Csk is constitutively associated with a 60-kDa tyrosine-phosphorylated protein in human T cells.
The protein-tyrosine kinase Csk is one of the main down-regulators of the Src family of kinases. Csk may be involved in the down-regulation of T cell receptor (TCR) signaling by C-terminal tyrosine phosphorylation of Lck and Fyn; however, it is not known how Csk activity is regulated or how it targets these Src family members. We used Jurkat T cells and normal human T cells to examine proteins that bind to the SH2 domain of Csk. In both Jurkat and normal T cells, the Src homology 2 (SH2) domain of Csk bound constitutively to a tyrosine-phosphorylated protein of 60 kDa (p60). The 60-kDa protein was detected in Csk immunoprecipitates from both unstimulated and CD3-stimulated cells. In addition to p60, a protein of 190 kDa coprecipitated with Csk, and both proteins were phosphorylated on tyrosine residues by the immunocomplex. Small amounts of GTPase-activating protein (GAP) were detected in anti-Csk immunoprecipitates, suggesting that p60 may be a GAP-associated protein. Our data demonstrate that the SH2 domain of Csk specifically associates with at least two tyrosine-phosphorylated proteins in normal human T cells, that this association is independent of TCR/CD3 activation, and that Csk may be a part of a multiprotein complex containing GAP.
Catipović
B
B
Division of Clinical Immunology, Johns Hopkins Asthma and Allergy Center, The John Hopkins University School of Medicine, Baltimore, Maryland 21224, USA.
Schneck
J P
JP
Brummet
M E
ME
Marsh
D G
DG
Rafnar
T
T
D000595:Amino Acid Sequence; D000081247:CSK Tyrosine-Protein Kinase; D002460:Cell Line; D004591:Electrophoresis, Polyacrylamide Gel; D005982:Glutathione Transferase / Q000096:biosynthesis / Q000302:isolation & purification; D006801:Humans; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010750:Phosphoproteins / Q000302:isolation & purification / Q000378:metabolism*; D019000:Phosphotyrosine; D011485:Protein Binding; D011505:Protein-Tyrosine Kinases / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism; D013601:T-Lymphocytes / Q000201:enzymology*; D014162:Transfection; D014407:Tumor Cells, Cultured; D019061:src-Family Kinases
D010750:Phosphoproteins; D011993:Recombinant Fusion Proteins; D019000:Phosphotyrosine; D005982:Glutathione Transferase; D011505:Protein-Tyrosine Kinases; D000081247:CSK Tyrosine-Protein Kinase; D019061:src-Family Kinases; C000631346:CSK protein, human