Activation of the Ras signalling pathway in human breast cancer cells overexpressing erbB-2.
The c-erbB-2 proto-oncogene encodes a receptor tyrosine kinase (RTK) closely related to the epidermal growth factor receptor (EGFR). Overexpression of erbB-2 occurs in approximately 20% of human breast tumours, where increased expression correlates with poor patient prognosis. The EGFR is coupled to the Ras signalling pathway by interaction with the adaptor protein Grb2, and Sos, a Ras GDP-GTP exchange factor. In this study, activation of the erbB-2 receptor and its association with Grb2 and Sos was investigated in breast cancer cell lines which overexpress erbB-2. The receptor was found to be tyrosine phosphorylated in all cell lines in which it is overexpressed. Western blotting of Grb2 and Sos immuneprecipitates from such cells revealed co-precipitation of erbB-2, demonstrating association of the Grb2/Sos complex with erbB-2 in vivo. Furthermore, a fusion protein containing only the SH2 domain of Grb2 bound to erbB-2 immobilized on nitrocellulose, indicating that association with Grb2 is direct and mediated by the SH2 domain of Grb2. The degree of association between the erbB-2 receptor and Grb2 in vivo was related to erbB-2 overexpression, and MAP kinase, which functions downstream from Ras, displayed markedly increased activity in cell lines overexpressing erbB-2. These results demonstrate that erbB-2 is coupled to Ras signalling via the Grb2/Sos complex, and that overexpression of this receptor in breast cancer cells leads to amplification of the Ras signalling pathway.
Janes
P W
PW
Cancer Biology Division, Garvan Institute of Medical Research, St Vincent's Hospital, Sydney, NSW, Australia.
Daly
R J
RJ
deFazio
A
A
Sutherland
R L
RL
D048868:Adaptor Proteins, Signal Transducing; D001943:Breast Neoplasms / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D051380:GRB2 Adaptor Protein; D015972:Gene Expression Regulation, Neoplastic; D018734:Genes, erbB-2; D020662:Guanine Nucleotide Exchange Factors; D006801:Humans; D019950:Mitogen-Activated Protein Kinase 1; D010641:Phenotype; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D011506:Proteins / Q000378:metabolism; D000090063:Proto-Oncogene Mas; D015398:Signal Transduction; D014407:Tumor Cells, Cultured; D020663:ras Guanine Nucleotide Exchange Factors
D048868:Adaptor Proteins, Signal Transducing; D051380:GRB2 Adaptor Protein; C495948:GRB2 protein, human; D020662:Guanine Nucleotide Exchange Factors; C000715470:MAS1 protein, human; D011506:Proteins; D000090063:Proto-Oncogene Mas; D020663:ras Guanine Nucleotide Exchange Factors; D011505:Protein-Tyrosine Kinases; D017346:Protein Serine-Threonine Kinases; D019950:Mitogen-Activated Protein Kinase 1