 [R. K. T., K. H. M., S. S., T. W. K., S. B.], and Center for Tsukuba Advanced Research Alliance and Institute of Basic Medical Sciences, University of Tsukuba, Tsukuba, 305-8577 Japan [M. Y.]

Electrophiles formed during metabolic activation of chemical carcinogens and reactive oxygen species generated from endogenous and exogenous sources play a significant role in carcinogenesis. Cancer chemoprevention by induction of phase 2 proteins to counteract the insults of these reactive intermediates has gained considerable attention. Nuclear factor E2 p45-related factor 2 (Nrf2), a bZIP transcription factor, plays a central role in the regulation (basal and or inducible expression) of phase 2 genes by binding to the "antioxidant response element" in their promoters. Identification of novel Nrf2-regulated genes is likely to provide insight into cellular defense systems against the toxicities of electrophiles and oxidants and may define effective targets for achieving cancer chemoprevention. Sulforaphane is a promising chemopreventive agent that exerts its effect by strong induction of phase 2 enzymes via activation of Nrf2. In the present study, a transcriptional profile of small intestine of wild-type (nrf2 +/+) and knock out (nrf2 -/-) mice treated with vehicle or sulforaphane (9 µmol/day for 1 week, p.o.) was generated using the Murine Genome U74Av2 oligonucleotide array (representing ~6000 well-characterized genes and nearly 6000 expressed sequence tags). Comparative analysis of gene expression changes between different treatment groups of wild-type and nrf2-deficient mice facilitated identification of numerous genes regulated by Nrf2 including previously reported Nrf2-regulated genes such as NAD(P)H:quinone reductase (NQO1), glutathione S-transferase (GST), {gamma}-glutamylcysteine synthetase (GCS), UDP-glucuronosyltransferases (UGT),epoxide hydrolase, as well as a number of new genes. Also identified were genes encoding for cellular NADPH regenerating enzymes (glucose 6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase, and malic enzyme), various xenobiotic metabolizing enzymes, antioxidants (glutathione peroxidase, glutathione reductase, ferritin, and haptaglobin), and biosynthetic enzymes of the glutathione and glucuronidation conjugation pathways. The data were validated by Northern blot analysis and enzyme assays of selected genes. This investigation expands the horizon of Nrf2-regulated genes, highlights the cross-talk between various metabolic pathways, and divulges the pivotal role played by Nrf2 in regulating cellular defenses against carcinogens and other toxins.




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A. G. Banerjee, I. Bhattacharyya, and J. K. Vishwanatha
Identification of genes and molecular pathways involved in the progression of premalignant oral epithelia
Mol. Cancer Ther., June 1, 2005; 4(6): 865 - 875.
[Abstract] [Full Text] [PDF]


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Mol. Cell. Biol.Home page
S. H. Ki, I. J. Cho, D. W. Choi, and S. G. Kim
Glucocorticoid Receptor (GR)-Associated SMRT Binding to C/EBP{beta} TAD and Nrf2 Neh4/5: Role of SMRT Recruited to GR in GSTA2 Gene Repression
Mol. Cell. Biol., May 15, 2005; 25(10): 4150 - 4165.
[Abstract] [Full Text] [PDF]


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J. Biol. Chem.Home page
H. Chen, E. Rubin, H. Zhang, S. Chung, C. C. Jie, E. Garrett, S. Biswal, and S. Sukumar
Identification of Transcriptional Targets of HOXA5
J. Biol. Chem., May 13, 2005; 280(19): 19373 - 19380.
[Abstract] [Full Text] [PDF]


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Physiol. GenomicsHome page
J. Li, M. L. Spletter, and J. A. Johnson
Dissecting tBHQ induced ARE-driven gene expression through long and short oligonucleotide arrays
Physiol Genomics, March 21, 2005; 21(1): 43 - 58.
[Abstract] [Full Text] [PDF]


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Proc. Natl. Acad. Sci. USAHome page
M. J. Calkins, R. J. Jakel, D. A. Johnson, K. Chan, Y. W. Kan, and J. A. Johnson
Protection from mitochondrial complex II inhibition in vitro and in vivo by Nrf2-mediated transcription
PNAS, January 4, 2005; 102(1): 244 - 249.
[Abstract] [Full Text] [PDF]


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Mol. Cell. Biol.Home page
D. D. Zhang, S.-C. Lo, J. V. Cross, D. J. Templeton, and M. Hannink
Keap1 Is a Redox-Regulated Substrate Adaptor Protein for a Cul3-Dependent Ubiquitin Ligase Complex
Mol. Cell. Biol., December 15, 2004; 24(24): 10941 - 10953.
[Abstract] [Full Text] [PDF]


