Involvement of regulatory and catalytic subunits of phosphoinositide 3-kinase in NF-kappaB activation.
Hypoxia, reoxygenation, and the tyrosine phosphatase inhibitor pervanadate activate the transcription factor NF-kappaB, involving phosphorylation of its inhibitor IkappaB-alpha on tyrosine 42. This modification does not lead to degradation of IkappaB by the proteasome/ubiquitin pathway, as is seen on stimulation of cells with proinflammatory cytokines. It is currently unknown how tyrosine-phosphorylated IkappaB is removed from NF-kappaB. Here we show that p85alpha, the regulatory subunit of PI3-kinase, specifically associates through its Src homology 2 domains with tyrosine-phosphorylated IkappaB-alpha in vitro and in vivo after stimulation of T cells with pervanadate. This association could provide a mechanism by which newly tyrosine-phosphorylated IkappaB is sequestered from NF-kappaB. Another mechanism by which PI3-kinase contributed to NF-kappaB activation in response to pervanadate appeared to involve its catalytic p110 subunit. This was evident from the inhibition of pervanadate-induced NF-kappaB activation and reporter gene induction by treatment of cells with nanomolar amounts of the PI3-kinase inhibitor wortmannin. The compound had virtually no effect on tumor necrosis factor- and interleukin-1-induced NF-kappaB activities. Wortmannin did not inhibit tyrosine phosphorylation of IkappaB-alpha or alter the stability of the PI3-kinase complex but inhibited Akt kinase activation in response to pervanadate. Our data suggest that both the regulatory and the catalytic subunit of PI3-kinase play a role in NF-kappaB activation by the tyrosine phosphorylation-dependent pathway.
Béraud
C
C
Tularik, Two Corporate Drive, South San Francisco, CA 94080, USA. beraud@cytokinetics.com
Henzel
W J
WJ
Baeuerle
P A
PA
D000730:Androstadienes / Q000494:pharmacology; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D020672:I-kappa B Proteins; D019169:Jurkat Cells; D016328:NF-kappa B / Q000378:metabolism*; D010455:Peptides / Q000378:metabolism; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000378:metabolism; D015533:Transcriptional Activation / Q000235:genetics; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology; D014638:Vanadates / Q000494:pharmacology; D000077191:Wortmannin; D018909:src Homology Domains / Q000235:genetics
D000730:Androstadienes; D004268:DNA-Binding Proteins; D004791:Enzyme Inhibitors; D020672:I-kappa B Proteins; D016328:NF-kappa B; D010455:Peptides; D014409:Tumor Necrosis Factor-alpha; C076715:pervanadate; D019000:Phosphotyrosine; D014638:Vanadates; D019869:Phosphatidylinositol 3-Kinases; D000077191:Wortmannin