Characterization of p59fyn-mediated signal transduction on T cell activation.
Protein tyrosine kinase p59fyn is associated with the TCR-CD3 complex and is suggested to play a role in T cell activation. To determine the molecular mechanism of p59fyn-mediated signal transduction in T cell activation, we established murine T cell hybridoma lines that expressed an elevated amount of wild-type or mutant fyns. Clones that expressed high levels of normal p59fyn and active p59fyn, encoded by wild-type and f-14 mutant fyn respectively, showed enhanced IL-2 production upon stimulation by anti-CD3 antibodies or natural antigen. On the other hand, clones that expressed kinase negative p59fyn and p59fyn with an SH2 (Src-homology 2) deletion encoded by t-1 mutant fyn showed little induction of IL-2 production upon stimulation. These data suggest that p59fyn is important in T cell signaling and that the SH2 sequence plays a critical role in the reaction. Induction of tyrosine phosphorylation of multiple proteins upon antigenic stimulation was augmented similarly in the cells that respectively expressed wild-type and f-14 mutant fyns at elevated levels. The proteins that became highly tyrosine-phosphorylated included phospholipase C (PLC-gamma 1), p95vav, ZAP-70, the MAP kinase, CD3 zeta and unidentified proteins of 120, 100 and 80 kDa. Tyrosine phosphorylation of the 120, 95 and 68 kDa proteins associated with PLC-gamma 1 was also observed in these cells upon stimulation. In contrast, only the 100 kDa protein and the MAP kinase were increasingly tyrosine phosphorylated in the antigen-stimulated cells expressing t-1 fyn. These data suggest that PLC-gamma 1, PLC-gamma 1 associated molecules, p95vav, the 80 kDa protein, ZAP-70 and the CD3 zeta chain may be substrates of p59fyn or of other tyrosine kinases regulated by p59fyn and be important in T cell signaling.
Fusaki
N
N
Department of Oncology, University of Tokyo, Japan.
Semba
K
K
Katagiri
T
T
Suzuki
G
G
Matsuda
S
S
Yamamoto
T
T
D000818:Animals; D000936:Antigen-Antibody Complex / Q000276:immunology / Q000378:metabolism; D005434:Flow Cytometry; D006825:Hybridomas / Q000276:immunology; D015151:Immunoblotting; D007376:Interleukin-2 / Q000096:biosynthesis*; D007527:Isoenzymes / Q000378:metabolism; D008213:Lymphocyte Activation / Q000276:immunology*; D051379:Mice; D051966:Phospholipase C gamma; D011233:Precipitin Tests; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D051076:Proto-Oncogene Proteins c-fyn; D017260:Receptor-CD3 Complex, Antigen, T-Cell / Q000276:immunology; D015398:Signal Transduction / Q000276:immunology*; D013601:T-Lymphocytes / Q000276:immunology*; D014162:Transfection; D010738:Type C Phospholipases / Q000378:metabolism
D000936:Antigen-Antibody Complex; D007376:Interleukin-2; D007527:Isoenzymes; D011518:Proto-Oncogene Proteins; D017260:Receptor-CD3 Complex, Antigen, T-Cell; D011505:Protein-Tyrosine Kinases; C494960:Fyn protein, mouse; D051076:Proto-Oncogene Proteins c-fyn; D010738:Type C Phospholipases; D051966:Phospholipase C gamma