Coexistence of phosphotyrosine-dependent and -independent interactions between Cbl and Bcr-Abl

I. Gaston, K.J. Johnson, T. Oda, A. Bhat, M. Reis, Wallace Langdon, L. Shen, M.W. Deininger, B.J. Druker

Research output: Contribution to journalArticlepeer-review

12 Citations (Scopus)


Objectives. Cbl is one of the major tyrosine-phosphorylated proteins in Bcr-Abi-expressing cells. A direct association between the SH2 domain of Bcr-Abl and tyrosine-phosphorylated Cbl has been demonstrated. The purpose of this study was to determine if and how unphosphorylated Cbl and Bcr-Abl may associate.Methods. Interactions between Cbl and Ber-Abl were investigated in yeast two- and three-hybrid systems, gel overlay assays, and immuno precipitates from mammalian cells expressing wild-type and the Y177F mutant of Ber-Abl.Results. No direct interaction between Bcr-Abl and unphosphorylated Cbl was observed. Bcr-Abl did, however, associate with Grb2, an adaptor protein that binds tyrosine 177 of Bcr-Abl. Additionally, Grb2 interacted with Cbl. In a yeast three-hybrid assay, Grb2 mediated an interaction between Cb1 and Bcr-Abl that was dependent on a functional Grb2 binding site. This interaction was confirmed in vitro using purified proteins. In cells expressing Bcr-Abl with a mutation in the Grb2 binding site, binding of Cbl to Bcr-Abl was significantly reduced, but Cbl tyrosine phosphorylation was maintained. Imatinib treatment of these cells further reduced but did not abrogate Cbl binding, reflecting residual kinase activity.Conclusion. Multiple phosphotyrosine-dependent and -independent interactions stabilize the interaction between Cbl and Abl. Grb2 or another, yet unidentified, protein may mediate an initial interaction between Cbl and Bcr-Abl that is independent of Cbl tyrosine phosphorylation. Following this initial interaction, Cbl can then become tyrosine phosphorylated and interact with the SH2 domain of Ber-Abl, further stabilizing the complex. (C) 2004 International Society for Experimental Hematology. Published by Elsevier Inc.
Original languageEnglish
Pages (from-to)113-121
JournalExperimental Hematology
Issue numberNA
Publication statusPublished - 2004


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