IL-4-induced expression of the IL-1 receptor antagonist gene is mediated by STAT6.
IL-4 alone or in cooperation with LPS can induce the expression of the gene encoding the secreted-type IL-1 receptor antagonist (sIL-1ra) in mononuclear phagocytes. To determine the nuclear signaling mechanisms involved in this response, the region flanking the transcription start site of the human sIL-1ra gene was placed upstream of the luciferase reporter gene, and the function of specific sequence elements was analyzed following transient transfection in the macrophage-like cell line RAW264.7. A region located between -250 and -200 bases relative to the transcription start site was necessary for response to IL-4 alone and for cooperation between IL-4 and LPS. This 50-bp region contains two inverted repeat elements that represent potential binding sites for members of the signal transducer and activator of transcription (STAT) gene family (STAT-binding elements (SBEs)). Site-directed mutagenesis of the distal SBE abolished IL-4 responsiveness, and multiple copies of this motif were able to confer IL-4 sensitivity to luciferase expression in the context of a heterologous (herpes virus thymidine kinase) promoter. Mutation of the proximal SBE in the intact IL-1ra promoter had little or no effect on response to IL-4, and this sequence motif was inactive when examined alone. Electrophoretic mobility shift assays using an oligonucleotide corresponding to the distal SBE identified a single binding activity that was detected in nuclei within 15 min of IL-4 treatment and that was recognized by Ab to STAT6. These results indicate that IL-4-induced STAT6 is required for IL-4-induced transcriptional activation of the sIL-1ra gene.
Ohmori
Y
Y
Department of Immunology, Cleveland Clinic Foundation, Ohio 44195, USA.
Smith
M F
MF
Hamilton
T A
TA
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D004742:Enhancer Elements, Genetic / Q000276:immunology; D005786:Gene Expression Regulation / Q000187:drug effects*; D006801:Humans; D053590:Interleukin 1 Receptor Antagonist Protein; D007375:Interleukin-1 / Q000378:metabolism*; D015847:Interleukin-4 / Q000494:pharmacology*; D008264:Macrophages / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic / Q000276:immunology; D011485:Protein Binding / Q000235:genetics / Q000276:immunology; D018123:Receptors, Interleukin / Q000037:antagonists & inhibitors*; D050800:STAT6 Transcription Factor; D012795:Sialoglycoproteins / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D015398:Signal Transduction / Q000235:genetics / Q000276:immunology; D015534:Trans-Activators / Q000502:physiology*
C507763:IL1RN protein, human; C507764:Il1rn protein, mouse; D053590:Interleukin 1 Receptor Antagonist Protein; D007375:Interleukin-1; D018123:Receptors, Interleukin; D050800:STAT6 Transcription Factor; C494101:STAT6 protein, human; D012795:Sialoglycoproteins; C494100:Stat6 protein, mouse; D015534:Trans-Activators; D015847:Interleukin-4