Molecular cloning of SLP-76, a 76-kDa tyrosine phosphoprotein associated with Grb2 in T cells.
The activation of protein tyrosine kinases is a critical event in T cell antigen receptor (TCR)-mediated signaling. One substrate of the TCR-activated protein tyrosine kinase pathway is a 76-kDa protein (pp76) that associates with the adaptor protein Grb2. In this report we describe the purification of pp76 and the molecular cloning of its cDNA, which encodes a novel 533-amino acid protein with a single carboxyl-terminal Src homology 2 (SH2) domain. Although no recognizable motifs related to tyrosine, serine/threonine, or lipid kinase domains are present in the predicted amino acid sequence, it contains several potential motifs recognized by SH2 and SH3 domains. A cDNA encoding the murine homologue of pp76 was also isolated and predicts a protein with 84% amino acid identity to human pp76. Northern analysis demonstrates that pp76 mRNA is expressed solely in peripheral blood leukocytes, thymus, and spleen; and in human T cell, B cell and monocytic cell lines. In vitro translation of pp76 cDNA gives rise to a single product of 76 kDa that associates with a GST/Grb2 fusion protein, demonstrating a direct association between these two molecules. Additionally, a GST fusion protein consisting of the predicted SH2 domain of pp76 precipitates two tyrosine phosphoproteins from Jurkat cell lysates, and antiserum directed against phospholipase C-gamma 1 coprecipitates a tyrosine phosphoprotein with an electrophoretic mobility identical to that of pp76. These results demonstrate that this novel protein, which we term SLP-76 (SH2 domain-containing Leukocyte Protein of 76 kDa), is likely to play an important role in TCR-mediated intracellular signal transduction.
Jackman
J K
JK
Institute of Biochemistry and Cell Biology, Syntex Discovery Research, Palo Alto, California 94304, USA.
Motto
D G
DG
Sun
Q
Q
Tanemoto
M
M
Turck
C W
CW
Peltz
G A
GA
Koretzky
G A
GA
Findell
P R
PR
D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D000818:Animals; D001402:B-Lymphocytes / Q000378:metabolism; D001483:Base Sequence; D002460:Cell Line; D003001:Cloning, Molecular; D066246:ErbB Receptors / Q000378:metabolism; D051380:GRB2 Adaptor Protein; D015870:Gene Expression; D006367:HeLa Cells; D006801:Humans; D007962:Leukocytes / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009838:Oligodeoxyribonucleotides; D009928:Organ Specificity; D010750:Phosphoproteins / Q000096:biosynthesis / Q000097:blood* / Q000302:isolation & purification / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D011506:Proteins / Q000302:isolation & purification / Q000378:metabolism*; D012333:RNA, Messenger / Q000032:analysis / Q000096:biosynthesis; D017386:Sequence Homology, Amino Acid; D013154:Spleen / Q000378:metabolism; D013601:T-Lymphocytes / Q000378:metabolism; D013950:Thymus Gland / Q000378:metabolism; D014407:Tumor Cells, Cultured; D014443:Tyrosine
D048868:Adaptor Proteins, Signal Transducing; D051380:GRB2 Adaptor Protein; C495948:GRB2 protein, human; C495949:Grb2 protein, mouse; D009838:Oligodeoxyribonucleotides; D010750:Phosphoproteins; D011506:Proteins; D012333:RNA, Messenger; C092947:SLP-76 signal Transducing adaptor proteins; D014443:Tyrosine; D066246:ErbB Receptors; D011505:Protein-Tyrosine Kinases