Skip to main content
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1990 Apr;87(7):2569–2573. doi: 10.1073/pnas.87.7.2569

Agonistic antibodies stimulate the kinase encoded by the neu protooncogene in living cells but the oncogenic mutant is constitutively active.

Y Yarden 1
PMCID: PMC53731  PMID: 1969636

Abstract

The neu protooncogene (also called c-erbB2 and HER-2) undergoes oncogenic activation through a single mutation. The product of the protooncogene, p185neu, probably functions as a receptor for a peptide growth factor. To circumvent the absence of a well-characterized ligand, I generated ligand-mimicking monoclonal antibodies directed to the presumed receptor. These antibodies stimulated tyrosine phosphorylation of p185neu in living cells and also accelerated the rate of endocytosis and degradation of p185neu. A monovalent Fab fragment of such an antibody was ineffective, suggesting a role for receptor dimerization in signal transduction. Unlike the product of the protooncogene, the transforming mutant was not affected by the ligand-like antibodies. However, it undergoes constitutively high phosphorylation on tyrosine residues in living cells, and its turnover rate is remarkably rapid. Nevertheless, the pattern of phosphorylation of the mutant protein is similar to the one exhibited by an antibody-stimulated p185neu, suggesting that the mutation mimics activation by the antibody. These results suggest that the kinase of p185neu is under allosteric control that may involve ligand-induced dimerization of receptors. This mechanism is deregulated in the oncogenic mutant, which is functionally equivalent to ligand-stimulated receptor.

