T cell activation up-regulates the expression of the focal adhesion kinase Pyk2: opposing roles for the activation of protein kinase C and the increase in intracellular Ca2+.
T cell activation initiates signals that control gene expression of molecules important for T cell function. The focal adhesion kinase Pyk2 has been implicated in T cell signaling. To further analyze the involvement of Pyk2 in T cell processes, we examined the effect of T cell stimulation on the expression of Pyk2. We found that TCR ligation or PMA increased Pyk2 expression in Jurkat T cells and in normal T cells. In contrast, TCR ligation and PMA failed to induce any detectable increase in the expression of the other member of the focal adhesion kinase family, Fak, in Jurkat T cells and induced only a weak increase in Fak expression in normal T cells. The serine/threonine kinases, protein kinase C and mitogen-activated protein/extracellular signal-related kinase kinase (MEK), regulated Pyk2 expression, as inhibitors of these kinases blocked stimulus-induced Pyk2 expression. Cyclosporin A, FK506, and KN-62 did not block Pyk2 expression; thus, calcineurin and Ca2+/calmodulin-activated kinases are not critical for augmenting Pyk2 expression. TCR ligation increased Pyk2 mRNA, and the transcriptional inhibitor actinomycin D blocked Pyk2 expression. Strikingly, Ca2+ ionophores, at concentrations that in combination with other stimuli induced IL-2 expression, blocked TCR- and PMA-induced up-regulation of Pyk2 expression. Thus, the increase in Ca2+ has opposing effects on IL-2 and Pyk2 expression. Cyclosporin A and FK506, but not KN-62, blocked Ca2+ ionophore-mediated inhibition of Pyk2 expression, implicating calcineurin in down-regulating Pyk2 expression. These results show that TCR-triggered intracellular signals increase Pyk2 expression and shed light on the molecular mechanisms that regulate Pyk2 expression in T cells.
Tsuchida
M
M
Department of Surgery, Division of Transplantation, University of Wisconsin, Madison 53792, USA.
Manthei
E R
ER
Alam
T
T
Knechtle
S J
SJ
Hamawy
M M
MM
D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine / Q000031:analogs & derivatives / Q000494:pharmacology; D018106:CD28 Antigens / Q000502:physiology; D000001:Calcimycin / Q000037:antagonists & inhibitors / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000037:antagonists & inhibitors; D015815:Cell Adhesion Molecules / Q000096:biosynthesis; D016572:Cyclosporine / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects / Q000235:genetics / Q000276:immunology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D051417:Focal Adhesion Kinase 1; D051418:Focal Adhesion Kinase 2; D051416:Focal Adhesion Protein-Tyrosine Kinases; D006801:Humans; D007424:Intracellular Fluid / Q000378:metabolism*; D007476:Ionophores / Q000037:antagonists & inhibitors / Q000494:pharmacology; D019169:Jurkat Cells; D008213:Lymphocyte Activation / Q000187:drug effects / Q000276:immunology*; D048728:MAP Kinase Kinase Kinase 1; D020930:MAP Kinase Kinase Kinases / Q000502:physiology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism* / Q000502:physiology; D017346:Protein Serine-Threonine Kinases; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors / Q000096:biosynthesis* / Q000235:genetics / Q000502:physiology; D011948:Receptors, Antigen, T-Cell / Q000276:immunology / Q000378:metabolism / Q000502:physiology; D015398:Signal Transduction / Q000276:immunology; D013601:T-Lymphocytes / Q000187:drug effects / Q000201:enzymology / Q000276:immunology* / Q000378:metabolism; D016559:Tacrolimus / Q000494:pharmacology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014158:Transcription, Genetic / Q000276:immunology; D015854:Up-Regulation / Q000187:drug effects / Q000235:genetics / Q000276:immunology*
D018106:CD28 Antigens; D015815:Cell Adhesion Molecules; D004791:Enzyme Inhibitors; D007476:Ionophores; D011948:Receptors, Antigen, T-Cell; D000001:Calcimycin; C063302:KN 62; D016572:Cyclosporine; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D011505:Protein-Tyrosine Kinases; D051417:Focal Adhesion Kinase 1; D051418:Focal Adhesion Kinase 2; D051416:Focal Adhesion Protein-Tyrosine Kinases; C496050:PTK2 protein, human; D017346:Protein Serine-Threonine Kinases; D011493:Protein Kinase C; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D048728:MAP Kinase Kinase Kinase 1; D020930:MAP Kinase Kinase Kinases; C482576:MAP3K1 protein, human; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium; D016559:Tacrolimus