Stimulation through the T Cell Receptor Induces Cbl Association with Crk Proteins and the Guanine Nucleotide Exchange Protein C3G

• Published in: The Journal of biological chemistry Vol. 271; no. 14; pp. 8435 - 8442
• Main Authors: Reedquist, K A, Fukazawa, T, Panchamoorthy, G, Langdon, W Y, Shoelson, S E, Druker, B J, Band, H
• Format: Journal Article
• Language: English
• Published: United States American Society for Biochemistry and Molecular Biology 05.04.1996
• Subjects: Adaptor Proteins, Signal Transducing; Amino Acid Sequence; Crk-Associated Substrate Protein; Guanine Nucleotide Exchange Factors; Humans; Leucine Zippers; Lymphocyte Activation; Molecular Sequence Data; Nuclear Proteins - metabolism; Phosphoproteins - metabolism; Phosphotyrosine - metabolism; Protein Binding; Proteins - metabolism; Proto-Oncogene Proteins - metabolism; Proto-Oncogene Proteins c-cbl; Receptors, Antigen, T-Cell - physiology; Recombinant Proteins; Retinoblastoma-Like Protein p130; Signal Transduction; src Homology Domains; T-Lymphocytes - metabolism; Tumor Cells, Cultured; Ubiquitin-Protein Ligases
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.271.14.8435

We and others have recently identified Cbl, the protein product of the c- cbl protooncogene, as an early tyrosine kinase substrate upon T cell activation and have shown that Cbl forms in vivo complexes with Src family tyrosine kinases, Grb2 adaptor protein, and the p85 subunit of PI-3 kinase. Here we show that Cbl associates with all three forms of the human Crk protein, predominantly CrkL, following T cell receptor activation of Jurkat T cells. Association between Cbl and Crk proteins was confirmed in normal human peripheral blood-derived T cells. In vitro , Cbl was able to interact with the Crk SH2 domain but not the SH3 domain. A phosphopeptide corresponding to a potential Crk SH2 domain-binding motif in Cbl (pYDVP) specifically inhibited binding between Cbl and Crk SH2 domain. Anti-Cbl antibody completely immunodepleted the CrkL-associated 120kDa phosphotyrosyl polypeptide, suggesting that the recently described p130 -related Crk-associated p116 of T cells may be Cbl. Consistent with this possibility, the 4F4 antibody used to characterize the p116 polypeptide cross-reacted with Cbl protein when it was resolved on one- or two-dimensional gels. CrkL was constitutively associated with a substantial amount of the guanine nucleotide exchange protein C3G, and a fraction of the C3G protein was coimmunoprecipitated with Cbl in activated Jurkat T cells. These results suggest the possibility that Cbl may participate in a signaling pathway that regulates guanine nucleotide exchange on small G-proteins in T cells.