Abstract
Receptor tyrosine kinases (RTKs) such as the fibroblast growth factor receptor (FGFR) and the epidermal growth factor receptor are overexpressed in a variety of cancers. In addition to overexpression, the FGFRs are found mutated in some cancers. The Src homology 2 domain-containing phosphotyrosine phosphatase (SHP2) is a critical mediator of RTK signaling, but its role in oncogenic RTK-induced cell transformation and cancer development is largely unknown. In the current report, we demonstrate that constitutively activated FGFR3 (K/E-FR3) transforms NIH-3T3 cells, and that SHP2 is a critical mediator of this transformation. Infection of K/E-FR3-transformed 3T3 cells with a retrovirus carrying a dominant-negative mutant of SHP2 (C/S-SHP2) retarded cell growth, reversed the transformation phenotype and inhibited focus-forming ability. Furthermore, treatment of K/E-FR3-transformed NIH-3T3 cells with PD98059 or LY294002, specific inhibitors of MEK and PI3K, respectively, inhibited focus formation. Biochemical analysis showed that K/E-FR3 activates the Ras-ERK and the PI3K signaling pathways, and that the C/S SHP2 mutant suppressed this effect via competitive displacement of interaction of the endogenous SHP2 with FRS2. However, the C/S SHP2 protein did not show any effect on receptor autophosphorylation, FRS2 tyrosine phosphorylation or interaction of Grb2 with K/E-FR3 or FRS2. Together, the results show that K/E-FR3 is transforming and that the Ras-ERK and the PI3K-Akt signaling pathways, which are positively regulated by SHP2, are important for K/E-FR3-induced transformation.
Publication types
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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3T3 Cells
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Adaptor Proteins, Signal Transducing*
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Animals
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COS Cells
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Calcium-Calmodulin-Dependent Protein Kinases / antagonists & inhibitors
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Carrier Proteins / genetics
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Carrier Proteins / metabolism*
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Cell Line, Transformed
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Cell Transformation, Neoplastic
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Cell Transformation, Viral*
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Cells, Cultured
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Chlorocebus aethiops
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Chromones / pharmacology
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Enzyme Activation
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Enzyme Inhibitors / pharmacology
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Fibroblast Growth Factors / metabolism
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Flavonoids / pharmacology
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Gene Expression Regulation
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Membrane Glycoproteins
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Membrane Proteins / genetics
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Membrane Proteins / metabolism*
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Mice
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Mitogen-Activated Protein Kinase 1 / metabolism
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Mitogen-Activated Protein Kinase 3
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Mitogen-Activated Protein Kinases / metabolism
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Morpholines / pharmacology
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Mutagenesis, Site-Directed
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Phosphatidylinositol 3-Kinases / genetics
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Phosphoinositide-3 Kinase Inhibitors
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Protein Serine-Threonine Kinases*
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Protein-Tyrosine Kinases*
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Proto-Oncogene Proteins / metabolism
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Proto-Oncogene Proteins c-akt
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Receptor, Fibroblast Growth Factor, Type 3
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Receptors, Fibroblast Growth Factor / genetics
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Receptors, Fibroblast Growth Factor / metabolism*
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Retroviridae / genetics
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Signal Transduction / drug effects
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ras Proteins / metabolism
Substances
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Adaptor Proteins, Signal Transducing
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Carrier Proteins
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Chromones
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Enzyme Inhibitors
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Flavonoids
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Membrane Glycoproteins
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Membrane Proteins
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Morpholines
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Phosphoinositide-3 Kinase Inhibitors
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Proto-Oncogene Proteins
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Receptors, Fibroblast Growth Factor
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SIT1 protein, human
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2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one
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Fibroblast Growth Factors
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Fgfr3 protein, mouse
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Protein-Tyrosine Kinases
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Receptor, Fibroblast Growth Factor, Type 3
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Protein Serine-Threonine Kinases
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Proto-Oncogene Proteins c-akt
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Calcium-Calmodulin-Dependent Protein Kinases
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Mitogen-Activated Protein Kinase 1
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Mitogen-Activated Protein Kinase 3
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Mitogen-Activated Protein Kinases
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ras Proteins
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2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one