Induction of neutrophil gelatinase-associated lipocalin expression by co-stimulation with interleukin-17 and tumor necrosis factor-alpha is controlled by IkappaB-zeta but neither by C/EBP-beta nor C/EBP-delta.
Neutrophil gelatinase-associated lipocalin (NGAL) is a siderophore-binding antimicrobial protein that is up-regulated in epithelial tissues during inflammation. We demonstrated previously that the gene encoding NGAL (LCN2) is strongly up-regulated by interleukin (IL)-1beta in an NF-kappaB-dependent manner but not by tumor necrosis factor (TNF)-alpha, another potent activator of NF-kappaB. This is due to an IL-1beta-specific synthesis of the NF-kappaB-binding co-factor IkappaB-zeta, which is essential for NGAL induction. We demonstrate here that NGAL is strongly induced by stimulation with TNF-alpha in the presence of IL-17, a pro-inflammatory cytokine produced by the newly discovered subset of CD4(+) T helper cells, T(H)-17. In contrast to the murine NGAL orthologue, 24p3/lipocalin 2, we found no requirement for C/EBP-beta or C/EBP-delta for NGAL induction by IL-17 and TNF-alpha as neither small interfering RNAs against the two C/EBP mRNAs nor mutation of the C/EBP sites in the LCN2 promoter abolished IL-17- and TNF-alpha-induced up-regulation of NGAL. NGAL induction is governed solely by NF-kappaB and its co-factor IkappaB-zeta. This was demonstrated by a pronounced reduction in the amount of NGAL mRNA and NGAL protein synthesized in cells treated with small interfering RNA against IkappaB-zeta and a total lack of activation of an LCN2 promoter construct with a mutated NF-kappaB site. As IL-17 stimulation stabilizes the IkappaB-zeta transcript, we propose a model where TNF-alpha induces activation and binding of NF-kappaB to the promoters of both NFKBIZ and LCN2 genes but induce only transcription of IkappaB-zeta. Co-stimulation with IL-17 leads to accumulation of IkappaB-zeta mRNA and IkappaB-zeta protein, which can bind to NF-kappaB on the LCN2 promoter and thus induce NGAL expression.
Karlsen
Joachim R
JR
Department of Hematology (93.2.2), Granulocyte Research Laboratory, University of Copenhagen, Rigshospitalet, 9 Blegdamsvej, DK-2100 Copenhagen, Denmark.
Borregaard
Niels
N
Cowland
Jack B
JB
D000209:Acute-Phase Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D048868:Adaptor Proteins, Signal Transducing; D022782:CCAAT-Enhancer-Binding Protein-beta / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D051743:CCAAT-Enhancer-Binding Protein-delta / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D024202:Electrophoretic Mobility Shift Assay; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D020672:I-kappa B Proteins; D020381:Interleukin-17 / Q000494:pharmacology*; D000071068:Lipocalin-2; D054834:Lipocalins / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D008156:Luciferases / Q000378:metabolism; D008175:Lung Neoplasms / Q000188:drug therapy* / Q000378:metabolism; D016328:NF-kappa B / Q000235:genetics / Q000378:metabolism; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D011518:Proto-Oncogene Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000494:pharmacology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection; D014407:Tumor Cells, Cultured; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
D000209:Acute-Phase Proteins; D048868:Adaptor Proteins, Signal Transducing; D022782:CCAAT-Enhancer-Binding Protein-beta; C545012:CEBPB protein, human; C497252:CEBPD protein, human; D020672:I-kappa B Proteins; D020381:Interleukin-17; C489506:LCN2 protein, human; D000071068:Lipocalin-2; D054834:Lipocalins; D016328:NF-kappa B; C417234:NFKBIZ protein, human; D009687:Nuclear Proteins; D011518:Proto-Oncogene Proteins; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D014409:Tumor Necrosis Factor-alpha; D051743:CCAAT-Enhancer-Binding Protein-delta; D008156:Luciferases