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J. Nutr.Home page
G. K. McWalter, L. G. Higgins, L. I. McLellan, C. J. Henderson, L. Song, P. J. Thornalley, K. Itoh, M. Yamamoto, and J. D. Hayes
Transcription Factor Nrf2 Is Essential for Induction of NAD(P)H:Quinone Oxidoreductase 1, Glutathione S-Transferases, and Glutamate Cysteine Ligase by Broccoli Seeds and Isothiocyanates
J. Nutr., December 1, 2004; 134(12): 3499S - 3506S.
[Abstract] [Full Text] [PDF]


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J AndrolHome page
W. Li, H. Amri, H. Huang, C. Wu, and V. Papadopoulos
Gene and Protein Profiling of the Response of MA-10 Leydig Tumor Cells to Human Chorionic Gonadotropin
J Androl, November 1, 2004; 25(6): 900 - 913.
[Abstract] [Full Text] [PDF]


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Cancer Res.Home page
I. Bae, S. Fan, Q. Meng, J. K. Rih, H. J. Kim, H. J. Kang, J. Xu, I. D. Goldberg, A. K. Jaiswal, and E. M. Rosen
BRCA1 Induces Antioxidant Gene Expression and Resistance to Oxidative Stress
Cancer Res., November 1, 2004; 64(21): 7893 - 7909.
[Abstract] [Full Text] [PDF]


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J. Immunol.Home page
N. Li, J. Alam, M. I. Venkatesan, A. Eiguren-Fernandez, D. Schmitz, E. Di Stefano, N. Slaughter, E. Killeen, X. Wang, A. Huang, et al.
Nrf2 Is a Key Transcription Factor That Regulates Antioxidant Defense in Macrophages and Epithelial Cells: Protecting against the Proinflammatory and Oxidizing Effects of Diesel Exhaust Chemicals
J. Immunol., September 1, 2004; 173(5): 3467 - 3481.
[Abstract] [Full Text] [PDF]


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CarcinogenesisHome page
V. Svehlikova, S. Wang, J. Jakubikova, G. Williamson, R. Mithen, and Y. Bao
Interactions between sulforaphane and apigenin in the induction of UGT1A1 and GSTA1 in CaCo-2 cells
Carcinogenesis, September 1, 2004; 25(9): 1629 - 1637.
[Abstract] [Full Text] [PDF]


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Cancer Res.Home page
M. C. Myzak, P. A. Karplus, F.-L. Chung, and R. H. Dashwood
A Novel Mechanism of Chemoprotection by Sulforaphane: Inhibition of Histone Deacetylase
Cancer Res., August 15, 2004; 64(16): 5767 - 5774.
[Abstract] [Full Text] [PDF]


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J. Pharmacol. Exp. Ther.Home page
R. Hu, V. Hebbar, B.-R. Kim, C. Chen, B. Winnik, B. Buckley, P. Soteropoulos, P. Tolias, R. P. Hart, and A.-N. T. Kong
In Vivo Pharmacokinetics and Regulation of Gene Expression Profiles by Isothiocyanate Sulforaphane in the Rat
J. Pharmacol. Exp. Ther., July 1, 2004; 310(1): 263 - 271.
[Abstract] [Full Text] [PDF]


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GENES CELLSHome page
T. Yanagawa, K. Itoh, J. Uwayama, Y. Shibata, A. Yamaguchi, T. Sano, T. Ishii, H. Yoshida, and M. Yamamoto
Nrf2 deficiency causes tooth decolourization due to iron transport disorder in enamel organ
Genes Cells, July 1, 2004; 9(7): 641 - 651.
[Abstract] [Full Text] [PDF]


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Molecular Cancer TherapeuticsHome page
Y. Zhang and G. B. Gordon
A strategy for cancer prevention: Stimulation of the Nrf2-ARE signaling pathway
Mol. Cancer Ther., July 1, 2004; 3(7): 885 - 893.
[Abstract] [Full Text] [PDF]


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Proc. Natl. Acad. Sci. USAHome page
J.-M. Lee, K. Chan, Y. W. Kan, and J. A. Johnson
Targeted disruption of Nrf2 causes regenerative immune-mediated hemolytic anemia
PNAS, June 29, 2004; 101(26): 9751 - 9756.
[Abstract] [Full Text] [PDF]


