Oxidant stress stimulates expression of the human peroxiredoxin 6 gene by a transcriptional mechanism involving an antioxidant response element.
Peroxiredoxin 6 (Prdx6) is a unique antioxidant enzyme that can reduce phospholipid and other hydroperoxides. A549 cells, a human lung-derived cell line, express both Prdx6 and Nrf2, a transcription factor that binds to antioxidant-response elements (AREs) and promotes expression of antioxidant genes. Treatment of A549 cells with 500 microM H(2)O(2) increased Prdx6 mRNA levels 2.5-fold, whereas treatment with 400 microM H(2)O(2) or 200 microM tert-butylhydroquinone (t-BHQ) triggered a corresponding 2.5-fold increase in reporter gene activity in A549 cells transfected with the pSEAP2:Basic vector (BD Bioscience), containing 1524 nucleotides of the human Prdx6 promoter region. Deletion of a consensus ARE sequence present between positions 357 and 349 before the start of transcription led to a striking decrease in both basal and H(2)O(2)- or t-BHQ-induced activation in A549 cells and H(2)O(2)-induced activation in primary rat alveolar type II cells. Cotransfection with Nrf2 stimulated the Prdx6 promoter in an ARE-dependent manner, whereas it was negatively regulated by Nrf3. siRNA targeting Nrf2 down-regulated reporter gene expression, whereas siRNA targeting the Nrf2 repressor, Keap1, up-regulated it. Binding of Nrf2 to the ARE sequence in chromatin was confirmed by PCR after chromatin immunoprecipitation. These data demonstrate that the ARE within the Prdx6 promoter is a key regulator of basal transcription of the Prdx6 gene and of its inducibility under conditions of oxidative stress.
Chowdhury
Ibrul
I
Institute for Environmental Medicine, University of Pennsylvania School of Medicine, 1 John Morgan Building, 3620 Hamilton Walk, Philadelphia, PA 19104-6068, USA.
Mo
Yiqun
Y
Gao
Ling
L
Kazi
Altaf
A
Fisher
Aron B
AB
Feinstein
Sheldon I
SI
D000818:Animals; D000975:Antioxidants / Q000378:metabolism*; D050976:Basic-Leucine Zipper Transcription Factors / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D042002:Chromatin Assembly and Disassembly; D004790:Enzyme Induction / Q000187:drug effects / Q000235:genetics; D004847:Epithelial Cells / Q000166:cytology / Q000502:physiology*; D025461:Feedback, Physiological; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D006873:Hydroquinones / Q000494:pharmacology; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism; D000072019:Kelch-Like ECH-Associated Protein 1; D008168:Lung / Q000166:cytology; D008297:Male; D051267:NF-E2-Related Factor 2 / Q000235:genetics / Q000378:metabolism; D018384:Oxidative Stress / Q000502:physiology*; D054465:Peroxiredoxin VI / Q000235:genetics / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D011485:Protein Binding / Q000187:drug effects / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics; D051381:Rats; D017207:Rats, Sprague-Dawley; D020218:Response Elements / Q000187:drug effects / Q000235:genetics; D017384:Sequence Deletion; D015533:Transcriptional Activation / Q000187:drug effects / Q000502:physiology*; D014162:Transfection
D000975:Antioxidants; D050976:Basic-Leucine Zipper Transcription Factors; D006873:Hydroquinones; D047908:Intracellular Signaling Peptides and Proteins; C431364:KEAP1 protein, human; D000072019:Kelch-Like ECH-Associated Protein 1; D051267:NF-E2-Related Factor 2; C117963:NFE2L3 protein, human; D034741:RNA, Small Interfering; D006861:Hydrogen Peroxide; C018855:2-tert-butylhydroquinone; D054465:Peroxiredoxin VI