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. 1982 Aug;79(16):5026–5030. doi: 10.1073/pnas.79.16.5026

Identity of human epidermal growth factor (EGF) receptor with glycoprotein SA-7: evidence for differential phosphorylation of the two components of the EGF receptor from A431 cells.

C R Carlin, B B Knowles
PMCID: PMC346819  PMID: 6289330

Abstract

A 165-kilodalton (kDal) surface glycoprotein encoded by human chromosome 7 (SA-7) had been characterized by using antisera raised against human chromosome 7-containing somatic cell hybrids. We now present evidence that SA-7 is the human receptor for epidermal growth factor (EGF) and that these antisera recognize human-specific determinants. The gene coding for the human EGF receptor is localized to the p12 to p22 region of chromosome 7. We have characterized the 145-kDal/165-kDal EGF receptor doublet of A431 cells after immunoprecipitation of radiolabeled cell extracts with these antisera. We find that a protein with endogenous kinase activity copurifies with the A431 receptor doublet and that both components of the doublet contain phosphotyrosine and phosphothreonine and the 165-kDal component contains phosphoserine as well. Further, although each component of the receptor doublet has an average pI of 7, both display charge heterogeneity and appear to have unique charge isomers. The relationship between the two components of the A431 EGF receptor is discussed.

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Selected References

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

  1. Aden D. P., Mausner R., Knowles B. B. Production of HLA antibody in mice immunised with syngeneic mouse-human hybrid cells containing human chromosome 6. Nature. 1978 Jan 26;271(5643):375–377. doi: 10.1038/271375a0. [DOI] [PubMed] [Google Scholar]
  2. Aden D. P., Mausner R., Knowles B. B. Production of HLA antibody in mice immunised with syngeneic mouse-human hybrid cells containing human chromosome 6. Nature. 1978 Jan 26;271(5643):375–377. doi: 10.1038/271375a0. [DOI] [PubMed] [Google Scholar]
  3. COHEN S. Isolation of a mouse submaxillary gland protein accelerating incisor eruption and eyelid opening in the new-born animal. J Biol Chem. 1962 May;237:1555–1562. [PubMed] [Google Scholar]
  4. Carpenter G., Cohen S. Human epidermal growth factor and the proliferation of human fibroblasts. J Cell Physiol. 1976 Jun;88(2):227–237. doi: 10.1002/jcp.1040880212. [DOI] [PubMed] [Google Scholar]
  5. Carpenter G., King L., Jr, Cohen S. Epidermal growth factor stimulates phosphorylation in membrane preparations in vitro. Nature. 1978 Nov 23;276(5686):409–410. doi: 10.1038/276409a0. [DOI] [PubMed] [Google Scholar]
  6. Carpenter G., Lembach K. J., Morrison M. M., Cohen S. Characterization of the binding of 125-I-labeled epidermal growth factor to human fibroblasts. J Biol Chem. 1975 Jun 10;250(11):4297–4304. [PubMed] [Google Scholar]
  7. Carpenter G., Poliner L., King L., Jr Protein phosphorylation in human placenta. Stimulation by epidermal growth factor. Mol Cell Endocrinol. 1980 Jun;18(3):189–199. doi: 10.1016/0303-7207(80)90065-9. [DOI] [PubMed] [Google Scholar]
  8. Cohen S., Carpenter G., King L., Jr Epidermal growth factor-receptor-protein kinase interactions. Co-purification of receptor and epidermal growth factor-enhanced phosphorylation activity. J Biol Chem. 1980 May 25;255(10):4834–4842. [PubMed] [Google Scholar]
  9. Cohen S., Carpenter G., Lembach K. J. Interaction of epidermal growth factor (EGF) with cultured fibroblasts. Adv Metab Disord. 1975;8:265–284. doi: 10.1016/b978-0-12-027308-9.50024-x. [DOI] [PubMed] [Google Scholar]
