An acute phase response factor/NF-kappa B site downstream of the junB gene that mediates responsiveness to interleukin-6 in a murine plasmacytoma.
The immediate early gene, junB, is induced by interleukin-6 (IL-6) in plasmacytomas. In order to identify enhancers that mediate this effect, we cloned upstream and downstream sequences flanking the gene into a luciferase reporter gene vector containing the junB promoter and evaluated the IL-6 inducibility of these sequences by transient expression in an IL-6-dependent plasmacytoma cell line. Although a 6.5-kilobase fragment of upstream flanking sequence did not increase the IL-6 inducibility of the junB promoter, a 222-base pair fragment was identified in 2.1 kilobases of down-stream flanking sequence that both up-regulates the promoter and confers inducibility by IL-6. Point mutation of an acute phase response factor (APRF) site within this region significantly reduced up-regulation of the promoter in cells grown continuously in IL-6, as well as inducibility upon restimulation of cells with IL-6 after withdrawal from the growth factor. Point mutation of an NF-kappa B site sharing five nucleotides with the APRF site reduced up-regulation of the promoter but not inducibility by IL-6, whereas mutation of two other NF-kappa B sites in the 222-base pair fragment had no effect on expression. Western blotting of nuclear proteins purified by DNA affinity chromatography revealed inducible binding of Stat3 and constitutive binding of NF-kappa B p65 to the APRF/NF-kappa B site.
Division of Monoclonal Antibodies, Center for Biologics Evaluation and Research, Bethesda, Maryland 20892, USA.
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Brown
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Ades
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Nordan
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004742:Enhancer Elements, Genetic; D015972:Gene Expression Regulation, Neoplastic / Q000502:physiology; D016758:Genes, jun; D015850:Interleukin-6 / Q000502:physiology*; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D016328:NF-kappa B / Q000378:metabolism*; D009687:Nuclear Proteins / Q000378:metabolism*; D009838:Oligodeoxyribonucleotides; D010954:Plasmacytoma / Q000235:genetics* / Q000378:metabolism; D011401:Promoter Regions, Genetic; D050796:STAT3 Transcription Factor; D015534:Trans-Activators
D004268:DNA-Binding Proteins; D015850:Interleukin-6; D016328:NF-kappa B; D009687:Nuclear Proteins; D009838:Oligodeoxyribonucleotides; D050796:STAT3 Transcription Factor; C494087:Stat3 protein, mouse; D015534:Trans-Activators