The SH3 Domain-binding T Cell Tyrosyl Phosphoprotein p120

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.