Full text

PDF
2569

Images in this article

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Bargmann C. I., Hung M. C., Weinberg R. A. Multiple independent activations of the neu oncogene by a point mutation altering the transmembrane domain of p185. Cell. 1986 Jun 6;45(5):649–657. doi: 10.1016/0092-8674(86)90779-8. [DOI] [PubMed] [Google Scholar]
  2. Bargmann C. I., Weinberg R. A. Increased tyrosine kinase activity associated with the protein encoded by the activated neu oncogene. Proc Natl Acad Sci U S A. 1988 Aug;85(15):5394–5398. doi: 10.1073/pnas.85.15.5394. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bargmann C. I., Weinberg R. A. Oncogenic activation of the neu-encoded receptor protein by point mutation and deletion. EMBO J. 1988 Jul;7(7):2043–2052. doi: 10.1002/j.1460-2075.1988.tb03044.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Coussens L., Yang-Feng T. L., Liao Y. C., Chen E., Gray A., McGrath J., Seeburg P. H., Libermann T. A., Schlessinger J., Francke U. Tyrosine kinase receptor with extensive homology to EGF receptor shares chromosomal location with neu oncogene. Science. 1985 Dec 6;230(4730):1132–1139. doi: 10.1126/science.2999974. [DOI] [PubMed] [Google Scholar]
  5. Defize L. H., Moolenaar W. H., van der Saag P. T., de Laat S. W. Dissociation of cellular responses to epidermal growth factor using anti-receptor monoclonal antibodies. EMBO J. 1986 Jun;5(6):1187–1192. doi: 10.1002/j.1460-2075.1986.tb04345.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Galfre G., Howe S. C., Milstein C., Butcher G. W., Howard J. C. Antibodies to major histocompatibility antigens produced by hybrid cell lines. Nature. 1977 Apr 7;266(5602):550–552. doi: 10.1038/266550a0. [DOI] [PubMed] [Google Scholar]
  7. Gilmore T., DeClue J. E., Martin G. S. Protein phosphorylation at tyrosine is induced by the v-erbB gene product in vivo and in vitro. Cell. 1985 Mar;40(3):609–618. doi: 10.1016/0092-8674(85)90209-0. [DOI] [PubMed] [Google Scholar]
  8. Glenney J. R., Jr, Chen W. S., Lazar C. S., Walton G. M., Zokas L. M., Rosenfeld M. G., Gill G. N. Ligand-induced endocytosis of the EGF receptor is blocked by mutational inactivation and by microinjection of anti-phosphotyrosine antibodies. Cell. 1988 Mar 11;52(5):675–684. doi: 10.1016/0092-8674(88)90405-9. [DOI] [PubMed] [Google Scholar]
  9. Honegger A. M., Dull T. J., Felder S., Van Obberghen E., Bellot F., Szapary D., Schmidt A., Ullrich A., Schlessinger J. Point mutation at the ATP binding site of EGF receptor abolishes protein-tyrosine kinase activity and alters cellular routing. Cell. 1987 Oct 23;51(2):199–209. doi: 10.1016/0092-8674(87)90147-4. [DOI] [PubMed] [Google Scholar]
  10. Huhn R. D., Posner M. R., Rayter S. I., Foulkes J. G., Frackelton A. R., Jr Cell lines and peripheral blood leukocytes derived from individuals with chronic myelogenous leukemia display virtually identical proteins phosphorylated on tyrosine residues. Proc Natl Acad Sci U S A. 1987 Jul;84(13):4408–4412. doi: 10.1073/pnas.84.13.4408. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Hung M. C., Schechter A. L., Chevray P. Y., Stern D. F., Weinberg R. A. Molecular cloning of the neu gene: absence of gross structural alteration in oncogenic alleles. Proc Natl Acad Sci U S A. 1986 Jan;83(2):261–264. doi: 10.1073/pnas.83.2.261. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. King C. R., Borrello I., Bellot F., Comoglio P., Schlessinger J. Egf binding to its receptor triggers a rapid tyrosine phosphorylation of the erbB-2 protein in the mammary tumor cell line SK-BR-3. EMBO J. 1988 Jun;7(6):1647–1651. doi: 10.1002/j.1460-2075.1988.tb02991.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Kurkela R., Vuolas L., Vihko P. Preparation of F(ab')2 fragments from monoclonal mouse IgG1 suitable for use in radioimaging. J Immunol Methods. 1988 Jun 13;110(2):229–236. doi: 10.1016/0022-1759(88)90108-1. [DOI] [PubMed] [Google Scholar]
  14. Padhy L. C., Shih C., Cowing D., Finkelstein R., Weinberg R. A. Identification of a phosphoprotein specifically induced by the transforming DNA of rat neuroblastomas. Cell. 1982 Apr;28(4):865–871. doi: 10.1016/0092-8674(82)90065-4. [DOI] [PubMed] [Google Scholar]
  15. Park M., Dean M., Kaul K., Braun M. J., Gonda M. A., Vande Woude G. Sequence of MET protooncogene cDNA has features characteristic of the tyrosine kinase family of growth-factor receptors. Proc Natl Acad Sci U S A. 1987 Sep;84(18):6379–6383. doi: 10.1073/pnas.84.18.6379. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Roussel M. F., Downing J. R., Rettenmier C. W., Sherr C. J. A point mutation in the extracellular domain of the human CSF-1 receptor (c-fms proto-oncogene product) activates its transforming potential. Cell. 1988 Dec 23;55(6):979–988. doi: 10.1016/0092-8674(88)90243-7. [DOI] [PubMed] [Google Scholar]
  17. Slamon D. J., Clark G. M., Wong S. G., Levin W. J., Ullrich A., McGuire W. L. Human breast cancer: correlation of relapse and survival with amplification of the HER-2/neu oncogene. Science. 1987 Jan 9;235(4785):177–182. doi: 10.1126/science.3798106. [DOI] [PubMed] [Google Scholar]
  18. Sprenger F., Stevens L. M., Nüsslein-Volhard C. The Drosophila gene torso encodes a putative receptor tyrosine kinase. Nature. 1989 Apr 6;338(6215):478–483. doi: 10.1038/338478a0. [DOI] [PubMed] [Google Scholar]
  19. Stern D. F., Kamps M. P., Cao H. Oncogenic activation of p185neu stimulates tyrosine phosphorylation in vivo. Mol Cell Biol. 1988 Sep;8(9):3969–3973. doi: 10.1128/mcb.8.9.3969. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Stern D. F., Kamps M. P. EGF-stimulated tyrosine phosphorylation of p185neu: a potential model for receptor interactions. EMBO J. 1988 Apr;7(4):995–1001. doi: 10.1002/j.1460-2075.1988.tb02906.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Yarden Y., Kuang W. J., Yang-Feng T., Coussens L., Munemitsu S., Dull T. J., Chen E., Schlessinger J., Francke U., Ullrich A. Human proto-oncogene c-kit: a new cell surface receptor tyrosine kinase for an unidentified ligand. EMBO J. 1987 Nov;6(11):3341–3351. doi: 10.1002/j.1460-2075.1987.tb02655.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Yarden Y., Ullrich A. Growth factor receptor tyrosine kinases. Annu Rev Biochem. 1988;57:443–478. doi: 10.1146/annurev.bi.57.070188.002303. [DOI] [PubMed] [Google Scholar]
  23. Yarden Y., Weinberg R. A. Experimental approaches to hypothetical hormones: detection of a candidate ligand of the neu protooncogene. Proc Natl Acad Sci U S A. 1989 May;86(9):3179–3183. doi: 10.1073/pnas.86.9.3179. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Zick Y., Rees-Jones R. W., Taylor S. I., Gorden P., Roth J. The role of antireceptor antibodies in stimulating phosphorylation of the insulin receptor. J Biol Chem. 1984 Apr 10;259(7):4396–4400. [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES