A novel phosphotyrosine-binding domain in the N-terminal transforming region of Cbl interacts directly and selectively with ZAP-70 in T cells.
The protooncogene product Cbl has emerged as a novel signal transduction protein downstream of a number of cell surface receptors coupled to tyrosine kinases. Recently, we and others have reported the activation-dependent association of Cbl with the Syk and ZAP-70 tyrosine kinases through presently undefined mechanisms. Potential Src homology 2 and 3 domain binding sites within the C-terminal half of Cbl mediate in vivo interactions with several signaling proteins; however, the N-terminal transforming region (Cbl-N) lacks recognizable catalytic or protein interaction motifs. Here, we show that in vitro Cbl-N (amino acids 1-357) but not Cbl-C (amino acids 358-906) binds to ZAP-70 in a T cell-activation-dependent manner. A point mutation in Cbl-N, G306E, corresponding to a loss-of-function mutation in the Caenorhabditis elegans Cbl homologue, SLI-1, severely compromised Cbl-N/ZAP-70 binding. Cbl-N/ZAP-70 binding was direct and phosphotyrosine-dependent, thus identifying a phosphotyrosine-binding domain within the transforming region of Cbl. In vivo, Cbl-N expressed in T cells selectively associated with the ZAP-70/zeta complex. These results identify a novel mechanism for the direct participation of the N-terminal region of Cbl in ZAP-70 signal transduction, and suggest a biochemical mechanism for the leukemogenicity of the oncogene v-cbl through potential interaction with proliferation-related phosphotyrosyl proteins.
Lupher
M L
ML
Department of Medicine, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts 02115, USA.
Reedquist
K A
KA
Miyake
S
S
Langdon
W Y
WY
Band
H
H
D002460:Cell Line; D002471:Cell Transformation, Neoplastic; D006801:Humans; D010750:Phosphoproteins / Q000378:metabolism; D019000:Phosphotyrosine / Q000378:metabolism*; D011485:Protein Binding; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000737:chemistry / Q000378:metabolism*; D050721:Proto-Oncogene Proteins c-cbl; D015398:Signal Transduction; D013329:Structure-Activity Relationship; D013601:T-Lymphocytes / Q000378:metabolism*; D044767:Ubiquitin-Protein Ligases; D051746:ZAP-70 Protein-Tyrosine Kinase
D010750:Phosphoproteins; D011518:Proto-Oncogene Proteins; D019000:Phosphotyrosine; D050721:Proto-Oncogene Proteins c-cbl; D044767:Ubiquitin-Protein Ligases; D011505:Protein-Tyrosine Kinases; D051746:ZAP-70 Protein-Tyrosine Kinase; C497261:ZAP70 protein, human; C067419:CBL protein, human