Differential transcriptional activation in vitro by NF-kappa B/Rel proteins.
Distinct NF-kappa B subunit combinations contribute to the specificity of NF-kappa B-mediated transcriptional activation and to the induction of multiple cytokine genes including interferon-beta (IFN-beta). To evaluate the regulatory influence of different homo- and heterodimers, NF-kappa B subunits were analyzed for transcriptional activity in vitro using test templates containing two types of NF-kappa B recognition elements (the human immunodeficiency virus type 1 enhancer and the IFN-beta-positive regulatory domain-II (PRDII) as well as IFN-beta PRDIII-PRDI-PRDII linked to the -56 minimal promoter of rabbit beta-globin. Recombinant NF-kappa B subunits (p50, p65, c-Rel, p52, and I kappa B alpha) and interferon regulatory factor 1 were produced from either Escherichia coli or baculovirus expression systems. Transcriptional analysis in vitro demonstrated that 1) various dimeric complexes of NF-kappa B differentially stimulated transcription through the human immunodeficiency virus enhancer or PRDII up to 20-fold; 2) recombinant I kappa B alpha specifically inhibited NF-kappa B-dependent transcription in vitro; and 3) different NF-kappa B complexes and interferon regulatory factor 1 cooperated to stimulate transcription in vitro through the PRDIII-PRDI-PRDII virus-inducible regulatory domains of the IFN-beta promoter. These results demonstrate the role of NF-kappa B protein dimerization in differential transcriptional activation in vitro and emphasize the role of cooperativity between transcription factor families as an additional regulatory level to maintain transcriptional specificity.
Terry Fox Molecular Oncology Group, Lady Davis Institute for Medical Research, Jewish General Hospital, Montreal, Quebec, Canada.
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Lin
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Gewert
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Hiscott
D000818:Animals; D016367:Baculoviridae; D001483:Base Sequence; D003001:Cloning, Molecular; D016207:Cytokines / Q000096:biosynthesis*; D017931:DNA Primers; D004268:DNA-Binding Proteins / Q000378:metabolism; D004926:Escherichia coli; D015870:Gene Expression; D005786:Gene Expression Regulation; D005914:Globins / Q000235:genetics; D016322:HIV Enhancer; D015497:HIV-1 / Q000235:genetics; D020672:I-kappa B Proteins; D016899:Interferon-beta / Q000096:biosynthesis*; D008969:Molecular Sequence Data; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000037:antagonists & inhibitors / Q000378:metabolism*; D015345:Oligonucleotide Probes; D011401:Promoter Regions, Genetic; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D020569:Proto-Oncogene Proteins c-rel; D011817:Rabbits; D011994:Recombinant Proteins / Q000378:metabolism; D012045:Regulatory Sequences, Nucleic Acid; D015183:Restriction Mapping; D013698:Templates, Genetic; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic; D015533:Transcriptional Activation
D016207:Cytokines; D017931:DNA Primers; D004268:DNA-Binding Proteins; D020672:I-kappa B Proteins; D016328:NF-kappa B; C000606204:NFKBIA protein, human; D015345:Oligonucleotide Probes; D011518:Proto-Oncogene Proteins; D020569:Proto-Oncogene Proteins c-rel; D011994:Recombinant Proteins; D014157:Transcription Factors; D000072000:NF-KappaB Inhibitor alpha; D016899:Interferon-beta; D005914:Globins