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J. Biol. Chem.Home page
S. V. Singh, A. Herman-Antosiewicz, A. V. Singh, K. L. Lew, S. K. Srivastava, R. Kamath, K. D. Brown, L. Zhang, and R. Baskaran
Sulforaphane-induced G2/M Phase Cell Cycle Arrest Involves Checkpoint Kinase 2-mediated Phosphorylation of Cell Division Cycle 25C
J. Biol. Chem., June 11, 2004; 279(24): 25813 - 25822.
[Abstract] [Full Text] [PDF]


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Toxicol SciHome page
W. H. M. Heijne, A. L. Slitt, P. J. van Bladeren, J. P. Groten, C. D. Klaassen, R. H. Stierum, and B. van Ommen
Bromobenzene-Induced Hepatotoxicity at the Transcriptome Level
Toxicol. Sci., June 1, 2004; 79(2): 411 - 422.
[Abstract] [Full Text] [PDF]


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J. Biol. Chem.Home page
B. G. Petrich, B. C. Eloff, D. L. Lerner, A. Kovacs, J. E. Saffitz, D. S. Rosenbaum, and Y. Wang
Targeted Activation of c-Jun N-terminal Kinase in Vivo Induces Restrictive Cardiomyopathy and Conduction Defects
J. Biol. Chem., April 9, 2004; 279(15): 15330 - 15338.
[Abstract] [Full Text] [PDF]


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CarcinogenesisHome page
S. Gebel, B. Gerstmayer, A. Bosio, H.-J. Haussmann, E. V. Miert, and T. Muller
Gene expression profiling in respiratory tissues from rats exposed to mainstream cigarette smoke
Carcinogenesis, February 1, 2004; 25(2): 169 - 178.
[Abstract] [Full Text] [PDF]


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CarcinogenesisHome page
A. V. Singh, D. Xiao, K. L. Lew, R. Dhir, and S. V. Singh
Sulforaphane induces caspase-mediated apoptosis in cultured PC-3 human prostate cancer cells and retards growth of PC-3 xenografts in vivo
Carcinogenesis, January 1, 2004; 25(1): 83 - 90.
[Abstract] [Full Text] [PDF]


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Mol. Cell. Biol.Home page
M.-K. Kwak, N. Wakabayashi, J. L. Greenlaw, M. Yamamoto, and T. W. Kensler
Antioxidants Enhance Mammalian Proteasome Expression through the Keap1-Nrf2 Signaling Pathway
Mol. Cell. Biol., December 1, 2003; 23(23): 8786 - 8794.
[Abstract] [Full Text] [PDF]


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Mol. Cell. Biol.Home page
D. D. Zhang and M. Hannink
Distinct Cysteine Residues in Keap1 Are Required for Keap1-Dependent Ubiquitination of Nrf2 and for Stabilization of Nrf2 by Chemopreventive Agents and Oxidative Stress
Mol. Cell. Biol., November 15, 2003; 23(22): 8137 - 8151.
[Abstract] [Full Text] [PDF]


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Mol. Pharmacol.Home page
I. R. Jowsey, Q. Jiang, K. Itoh, M. Yamamoto, and J. D. Hayes
Expression of the Aflatoxin B1-8,9-Epoxide-Metabolizing Murine Glutathione S-Transferase A3 Subunit Is Regulated by the Nrf2 Transcription Factor through an Antioxidant Response Element
Mol. Pharmacol., November 1, 2003; 64(5): 1018 - 1028.
[Abstract] [Full Text] [PDF]


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Nucleic Acids ResHome page
S. A. Haas, M. Hild, A. P. H. Wright, T. Hain, D. Talibi, and M. Vingron
Genome-scale design of PCR primers and long oligomers for DNA microarrays
Nucleic Acids Res., October 1, 2003; 31(19): 5576 - 5581.
[Abstract] [Full Text] [PDF]


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CarcinogenesisHome page
E.M.J. van der Logt, H.M.J. Roelofs, F.M. Nagengast, and W.H.M. Peters
Induction of rat hepatic and intestinal UDP-glucuronosyltransferases by naturally occurring dietary anticarcinogens
Carcinogenesis, October 1, 2003; 24(10): 1651 - 1656.
[Abstract] [Full Text] [PDF]


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J. Biol. Chem.Home page
J.-M. Lee, A. Y. Shih, T. H. Murphy, and J. A. Johnson
NF-E2-related Factor-2 Mediates Neuroprotection against Mitochondrial Complex I Inhibitors and Increased Concentrations of Intracellular Calcium in Primary Cortical Neurons
J. Biol. Chem., September 26, 2003; 278(39): 37948 - 37956.
[Abstract] [Full Text] [PDF]


