Transformation of myeloid leukemia cells to cytokine independence by Bcr-Abl is suppressed by kinase-defective Hck.
Bcr-Abl is the constitutively active protein-tyrosine kinase expressed as a result of the Philadelphia translocation in chronic myelogenous leukemia. Bcr-Abl is coupled to many of the same signaling pathways normally regulated by hematopoietic cytokines. Recent work shows that Hck, a member of the Src tyrosine kinase family with myeloid-restricted expression, associates with and is activated by Bcr-Abl. Here we investigated the mechanism of Hck interaction with Bcr-Abl and the requirement for Hck activation in Bcr-Abl transformation signaling. Binding studies demonstrated that the Hck SH3 and SH2 domains are sufficient for interaction with Bcr-Abl in vitro. Hck binding localizes to the Abl SH2, SH3, and kinase domains as well as the distal portion of the C-terminal tail. To address the requirement for endogenous Src family kinase activation in Bcr-Abl signaling, a kinase-defective mutant of Hck was stably expressed in the cytokine-dependent myeloid leukemia cell line DAGM. Kinase-defective Hck dramatically suppressed Bcr-Abl-induced outgrowth of these cells in the absence of cytokine compared with a control cell line expressing beta-galactosidase. In contrast, kinase-defective Hck did not affect cell proliferation in response to interleukin-3, suggesting that the effect is specific for Bcr-Abl. These data show that Hck interacts with Bcr-Abl through a complex mechanism involving kinase-dependent and -independent components and that interaction with Hck or other Src family members is essential for transformation signaling by Bcr-Abl.
Lionberger
J M
JM
Department of Molecular Genetics, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15261, USA.
Wilson
M B
MB
Smithgall
T E
TE
D000818:Animals; D002478:Cells, Cultured; D016207:Cytokines / Q000502:physiology*; D016044:Fusion Proteins, bcr-abl / Q000494:pharmacology*; D006801:Humans; D007951:Leukemia, Myeloid / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000494:pharmacology*; D011518:Proto-Oncogene Proteins / Q000494:pharmacology*; D051572:Proto-Oncogene Proteins c-hck; D015398:Signal Transduction; D018411:Spodoptera; D018909:src Homology Domains; D019061:src-Family Kinases / Q000378:metabolism
D016207:Cytokines; D011518:Proto-Oncogene Proteins; D011505:Protein-Tyrosine Kinases; D016044:Fusion Proteins, bcr-abl; C496631:HCK protein, human; D051572:Proto-Oncogene Proteins c-hck; D019061:src-Family Kinases