Abstract
Crk belongs to the adapter proteins that participate in many signalling pathways from cell surface receptors. We have characterised the CrkII-23 mutant that inhibits the transformation of NRK cells induced by epidermal growth factor (EGF) and transforming growth factor (TGF)-beta. To study the biochemical difference, cDNAs of the wild-type CrkII and the CrkII-23 mutant were introduced stably into NIH 3T3 cells expressing EGF receptor (EGFR). Both CrkII and CrkII-23 were phosphorylated on tyrosine upon EGF simulation with similar time course and dose dependency. Whereas the wild-type CrkII bound to EGFR only after EGF stimulation, CrkII-23 bound to EGFR from before stimulation. Mutation in the Src homology (SH) 2 or amino-terminal SH3 domain did not abolish the binding of CrkII-23 to EGFR in the quiescent cells, suggesting that the binding is mediated by a novel mechanism. These CrkII-23-derived mutants, however, did not suppress transformation of NRK cells by EGF and TGF-beta. Hence, both the SH2 and amino-terminal SH3 domains are required to inhibit transformation of NRK cells. These results suggest that persistent signalling from CrkII-23 bound to EGFR suppresses transformation by EGF and TGF-beta in NRK23 cells.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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3T3 Cells
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Adaptor Proteins, Signal Transducing*
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Adaptor Proteins, Vesicular Transport*
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Animals
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COS Cells
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Cell Differentiation / drug effects
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Cell Transformation, Neoplastic / genetics*
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Cell Transformation, Neoplastic / metabolism
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DNA Mutational Analysis
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Epidermal Growth Factor / pharmacology*
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ErbB Receptors / metabolism*
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Eukaryotic Initiation Factor-2
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Guanine Nucleotide Exchange Factors
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Humans
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Mice
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Mutation*
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Neurons / cytology
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Neurons / drug effects
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PC12 Cells
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Phosphorylation
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Protein Binding
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Protein Kinases / genetics*
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Protein Kinases / metabolism*
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Protein Kinases / physiology
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Protein Structure, Tertiary
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Proteins / metabolism
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Proto-Oncogene Proteins / metabolism
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Proto-Oncogene Proteins c-abl / metabolism
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Proto-Oncogene Proteins c-cbl
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Proto-Oncogene Proteins c-crk
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Rats
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Shc Signaling Adaptor Proteins
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Src Homology 2 Domain-Containing, Transforming Protein 1
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Transforming Growth Factor beta / pharmacology*
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Tyrosine / metabolism
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Ubiquitin-Protein Ligases*
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src Homology Domains / genetics*
Substances
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Adaptor Proteins, Signal Transducing
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Adaptor Proteins, Vesicular Transport
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Crk protein, rat
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Eukaryotic Initiation Factor-2
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Guanine Nucleotide Exchange Factors
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Proteins
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Proto-Oncogene Proteins
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Proto-Oncogene Proteins c-crk
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SHC1 protein, human
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Shc Signaling Adaptor Proteins
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Shc1 protein, mouse
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Shc1 protein, rat
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Src Homology 2 Domain-Containing, Transforming Protein 1
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Transforming Growth Factor beta
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Tyrosine
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Epidermal Growth Factor
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Proto-Oncogene Proteins c-cbl
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Ubiquitin-Protein Ligases
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Protein Kinases
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ErbB Receptors
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Proto-Oncogene Proteins c-abl
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CBL protein, human
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Cbl protein, mouse