Cbl-mediated negative regulation of platelet-derived growth factor receptor-dependent cell proliferation. A critical role for Cbl tyrosine kinase-binding domain.
The Cbl proto-oncogene product has emerged as a novel negative regulator of receptor and non-receptor tyrosine kinases. Our previous observations that Cbl overexpression in NIH3T3 cells enhanced the ubiquitination and degradation of the platelet-derived growth factor receptor-alpha (PDGFRalpha) and that the expression of oncogenic Cbl mutants up-regulated the PDGFRalpha signaling machinery strongly suggested that Cbl negatively regulates PDGFRalpha signaling. Here, we show that, similar to PDGFRalpha, selective stimulation of PDGFRbeta induces Cbl phosphorylation, and its physical association with the receptor. Overexpression of wild type Cbl in NIH3T3 cells led to an enhancement of the ligand-dependent ubiquitination and subsequent degradation of the PDGFRbeta, as observed with PDGFRalpha. We show that Cbl-dependent negative regulation of PDGFRalpha and beta results in a reduction of PDGF-induced cell proliferation and protection against apoptosis. A point mutation (G306E) that inactivates the tyrosine kinase binding domain in the N-terminal transforming region of Cbl compromised the PDGF-inducible tyrosine phosphorylation of Cbl although this mutant could still associate with the PDGFR. More importantly, the G306E mutation abrogated the ability of Cbl to enhance the ligand-induced ubiquitination and degradation of the PDGFR and to inhibit the PDGF-dependent cell proliferation and protection from apoptosis. These results demonstrate that Cbl can negatively regulate PDGFR-dependent biological responses and that this function requires the conserved tyrosine kinase binding domain of Cbl.
Miyake
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Lymphocyte Biology Section, Division of Rheumatology, Immunology and Allergy, Department of Medicine, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts 02115, USA.
Mullane-Robinson
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KP
Lill
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Douillard
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P
Band
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H
D016475:3T3 Cells; D000818:Animals; D017209:Apoptosis; D001665:Binding Sites; D002455:Cell Division; D008024:Ligands; D051379:Mice; D010766:Phosphorylation; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D050721:Proto-Oncogene Proteins c-cbl; D017479:Receptors, Platelet-Derived Growth Factor / Q000502:physiology*; D015398:Signal Transduction; D014443:Tyrosine / Q000378:metabolism; D044767:Ubiquitin-Protein Ligases; D014452:Ubiquitins / Q000378:metabolism
D008024:Ligands; D010982:Platelet-Derived Growth Factor; D011518:Proto-Oncogene Proteins; D014452:Ubiquitins; C075278:platelet-derived growth factor A; D014443:Tyrosine; D050721:Proto-Oncogene Proteins c-cbl; D044767:Ubiquitin-Protein Ligases; D011505:Protein-Tyrosine Kinases; D017479:Receptors, Platelet-Derived Growth Factor; C493849:Cbl protein, mouse