title: Global effects of BCR/ABL and TEL/PDGFRbeta expression on the proteome and phosphoproteome: identification of the Rho pathway as a target of BCR/ABL.
abstract: Many leukemic oncogenes form as a consequence of gene fusions or mutation that result in the activation or overexpression of a tyrosine kinase. To identify commonalities and differences in the action of two such kinases, breakpoint cluster region (BCR)/ABL and TEL/PDGFRbeta, two-dimensional gel electrophoresis was employed to characterize their effects on the proteome. While both oncogenes affected expression of specific proteins, few common effects were observed. A number of proteins whose expression is altered by BCR/ABL, including gelsolin and stathmin, are related to cytoskeletal function whereas no such changes were seen in TEL/PDGFRbeta-transfected cells. Treatment of cells with the kinase inhibitor STI571 for 4-h reversed changes in expression of some of these cytoskeletal proteins. Correspondingly, BCR/ABL-transfected cells were less responsive to chemotactic and chemokinetic stimuli than non-transfected cells and TEL/PDGFRbeta-transfected Ba/F3 cells. Decreased motile response was reversed by a 16-h treatment with STI571. A phosphoprotein-specific gel stain was used to identify TEL/PDGFRbeta and BCR/ABL-mediated changes in the phosphoproteome. These included changes on Crkl, Ras-GAP-binding protein 1, and for BCR/ABL, cytoskeletal proteins such as tubulin, and Nedd5. Decreased phosphorylation of Rho-GTPase dissociation inhibitor (Rho GDI) was also observed in BCR/ABL-transfected cells. This results in the activation of the Rho pathway, and treatment of cells with Y27632, an inhibitor of Rho kinase, inhibited DNA synthesis in BCR/ABL-transfected Ba/F3 cells but not TEL/PDGFRbeta-expressing cells. Expression of a dominant-negative RhoA inhibited both DNA synthesis and transwell migration, demonstrating the significance of this pathway in BCR/ABL-mediated transformation.
authors.affiliation: Faculty of Medical and Human Sciences, University of Manchester.
authors.forename: Richard D
authors.initials: RD
authors.lastname: Unwin
authors.forename: David W
authors.initials: DW
authors.lastname: Sternberg
authors.forename: Yuning
authors.initials: Y
authors.lastname: Lu
authors.forename: Andrew
authors.initials: A
authors.lastname: Pierce
authors.forename: D Gary
authors.initials: DG
authors.lastname: Gilliland
authors.forename: Anthony D
authors.initials: AD
authors.lastname: Whetton
mesh_terms: D000818:Animals; D001549:Benzamides; D002460:Cell Line; D002471:Cell Transformation, Neoplastic / Q000235:genetics; D015180:Electrophoresis, Gel, Two-Dimensional; D016044:Fusion Proteins, bcr-abl / Q000235:genetics*; D020869:Gene Expression Profiling; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects / Q000235:genetics*; D000068877:Imatinib Mesylate; D051379:Mice; D015514:Oncogene Proteins, Fusion / Q000235:genetics*; D010750:Phosphoproteins / Q000235:genetics*; D010879:Piperazines / Q000494:pharmacology; D020543:Proteome; D011743:Pyrimidines / Q000494:pharmacology; D014162:Transfection
chemical_list: D001549:Benzamides; D015514:Oncogene Proteins, Fusion; D010750:Phosphoproteins; D010879:Piperazines; D020543:Proteome; D011743:Pyrimidines; C120888:TEL-PDGFRbeta fusion protein, human; D000068877:Imatinib Mesylate; D016044:Fusion Proteins, bcr-abl