Nuclear uptake control of NF-kappa B by MAD-3, an I kappa B protein present in the nucleus.
I kappa B proteins specifically inhibit the DNA binding of NF-kappa B/Rel transcription factors. An additional role as inhibitors of nuclear uptake was supposed by subcellular fractionation and enucleation experiments. Using indirect immunofluorescence labeling of cells, we show here that the DNA-binding p50 and p65 subunits of NF-kappa B, as well as the I kappa B protein MAD-3, all occur in the nucleus when overexpressed on their own. Nuclear uptake of p65 and, to a lesser extent of p50, was, however, suppressed when MAD-3 was coexpressed. Likewise, nuclear uptake of MAD-3 was blocked by overexpressed p65 or p50. This directly demonstrates that I kappa B is a nuclear uptake regulatory protein and that the various subunits of NF-kappa B can mutually control their access to the nucleus. In the presence of MAD-3, antibodies specific for peptides overlapping the nuclear location signal (NLS) sequences of p65 and p50 could not recognize their epitopes on NF-kappa B, suggesting that the I kappa B protein rendered the signals inaccessible for NLS receptors.
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Laboratory for Molecular Biology, Ludwig-Maximilians-University, Munich, Martinsried, Germany.
Henkel
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Silva
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Baeuerle
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PA
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies; D000952:Antigens, Polyomavirus Transforming / Q000032:analysis / Q000235:genetics; D001483:Base Sequence; D001692:Biological Transport; D002467:Cell Nucleus / Q000378:metabolism*; D003001:Cloning, Molecular; D004268:DNA-Binding Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D000939:Epitopes / Q000032:analysis; D004926:Escherichia coli / Q000235:genetics; D005455:Fluorescent Antibody Technique; D005822:Genetic Vectors; D020672:I-kappa B Proteins; D008969:Molecular Sequence Data; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000378:metabolism*; D009687:Nuclear Proteins / Q000378:metabolism; D009838:Oligodeoxyribonucleotides; D010957:Plasmids; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000032:analysis / Q000096:biosynthesis; D011994:Recombinant Proteins / Q000032:analysis / Q000378:metabolism; D013539:Simian virus 40 / Q000235:genetics; D013347:Subcellular Fractions / Q000378:metabolism / Q000648:ultrastructure; D014158:Transcription, Genetic; D014709:Vero Cells
D000906:Antibodies; D000952:Antigens, Polyomavirus Transforming; D004268:DNA-Binding Proteins; D000939:Epitopes; D020672:I-kappa B Proteins; D016328:NF-kappa B; D009687:Nuclear Proteins; D009838:Oligodeoxyribonucleotides; D012333:RNA, Messenger; D011994:Recombinant Proteins; D000072000:NF-KappaB Inhibitor alpha