Phosphorylation of RelA/p65 on serine 536 defines an I{kappa}B{alpha}-independent NF-{kappa}B pathway.
The association of the NF-kappaB p65/p50 dimer with IkappaBalpha plays a pivotal role in regulating its nuclear translocation and gene transcription. In addition, serine phosphorylation at various sites of the p65 subunit has been shown to be important in initiating transcription. Here we demonstrate that the regulation of nuclear translocation of p65 phosphorylated at serine 536 is not dependent on IkappaBalpha. Stimulation of either Jurkat or normal human T cells resulted in the nuclear translocation of phospho-p65 (Ser(536)). In addition, the phospho-p65 (Ser(536)) was not associated with either IkappaBalpha or p50, and the nuclear translocation of phospho-p65 (Ser(536)), but not total p65, was unaffected by the proteosome inhibitor MG-132, which blocks IkappaB protein degradation and prevents p65/p50 dimer nuclear translocation. Accordingly, the co-expression of a dominant negative mutant of IkappaBalpha blocked the transcriptional activity mediated by wild type but not the dominant positive p65 mutant (S536D). Furthermore, the transfection of the S536D form of p65 led to the induction of interleukin-8 transcription following stimulation, whereas the S536A form, which cannot be phosphorylated at this site, did not. Together, the findings suggest that p65 phosphorylated on serine 536 is not associated with or regulated by IkappaBalpha, that it has a distinct set of target genes, and that it may represent a noncanonical NF-kappaB pathway that is independent of IkappaBalpha regulation.
Sasaki
Carl Y
CY
Laboratory of Immunology, NIA, National Institutes of Health, Baltimore, Maryland 21224, USA. sasakic@grc.nia.nih.gov
Barberi
Theresa J
TJ
Ghosh
Paritosh
P
Longo
Dan L
DL
D021581:Active Transport, Cell Nucleus; D002467:Cell Nucleus / Q000378:metabolism; D047369:Chromatin Immunoprecipitation; D003593:Cytoplasm / Q000378:metabolism; D019281:Dimerization; D005786:Gene Expression Regulation; D005799:Genes, Dominant; D006367:HeLa Cells; D006801:Humans; D020672:I-kappa B Proteins / Q000378:metabolism*; D015151:Immunoblotting; D047468:Immunoprecipitation; D018799:Intercellular Adhesion Molecule-1 / Q000378:metabolism; D016209:Interleukin-8 / Q000378:metabolism; D019169:Jurkat Cells; D007976:Leupeptins / Q000494:pharmacology; D009154:Mutation; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000378:metabolism*; D010766:Phosphorylation; D010957:Plasmids / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011480:Protease Inhibitors / Q000494:pharmacology; D061988:Proteasome Inhibitors; D021381:Protein Transport; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012694:Serine / Q000737:chemistry*; D013997:Time Factors; D051996:Transcription Factor RelA / Q000737:chemistry / Q000378:metabolism*; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D014162:Transfection
D020672:I-kappa B Proteins; D016209:Interleukin-8; D007976:Leupeptins; D016328:NF-kappa B; C000606204:NFKBIA protein, human; D011480:Protease Inhibitors; D061988:Proteasome Inhibitors; D051996:Transcription Factor RelA; D018799:Intercellular Adhesion Molecule-1; D000072000:NF-KappaB Inhibitor alpha; D012694:Serine; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde