The Src family kinase Hck interacts with Bcr-Abl by a kinase-independent mechanism and phosphorylates the Grb2-binding site of Bcr.
bcr-abl, the oncogene causing chronic myeloid leukemia, encodes a fusion protein with constitutively active tyrosine kinase and transforming capacity in hematopoietic cells. Various intracellular signaling intermediates become activated and/or associate by/with Bcr-Abl, including the Src family kinase Hck. To elucidate some of the structural requirements and functional consequences of the association of Bcr-Abl with Hck, their interaction was investigated in transiently transfected COS7 cells. Neither the complex formation of Hck kinase with Bcr-Abl nor the activation of Hck by Bcr-Abl was dependent on the Abl kinase activity. Both inactivating point mutations of Hck and dephosphorylation of Hck enhanced its complex formation with Bcr-Abl, indicating that their physical interaction was negatively regulated by Hck (auto)phosphorylation. Finally, experiments with a series of kinase negative Bcr-Abl mutants showed that Hck phosphorylated Bcr-Abl and induced the binding of Grb2 to Tyr177 of Bcr-Abl. Taken together, our results suggest that Bcr-Abl preferentially binds inactive forms of Hck by an Abl kinase-independent mechanism. This physical interaction stimulates the Hck tyrosine kinase, which may then phosphorylate the Grb2-binding site in Bcr-Abl.
Warmuth
M
M
Medizinische Klinik, Klinikum Innenstadt, Universität München, D-80336 München, Germany.
Bergmann
M
M
Priess
A
A
Häuslmann
K
K
Emmerich
B
B
Hallek
M
M
D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D001665:Binding Sites; D019556:COS Cells; D066246:ErbB Receptors / Q000378:metabolism*; D016044:Fusion Proteins, bcr-abl / Q000378:metabolism*; D051380:GRB2 Adaptor Protein; D006801:Humans; D010766:Phosphorylation; D017354:Point Mutation; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D011506:Proteins / Q000235:genetics / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D051572:Proto-Oncogene Proteins c-hck; D016392:Proto-Oncogene Proteins pp60(c-src) / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; D051380:GRB2 Adaptor Protein; C495948:GRB2 protein, human; D011506:Proteins; D011518:Proto-Oncogene Proteins; D066246:ErbB Receptors; D011505:Protein-Tyrosine Kinases; D016044:Fusion Proteins, bcr-abl; C496631:HCK protein, human; D051572:Proto-Oncogene Proteins c-hck; D016392:Proto-Oncogene Proteins pp60(c-src)