Expression of IkappaBalpha in the nucleus of human peripheral blood T lymphocytes.
According to current models the inhibitory capacity of I(kappa)B(alpha) would be mediated through the retention of Rel/NF-kappaB proteins in the cytosol. However, I(kappa)B(alpha) has also been detected in the nucleus of cell lines and when overexpressed by transient transfection. To gain better insight into the potential role of nuclear I(kappa)B(alpha) in a physiological context we have analysed its presence in the nucleus of human peripheral blood T lymphocytes (PBL). We demonstrate the nuclear localization of I(kappa)B(alpha) in PBL by different techniques: Western blot, indirect immunofluorescence and electron microscopy. Low levels of nuclear I(kappa)B(alpha) were detected in resting cells whereas a superinduction was obtained after PMA activation. The nuclear pool of I(kappa)B(alpha) showed a higher stability than cytosolic I(kappa)B(alpha) and was partially independent of the resynthesis of the protein. Unexpectedly, the presence of nuclear I(kappa)B(alpha) did not inhibit NF-kappaB binding to DNA and this phenomenon was not due to the presence of IkappaBbeta at the nuclear level. Immunoprecipitation experiments failed to demonstrate an association between nuclear I(kappa)B(alpha) and NF-kappaB proteins. Our results demonstrate that in resting and PMA-activated human PBL, I(kappa)B(alpha) is present in the nucleus in an apparently inactive form unable to disrupt NF-kappaB binding from DNA.
Laín de Lera
T
T
Servicio de Microbiología, Centro de Investigación, Hospital 12 de Octubre, Madrid, Spain.
Folgueira
L
L
Martín
A G
AG
Dargemont
C
C
Pedraza
M A
MA
Bermejo
M
M
Bonay
P
P
Fresno
M
M
Alcami
J
J
D001773:Blood Cells; D015153:Blotting, Western; D002467:Cell Nucleus / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000096:biosynthesis* / Q000235:genetics; D019084:Fluorescent Antibody Technique, Indirect; D006801:Humans; D020672:I-kappa B Proteins; D019169:Jurkat Cells / Q000378:metabolism; D008213:Lymphocyte Activation; D018613:Microscopy, Confocal; D008854:Microscopy, Electron; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D020569:Proto-Oncogene Proteins c-rel; D013601:T-Lymphocytes / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014158:Transcription, Genetic
D004268:DNA-Binding Proteins; D020672:I-kappa B Proteins; D016328:NF-kappa B; C000606204:NFKBIA protein, human; D011518:Proto-Oncogene Proteins; D020569:Proto-Oncogene Proteins c-rel; D000072000:NF-KappaB Inhibitor alpha; D004247:DNA; D013755:Tetradecanoylphorbol Acetate