Lipopolysaccharide-mediated induction of the bovine interleukin-6 gene in monocytes requires both NF-kappa B and C/EBP binding sites.
The interleukin-6 (IL-6) gene expression in bovine monocytes is highly induced following bacterial lipopolysaccharide (LPS) stimulation. To identify the promoter element(s) involved in the inducible transcription of IL-6, a 5'-flanking region containing 230 bp of the bovine IL-6 gene was linked to a reporter gene coding for bacterial chloramphenicol acetyltransferase (CAT) and analyzed for its ability to confer LPS-responsiveness to the reporter CAT gene in monocytic cells. Using mutant reporter genes, we demonstrate that although mutation in the NF-kappa B element produces the major loss of induction, both NF-kappa B and C/EBP elements are necessary for maximal transcriptional activation of the bovine IL-6 gene. Gel electrophoretic mobility-shift assays have detected induced DNA-binding activities in the LPS-stimulated monocytes. Further characterization has revealed the activation and interaction of C/EBP-alpha, C/EBP-beta (NF-IL6), NFKB1 (p50), and RelA (p65) to their specific binding elements present in the bovine IL-6 gene. These results suggest a model in which induction of C/EBP-alpha in differentiating monocytes contributes and synergizes with induced C/EBP-beta and NF-kappa B, which are activated following LPS stimulation, to mediate a high rate of IL-6 transcription under inflammatory conditions.
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Department of Veterinary Microbiology, University of Missouri, Columbia 65211, USA.
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D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D022762:CCAAT-Enhancer-Binding Proteins; D002417:Cattle; D015342:DNA Probes; D004268:DNA-Binding Proteins / Q000378:metabolism*; D015870:Gene Expression / Q000187:drug effects; D015850:Interleukin-6 / Q000235:genetics* / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology*; D008969:Molecular Sequence Data; D009000:Monocytes / Q000378:metabolism*; D016328:NF-kappa B / Q000378:metabolism*; D009687:Nuclear Proteins / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D014157:Transcription Factors / Q000378:metabolism*
D022762:CCAAT-Enhancer-Binding Proteins; D015342:DNA Probes; D004268:DNA-Binding Proteins; D015850:Interleukin-6; D008070:Lipopolysaccharides; D016328:NF-kappa B; D009687:Nuclear Proteins; D014157:Transcription Factors