In vivo control of NF-kappa B activation by I kappa B alpha.
The transcription factor NF-kappa B is stored in the cytoplasm in complexes with the inhibitor protein I kappa B alpha. It has been shown in vitro that dissociation of I kappa B alpha from these complexes results in active NF-kappa B. In this report we show that lipopolysaccharide (LPS)-induced activation of B or pre-B cells results in loss of I kappa B alpha from NF-kappa B complexes in vivo. Many liberated NF-kappa B dimers reached the nucleus, where increased c-rel, p65 and p50 were detected by immunoblotting and by DNA binding assays. Some liberated dimers were retained in the cytoplasm, however, through binding to newly synthesized I kappa B alpha, a finding which strongly suggests (i) that the LPS-induced signal causes dissociation of complexes rather than preventing their association and (ii) that dissociation results from modification of I kappa B alpha and not of c-rel or p65. No effect of LPS treatment was detected on p105 or p100, which also retain rel family members in the cytoplasm. Quite unexpectedly, we also found that in unstimulated cells there is a constant ongoing process of degradation and replacement of complexed I kappa B alpha. We propose that this turnover results in the low level of active NF-kappa B presumably necessary even in the unstimulated cell, and that the high rate of synthesis of I kappa B alpha provides the ability to turn off NF-kappa B activity rapidly as soon as the activating signal ceases.
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Laboratory of Molecular Virology and Carcinogenesis, NCI-Frederick Cancer Research and Development Center, Frederick, MD 21702-1201.
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D000595:Amino Acid Sequence; D000818:Animals; D001402:B-Lymphocytes / Q000378:metabolism; D001483:Base Sequence; D002460:Cell Line; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D006207:Half-Life; D006801:Humans; D020672:I-kappa B Proteins; D015151:Immunoblotting; D008070:Lipopolysaccharides / Q000494:pharmacology; D051379:Mice; D008969:Molecular Sequence Data; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D020569:Proto-Oncogene Proteins c-rel; D014162:Transfection
D004268:DNA-Binding Proteins; D020672:I-kappa B Proteins; D008070:Lipopolysaccharides; D016328:NF-kappa B; C000606204:NFKBIA protein, human; C000606205:Nfkbia protein, mouse; D011518:Proto-Oncogene Proteins; D020569:Proto-Oncogene Proteins c-rel; D000072000:NF-KappaB Inhibitor alpha; D004247:DNA