title: Zap-70-independent Ca(2+) mobilization and Erk activation in Jurkat T cells in response to T-cell antigen receptor ligation.
abstract: The tyrosine kinase ZAP-70 has been implicated as a critical intermediary between T-cell antigen receptor (TCR) stimulation and Erk activation on the basis of the ability of dominant negative ZAP-70 to inhibit TCR-stimulated Erk activation, and the reported inability of anti-CD3 antibodies to activate Erk in ZAP-70-negative Jurkat cells. However, Erk is activated in T cells receiving a partial agonist signal, despite failing to activate ZAP-70. This discrepancy led us to reanalyze the ZAP-70-negative Jurkat T-cell line P116 for its ability to support Erk activation in response to TCR/CD3 stimulation. Erk was activated by CD3 cross-linking in P116 cells. However, this response required a higher concentration of anti-CD3 antibody and was delayed and transient compared to that in Jurkat T cells. Activation of Raf-1 and MEK-1 was coincident with Erk activation. Remarkably, the time course of Ras activation was comparable in the two cell lines, despite proceeding in the absence of LAT tyrosine phosphorylation in the P116 cells. CD3 stimulation of P116 cells also induced tyrosine phosphorylation of phospholipase C-gamma1 (PLCgamma1) and increased the intracellular Ca(2+) concentration. Protein kinase C (PKC) inhibitors blocked CD3-stimulated Erk activation in P116 cells, while parental Jurkat cells were refractory to PKC inhibition. The physiologic relevance of these signaling events is further supported by the finding of PLCgamma1 tyrosine phosphorylation, Erk activation, and CD69 upregulation in P116 cells on stimulation with superantigen and antigen-presenting cells. These results demonstrate the existence of two pathways leading to TCR-stimulated Erk activation in Jurkat T cells: a ZAP-70-independent pathway requiring PKC and a ZAP-70-dependent pathway that is PKC independent.
authors.affiliation: Laboratory of Cellular and Molecular Biology, National Institute on Aging, National Institutes of Health, Baltimore, Maryland 21224-6825, USA.
authors.forename: X
authors.initials: X
authors.lastname: Shan
authors.forename: R
authors.initials: R
authors.lastname: Balakir
authors.forename: G
authors.initials: G
authors.lastname: Criado
authors.forename: J S
authors.initials: JS
authors.lastname: Wood
authors.forename: M C
authors.initials: MC
authors.lastname: Seminario
authors.forename: J
authors.initials: J
authors.lastname: Madrenas
authors.forename: R L
authors.initials: RL
authors.lastname: Wange
mesh_terms: D017252:CD3 Complex / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D003432:Cross-Linking Reagents / Q000494:pharmacology; D004268:DNA-Binding Proteins / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005434:Flow Cytometry; D017930:Genes, Reporter; D006801:Humans; D007527:Isoenzymes / Q000378:metabolism; D019169:Jurkat Cells; D007700:Kinetics; D048369:MAP Kinase Kinase 1; D020929:Mitogen-Activated Protein Kinase Kinases / Q000378:metabolism; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D008954:Models, Biological; D050778:NFATC Transcription Factors; D009687:Nuclear Proteins; D051966:Phospholipase C gamma; D010766:Phosphorylation; D011233:Precipitin Tests; D011485:Protein Binding; D011493:Protein Kinase C / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000378:metabolism* / Q000502:physiology*; D019908:Proto-Oncogene Proteins c-raf / Q000378:metabolism; D013997:Time Factors; D014157:Transcription Factors / Q000378:metabolism; D010738:Type C Phospholipases / Q000378:metabolism; D015854:Up-Regulation; D051746:ZAP-70 Protein-Tyrosine Kinase
chemical_list: D017252:CD3 Complex; D003432:Cross-Linking Reagents; D004268:DNA-Binding Proteins; D004791:Enzyme Inhibitors; D007527:Isoenzymes; D050778:NFATC Transcription Factors; D009687:Nuclear Proteins; D014157:Transcription Factors; D011505:Protein-Tyrosine Kinases; D051746:ZAP-70 Protein-Tyrosine Kinase; C497261:ZAP70 protein, human; D017346:Protein Serine-Threonine Kinases; D019908:Proto-Oncogene Proteins c-raf; D011493:Protein Kinase C; D020928:Mitogen-Activated Protein Kinases; D048369:MAP Kinase Kinase 1; C482125:MAP2K1 protein, human; D020929:Mitogen-Activated Protein Kinase Kinases; D010738:Type C Phospholipases; D051966:Phospholipase C gamma; D002118:Calcium