  10. Cooper J. A., Hunter T. Changes in protein phosphorylation in Rous sarcoma virus-transformed chicken embryo cells. Mol Cell Biol. 1981 Feb;1(2):165–178. doi: 10.1128/mcb.1.2.165. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Croce C. M., Koprowski H. Somatic cell hybrids between mouse peritoneal macrophages and SV40-transformed human cells. I. Positive control of the transformed phenotype by the human chromosome 7 carrying the SV40 genome. J Exp Med. 1974 Nov 1;140(5):1221–1229. doi: 10.1084/jem.140.5.1221. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Cullen S. E., Schwartz B. D. An improved method for isolation of H-2 and Ia alloantigens with immunoprecipitation induced by protein A-bearing staphylococci. J Immunol. 1976 Jul;117(1):136–142. [PubMed] [Google Scholar]
  13. Das M., Miyakawa T., Fox C. F., Pruss R. M., Aharonov A., Herschman H. R. Specific radiolabeling of a cell surface receptor for epidermal growth factor. Proc Natl Acad Sci U S A. 1977 Jul;74(7):2790–2794. doi: 10.1073/pnas.74.7.2790. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Davies R. L., Grosse V. A., Kucherlapati R., Bothwell M. Genetic analysis of epidermal growth factor action: assignment of human epidermal growth factor receptor gene to chromosome 7. Proc Natl Acad Sci U S A. 1980 Jul;77(7):4188–4192. doi: 10.1073/pnas.77.7.4188. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Eckhart W., Hutchinson M. A., Hunter T. An activity phosphorylating tyrosine in polyoma T antigen immunoprecipitates. Cell. 1979 Dec;18(4):925–933. doi: 10.1016/0092-8674(79)90205-8. [DOI] [PubMed] [Google Scholar]
  16. Erikson E., Shealy D. J., Erikson R. L. Evidence that viral transforming gene products and epidermal growth factor stimulate phosphorylation of the same cellular protein with similar specificity. J Biol Chem. 1981 Nov 25;256(22):11381–11384. [PubMed] [Google Scholar]
  17. Fabricant R. N., De Larco J. E., Todaro G. J. Nerve growth factor receptors on human melanoma cells in culture. Proc Natl Acad Sci U S A. 1977 Feb;74(2):565–569. doi: 10.1073/pnas.74.2.565. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Gregory H. Isolation and structure of urogastrone and its relationship to epidermal growth factor. Nature. 1975 Sep 25;257(5524):325–327. doi: 10.1038/257325a0. [DOI] [PubMed] [Google Scholar]
  19. HUNTER W. M., GREENWOOD F. C. Preparation of iodine-131 labelled human growth hormone of high specific activity. Nature. 1962 May 5;194:495–496. doi: 10.1038/194495a0. [DOI] [PubMed] [Google Scholar]
  20. Haigler H. T., Carpenter G. Production and characterization of antibody blocking epidermal growth factor:receptor interactions. Biochim Biophys Acta. 1980 May 23;598(2):314–325. doi: 10.1016/0005-2736(80)90009-7. [DOI] [PubMed] [Google Scholar]
  21. Haigler H., Ash J. F., Singer S. J., Cohen S. Visualization by fluorescence of the binding and internalization of epidermal growth factor in human carcinoma cells A-431. Proc Natl Acad Sci U S A. 1978 Jul;75(7):3317–3321. doi: 10.1073/pnas.75.7.3317. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Hock R. A., Nexø E., Hollenberg M. D. Isolation of the human placenta receptor for epidermal growth factor-urogastrone. Nature. 1979 Feb 1;277(5695):403–405. doi: 10.1038/277403a0. [DOI] [PubMed] [Google Scholar]
  23. Hollenberg M. D., Cuatrecasas P. Epidermal growth factor: receptors in human fibroblasts and modulation of action by cholera toxin. Proc Natl Acad Sci U S A. 1973 Oct;70(10):2964–2968. doi: 10.1073/pnas.70.10.2964. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Hollenberg M. D., Cuatrecasas P. Insulin and epidermal growth factor. Human fibroblast receptors related to deoxyribonucleic acid synthesis and amino acid uptake. J Biol Chem. 1975 May 25;250(10):3845–3853. [PubMed] [Google Scholar]
