The SH3 domain-binding T cell tyrosyl phosphoprotein p120. Demonstration of its identity with the c-cbl protooncogene product and in vivo complexes with Fyn, Grb2, and phosphatidylinositol 3-kinase.
Previously, we have identified p120 as a Fyn/Lck SH3 and SH2 domain-binding protein that is tyrosine phosphorylated rapidly after T cell receptor triggering. Here, we used direct protein purification, amino acid sequence analysis, reactivity with antibodies, and two-dimensional gel analyses to identify p120 as the human c-cbl protooncogene product. We demonstrate in vivo complexes of p120cbl with Fyn tyrosine kinase, the adaptor protein Grb2, and the p85 subunit of phosphatidylinositol (PI) 3-kinase. The association of p120cbl with Fyn and the p85 subunit of PI 3-kinase (together with PI 3-kinase activity) was markedly increased by T cell activation, consistent with in vitro binding of p120cbl to their SH2 as well as SH3 domains. In contrast, a large fraction of p120cbl was associated with Grb2 prior to activation, and this association did not change upon T cell activation. In vitro, p120cbl interacted with Grb2 exclusively through its SH3 domains. These results demonstrate a novel Grb2-p120cbl signaling complex in T cells, distinct from the previously analyzed Grb2-Sos complex. The association of p120cbl with ubiquitous signaling proteins strongly suggests a general signal transducing function for this enigmatic protooncogene with established leukemogenic potential but unknown physiological function.
Fukazawa
T
T
Department of Rheumatology and Immunology, Brigham and Women's Hospital, Boston, Massachusetts 02115, USA.
Reedquist
K A
KA
Trub
T
T
Soltoff
S
S
Panchamoorthy
G
G
Druker
B
B
Cantley
L
L
Shoelson
S E
SE
Band
H
H
D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D051380:GRB2 Adaptor Protein; D006801:Humans; D008213:Lymphocyte Activation; D008969:Molecular Sequence Data; D019869:Phosphatidylinositol 3-Kinases; D010766:Phosphorylation; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000378:metabolism*; D011506:Proteins / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D016315:Proto-Oncogene Proteins c-abl / Q000302:isolation & purification / Q000378:metabolism*; D051076:Proto-Oncogene Proteins c-fyn; D011948:Receptors, Antigen, T-Cell / Q000502:physiology*; D014443:Tyrosine / Q000378:metabolism*
D048868:Adaptor Proteins, Signal Transducing; D051380:GRB2 Adaptor Protein; C495948:GRB2 protein, human; D011506:Proteins; D011518:Proto-Oncogene Proteins; D011948:Receptors, Antigen, T-Cell; D014443:Tyrosine; D019869:Phosphatidylinositol 3-Kinases; D017853:Phosphotransferases (Alcohol Group Acceptor); C494957:FYN protein, human; D016315:Proto-Oncogene Proteins c-abl; D051076:Proto-Oncogene Proteins c-fyn