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.
Cancer Biology Division, Garvan Institute of Medical Research, St Vincent's Hospital, Sydney, NSW, Australia.
P W
PW
Janes
R J
RJ
Daly
A
A
deFazio
R L
RL
Sutherland
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