Regulation of the promoter for human immunoglobulin gamma3 germ-line transcription and its interaction with the 3'alpha enhancer.
The mechanism underlying the differential regulation of switching to human IgG subclasses is still largely unknown. We demonstrate that the region upstream of the initiation sites for gamma3 germ-line (GL) transcripts contains a functional promoter which is synergistically induced by IL-4, antibody to CD40 and phorbol dibutyrate in transient transfection assays in the human DG75 cell line. Linker-scanning mutations identified multiple elements in the 3' half of the evolutionarily conserved sequence that are required for inducibility. Electrophoretic mobility shift assays showed that Stat6 and NF-kappaB p50 / p65 are induced after stimulation, and bind to specific sequence motifs within the promoter. Overexpression of Stat6, NF-kappaB p50 / p65 and C / EBPgamma synergistically induced the GL gamma3 promoter. Insertion of DNA segments from the human 3' IgH regions, which may function as a locus control region for switch recombination, greatly activated the promoter in an orientation-independent manner. Duplication of the enhancer fragments resulted in a further increase of promoter activity. The greater enhancement of the HS1,2 fragment from the 3' alpha1 rather than the alpha2 locus may suggest a mechanistic explanation for the differential expression of various isotypes.
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Division of Clinical Immunology, IMPI, Karolinska Institutet at Huddinge Hospital, Huddinge, Sweden.
Petit-Frére
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Stavnezer
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Hammarström
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D001402:B-Lymphocytes / Q000187:drug effects / Q000378:metabolism; D001483:Base Sequence; D022762:CCAAT-Enhancer-Binding Proteins; D019013:CD40 Antigens / Q000378:metabolism; D004247:DNA / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D004357:Drug Synergism; D004742:Enhancer Elements, Genetic / Q000235:genetics*; D005803:Genes, Immunoglobulin / Q000235:genetics; D006801:Humans; D017578:Immunoglobulin Class Switching / Q000235:genetics; D007074:Immunoglobulin G / Q000235:genetics*; D015847:Interleukin-4 / Q000494:pharmacology; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional / Q000235:genetics; D016328:NF-kappa B / Q000235:genetics / Q000378:metabolism; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology; D011401:Promoter Regions, Genetic / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020218:Response Elements / Q000235:genetics; D050800:STAT6 Transcription Factor; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects / Q000235:genetics*; D015533:Transcriptional Activation / Q000187:drug effects; D014407:Tumor Cells, Cultured
D022762:CCAAT-Enhancer-Binding Proteins; D019013:CD40 Antigens; D004268:DNA-Binding Proteins; D007074:Immunoglobulin G; D016328:NF-kappa B; D009687:Nuclear Proteins; D012333:RNA, Messenger; D050800:STAT6 Transcription Factor; C494101:STAT6 protein, human; D015534:Trans-Activators; D015847:Interleukin-4; D015240:Phorbol 12,13-Dibutyrate; D004247:DNA