  25. Hunter T., Sefton B. M. Transforming gene product of Rous sarcoma virus phosphorylates tyrosine. Proc Natl Acad Sci U S A. 1980 Mar;77(3):1311–1315. doi: 10.1073/pnas.77.3.1311. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Hynes R. O. Alteration of cell-surface proteins by viral transformation and by proteolysis. Proc Natl Acad Sci U S A. 1973 Nov;70(11):3170–3174. doi: 10.1073/pnas.70.11.3170. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. King L. E., Jr, Carpenter G., Cohen S. Characterization by electrophoresis of epidermal growth factor stimulated phosphorylation using A-431 membranes. Biochemistry. 1980 Apr 1;19(7):1524–1528. doi: 10.1021/bi00548a040. [DOI] [PubMed] [Google Scholar]
  28. Knowles B. B., Solter D., Trinchieri G., Maloney K. M., Ford S. R., Aden D. P. Complement-mediated antiserum cytotoxic reactions to human chromosome 7 coded antigen(s): immunoselection of rearranged human chromosome 7 in human-mouse somatic cell hybrids. J Exp Med. 1977 Feb 1;145(2):314–326. doi: 10.1084/jem.145.2.314. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  30. Linsley P. S., Blifeld C., Wrann M., Fox C. F. Direct linkage of epidermal growth factor to its receptor. Nature. 1979 Apr 19;278(5706):745–748. doi: 10.1038/278745a0. [DOI] [PubMed] [Google Scholar]
  31. O'Farrell P. H. High resolution two-dimensional electrophoresis of proteins. J Biol Chem. 1975 May 25;250(10):4007–4021. [PMC free article] [PubMed] [Google Scholar]
  32. Plimmer R. H. Esters of phosphoric acid: Phosphoryl hydroxyamino-acids. Biochem J. 1941 Apr;35(4):461–469. doi: 10.1042/bj0350461. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Ross A. H., Baltimore D., Eisen H. N. Phosphotyrosine-containing proteins isolated by affinity chromatography with antibodies to a synthetic hapten. Nature. 1981 Dec 17;294(5842):654–656. doi: 10.1038/294654a0. [DOI] [PubMed] [Google Scholar]
  34. Sahyoun N., Hock R. A., Hollenberg M. D. Insulin and epidermal growth factor-urogastrone: affinity crosslinking to specific binding sites in rat liver membranes. Proc Natl Acad Sci U S A. 1978 Apr;75(4):1675–1679. doi: 10.1073/pnas.75.4.1675. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Savage C. R., Jr, Cohen S. Epidermal growth factor and a new derivative. Rapid isolation procedures and biological and chemical characterization. J Biol Chem. 1972 Dec 10;247(23):7609–7611. [PubMed] [Google Scholar]
  36. Schreiber A. B., Lax I., Yarden Y., Eshhar Z., Schlessinger J. Monoclonal antibodies against receptor for epidermal growth factor induce early and delayed effects of epidermal growth factor. Proc Natl Acad Sci U S A. 1981 Dec;78(12):7535–7539. doi: 10.1073/pnas.78.12.7535. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Shimizu N., Behzadian M. A., Shimizu Y. Genetics of cell surface receptors for bioactive polypeptides: binding of epidermal growth factor is associated with the presence of human chromosome 7 in human-mouse cell hybrids. Proc Natl Acad Sci U S A. 1980 Jun;77(6):3600–3604. doi: 10.1073/pnas.77.6.3600. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Ushiro H., Cohen S. Identification of phosphotyrosine as a product of epidermal growth factor-activated protein kinase in A-431 cell membranes. J Biol Chem. 1980 Sep 25;255(18):8363–8365. [PubMed] [Google Scholar]

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