Protein kinase Calpha (PKCalpha) acts upstream of PKCtheta to activate IkappaB kinase and NF-kappaB in T lymphocytes.
NF-kappaB is an ubiquitous transcription factor that is a key in the regulation of the immune response and inflammation. T-cell receptor (TCR) cross-linking leads to NF-kappaB activation, an IkappaB kinase (IKK)-dependent process. However, the upstream kinases that regulate IKK activity following TCR activation remain to be fully characterized. Herein, we demonstrate using genetic analysis, pharmacological inhibition, and RNA interference (RNAi) that the conventional protein kinase C (PKC) isoform PKCalpha, but not PKCbeta1, is required for the activation of the IKK complex following T-cell activation triggered by CD3/CD28 cross-linking. We find that in the presence of Ca(2+) influx, the catalytically active PKCalphaA25E induces IKK activity and NF-kappaB-dependent transcription; which is abrogated following the mutations of two aspartates at positions 246 and 248, which are required for Ca(2+) binding to PKCalpha and cell membrane recruitment. Kinetic studies reveal that an early phase (1 to 5 min) of IKK activation following TCR/CD28 cross-linking is PKCalpha dependent and that a later phase (5 to 25 min) of IKK activation is PKCtheta dependent. Activation of IKK- and NF-kappaB-dependent transcription by PKCalphaA25E is abrogated by the PKCtheta inhibitor rottlerin or the expression of the kinase-inactive form of PKCtheta. Taken together, our results suggest that PKCalpha acts upstream of PKCtheta to activate the IKK complex and NF-kappaB in T lymphocytes following TCR activation.
Trushin
Sergey A
SA
Department of Immunology, Mayo Clinic, Rochester, Minnesota 55905, USA.
Pennington
Kevin N
KN
Carmona
Eva M
EM
Asin
Susana
S
Savoy
Doris N
DN
Billadeau
Daniel D
DD
Paya
Carlos V
CV
D000098:Acetophenones / Q000494:pharmacology; D001224:Aspartic Acid / Q000235:genetics; D001578:Benzopyrans / Q000494:pharmacology; D018106:CD28 Antigens / Q000378:metabolism; D017252:CD3 Complex / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D015971:Gene Expression Regulation, Enzymologic; D006801:Humans; D051550:I-kappa B Kinase; D007376:Interleukin-2 / Q000235:genetics; D007527:Isoenzymes / Q000187:drug effects / Q000378:metabolism*; D019169:Jurkat Cells; D008213:Lymphocyte Activation; D016328:NF-kappa B / Q000378:metabolism*; D017354:Point Mutation; D011493:Protein Kinase C / Q000187:drug effects / Q000378:metabolism*; D051571:Protein Kinase C-alpha; D000074121:Protein Kinase C-theta; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D013601:T-Lymphocytes / Q000201:enzymology / Q000378:metabolism*; D014158:Transcription, Genetic; D016335:Zinc Fingers
D000098:Acetophenones; D001578:Benzopyrans; D018106:CD28 Antigens; D017252:CD3 Complex; D004791:Enzyme Inhibitors; D007376:Interleukin-2; D007527:Isoenzymes; D016328:NF-kappa B; D001224:Aspartic Acid; C085746:rottlerin; D017346:Protein Serine-Threonine Kinases; C496559:CHUK protein, human; D051550:I-kappa B Kinase; C496560:IKBKB protein, human; C496565:IKBKE protein, human; C496628:PRKCA protein, human; C076233:PRKCQ protein, human; D011493:Protein Kinase C; D051571:Protein Kinase C-alpha; D000074121:Protein Kinase C-theta; D002118:Calcium