Fyn and ZAP-70 Are Required for Vav Phosphorylation in T Cells Stimulated by Antigen-presenting Cells

In T cells, triggering of the T cell antigen receptor or of the co-stimulatory receptor CD28 can direct tyrosine phosphorylation of the signaling protein Vav. We investigated the role played by the protein tyrosine kinases Fyn, Lck, and ZAP-70 in these processes in a T cell hybridoma after physiolog...

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Bibliographic Details
Published in:The Journal of biological chemistry 1998-11, Vol.273 (48), p.31932-31938
Main Authors: Michel, Frédérique, Grimaud, Linda, Tuosto, Loretta, Acuto, Oreste
Format: Article
Language:English
Subjects:
Animals
Antigen-Presenting Cells - immunology
Antigen-Presenting Cells - physiology
CD28 Antigens - physiology
Cell Cycle Proteins
Hybridomas
L Cells
Lymphocyte Activation
Mice
Phosphorylation
Phosphotyrosine - metabolism
Protein-Tyrosine Kinases - metabolism
Proto-Oncogene Proteins - metabolism
Proto-Oncogene Proteins c-fyn
Proto-Oncogene Proteins c-vav
Receptors, Antigen, T-Cell - metabolism
Recombinant Proteins - metabolism
T-Lymphocytes - immunology
T-Lymphocytes - physiology
Transfection
ZAP-70 Protein-Tyrosine Kinase
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Summary:In T cells, triggering of the T cell antigen receptor or of the co-stimulatory receptor CD28 can direct tyrosine phosphorylation of the signaling protein Vav. We investigated the role played by the protein tyrosine kinases Fyn, Lck, and ZAP-70 in these processes in a T cell hybridoma after physiological stimulation of the T cell receptor (TCR) and CD28. A dominant-negative mutant approach based on overexpression of catalytically inactive alleles of these kinases showed that CD28-induced Vav phosphorylation preferentially requires Fyn, whereas ZAP-70 had no role. Consistently, Vav was strongly phosphorylated in Lck-deficient JCAM-1 cells after CD28 ligation. In contrast, ZAP-70 appeared to control TCR-directed Vav phosphorylation. However, overexpression of ZAP-70 carrying a mutated Tyr315, contained within a motif previously suggested to be a Vav Src homology 2 domain binding site, had little or no effect. Immunoprecipitation assays showed that phosphorylated Vav associated with Fyn after CD28 triggering and that this interaction, likely to involve binding of Fyn Src homology 2 domain to Vav, was more strongly detectable after concomitant CD28 and TCR stimulation. These data suggest that Fyn plays a major role in controlling Vav phosphorylation upon T cell activation and that the mechanism implicating ZAP-70 in this process may be more complex than previously anticipated.
ISSN:0021-9258
1083-351X
DOI:10.1074/jbc.273.48.31932