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J. Nutr.Home page
K. J. Hintze, K. A. Wald, H. Zeng, E. H. Jeffery, and J. W. Finley
Thioredoxin Reductase in Human Hepatoma Cells Is Transcriptionally Regulated by Sulforaphane and Other Electrophiles via an Antioxidant Response Element-
J. Nutr., September 1, 2003; 133(9): 2721 - 2727.
[Abstract] [Full Text] [PDF]


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Am. J. Respir. Cell Mol. Bio.Home page
S. Srisuma, S. S. Biswal, W. A. Mitzner, S. J. Gallagher, K. H. Mai, and E. M. Wagner
Identification of Genes Promoting Angiogenesis in Mouse Lung by Transcriptional Profiling
Am. J. Respir. Cell Mol. Biol., August 1, 2003; 29(2): 172 - 179.
[Abstract] [Full Text] [PDF]


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Genes Dev.Home page
J. H. An and T. K. Blackwell
SKN-1 links C. elegans mesendodermal specification to a conserved oxidative stress response
Genes & Dev., August 1, 2003; 17(15): 1882 - 1893.
[Abstract] [Full Text] [PDF]


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J. Biol. Chem.Home page
M. McMahon, K. Itoh, M. Yamamoto, and J. D. Hayes
Keap1-dependent Proteasomal Degradation of Transcription Factor Nrf2 Contributes to the Negative Regulation of Antioxidant Response Element-driven Gene Expression
J. Biol. Chem., June 6, 2003; 278(24): 21592 - 21600.
[Abstract] [Full Text] [PDF]


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J. Neurosci.Home page
A. Y. Shih, D. A. Johnson, G. Wong, A. D. Kraft, L. Jiang, H. Erb, J. A. Johnson, and T. H. Murphy
Coordinate Regulation of Glutathione Biosynthesis and Release by Nrf2-Expressing Glia Potently Protects Neurons from Oxidative Stress
J. Neurosci., April 15, 2003; 23(8): 3394 - 3406.
[Abstract] [Full Text] [PDF]


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CarcinogenesisHome page
E. M. Ellis, C. M. Slattery, and J. D. Hayes
Characterization of the rat aflatoxin B1 aldehyde reductase gene, AKR7A1. Structure and chromosomal localization of AKR7A1 as well as identification of antioxidant response elements in the gene promoter
Carcinogenesis, April 1, 2003; 24(4): 727 - 737.
[Abstract] [Full Text] [PDF]


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J. Biol. Chem.Home page
J.-M. Lee, M. J. Calkins, K. Chan, Y. W. Kan, and J. A. Johnson
Identification of the NF-E2-related Factor-2-dependent Genes Conferring Protection against Oxidative Stress in Primary Cortical Astrocytes Using Oligonucleotide Microarray Analysis
J. Biol. Chem., March 28, 2003; 278(14): 12029 - 12038.
[Abstract] [Full Text] [PDF]


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J. Biol. Chem.Home page
M.-K. Kwak, N. Wakabayashi, K. Itoh, H. Motohashi, M. Yamamoto, and T. W. Kensler
Modulation of Gene Expression by Cancer Chemopreventive Dithiolethiones through the Keap1-Nrf2 Pathway. IDENTIFICATION OF NOVEL GENE CLUSTERS FOR CELL SURVIVAL
J. Biol. Chem., February 28, 2003; 278(10): 8135 - 8145.
[Abstract] [Full Text] [PDF]


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J. Biol. Chem.Home page
S. Allen, P. R. Heath, J. Kirby, S. B. Wharton, M. R. Cookson, F. M. Menzies, R. E. Banks, and P. J. Shaw
Analysis of the Cytosolic Proteome in a Cell Culture Model of Familial Amyotrophic Lateral Sclerosis Reveals Alterations to the Proteasome, Antioxidant Defenses, and Nitric Oxide Synthetic Pathways
J. Biol. Chem., February 14, 2003; 278(8): 6371 - 6383.
[Abstract] [Full Text] [PDF]


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J. Biol. Chem.Home page
E. C. Pietsch, J. Y. Chan, F. M. Torti, and S. V. Torti
Nrf2 Mediates the Induction of Ferritin H in Response to Xenobiotics and Cancer Chemopreventive Dithiolethiones
J. Biol. Chem., January 17, 2003; 278(4): 2361 - 2369.
[Abstract] [Full Text] [PDF]




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