Article: article from journal or magazin.
Thy-1/CD3 coengagement promotes TCR signaling and enhances particularly tyrosine phosphorylation of the raft molecule LAT.
Publication types: Journal Article ; Research Support, Non-U.S. Gov't Publication Status: ppublish
Clustering of the glycosyl-phosphatidylinositol (GPI)-anchored protein Thy-1 on the cell surface leads to T cell activation. However, despite the similarity to TCR-mediated events, cell signaling triggered by Thy-1 crosslinking, reportedly occurs in a manner independent of the TCR/CD3 complex. To investigate the relationship between responses resulting from Thy-1 or TCR engagement, a biochemically well defined system employing only affinity purified antibodies was used to crosslink these surface molecules and activation was assessed by monitoring tyrosine phosphorylation, intracellular calcium influx and IL-2 production. By these criteria, anti-CD3 mAbs moderately activated EL-4 thymoma or 2B4 hybridoma cell lines, while costimulation with anti-Thy-1-mAb strongly enhanced TCR signaling. Furthermore, a Thy-1 loss mutant cell line, did not respond to stimulation through CD3 despite expressing all essential signaling molecules. Together these results emphasized the existence of a poorly appreciated mutual interdependence between Thy-1 and CD3 for efficient cellular signaling. Thy-1/CD3-mediated activation enhanced mostly tyrosine phosphorylation of a 40 kDa protein which was identified as a transmembrane protein lacking N-linked oligosaccharides. These biochemical properties are identical to those described for a recently cloned adaptor protein called 'Linker for Activation of T cells' (LAT). Indeed, polyclonal Abs raised against a LAT-peptide (amino acids 103-131) specifically recognized the 40 kDa protein. LAT is present in microdomains of the plasma membrane enriched in sphingolipids, cholesterol, GPI-anchored proteins and a variety of signaling molecules. By contrast, the TCR/CD3 complex is excluded from these domains at least until stimulation takes place. Hence, we propose that Thy-1 promotes TCR/CD3 dependent signaling by facilitating LAT phosphorylation on tyrosine and the subsequent recruitment of downstream effector molecules.
Adaptor Proteins, Signal Transducing, Animals, Antigens, CD3/metabolism, Antigens, Thy-1/genetics, Antigens, Thy-1/metabolism, Calcium Signaling, Carrier Proteins/metabolism, Cell Line, Cross-Linking Reagents, Hybridomas/immunology, Hybridomas/metabolism, Interleukin-2/secretion, Lymphocyte Activation, Membrane Proteins/metabolism, Mice, Mutation, Phosphoproteins/metabolism, Phosphorylation, Receptors, Antigen, T-Cell/metabolism, Signal Transduction, T-Lymphocytes/immunology, T-Lymphocytes/metabolism, Tyrosine/metabolism
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