Skip to main content
NCBI home page
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
. 1995 Mar 14;92(6):2229–2233. doi: 10.1073/pnas.92.6.2229

A region in the cytosolic domain of the epidermal growth factor receptor antithetically regulates the stimulatory and inhibitory guanine nucleotide-binding regulatory proteins of adenylyl cyclase.

H Sun 1, J M Seyer 1, T B Patel 1
PMCID: PMC42457  PMID: 7892252

Abstract

Epidermal growth factor (EGF) stimulates adenylyl cyclase in the heart via activation of the stimulatory GTP-binding protein Gs. Therefore, employing peptides corresponding to regions in the cytosolic domain of the EGF receptor, we have investigated the ability of sequences within the EGF receptor to activate Gs. A 13-aa peptide (EGFR-13) corresponding to the juxtamembrane region in the cytosolic domain of the EGF receptor stimulated GTP binding and GTPase activity of Gs. This peptide did not stimulate GTP binding to Gi but increased the GTPase activity of this protein. Additionally, phosphorylation of the protein kinase C site (threonine residue) within EGFR-13 decreased the ability of the peptide to stimulate Gs and increase GTPase activity of Gi. Further, in functional assays of Gs employing S49 cyc- cell membranes, EGFR-13 increased the ability of Gs to stimulate adenylyl cyclase; phospho-EGFR-13 and a 14-aa peptide corresponding to a sequence in the cytosolic domain of the EGF receptor did not alter the functional activity of Gs. Hence, the juxtamembrane region of the EGF receptor can activate Gs and, by stimulating GTPase activity of Gi, inactivates this latter G protein. Phosphorylation of the threonine residue within this region attenuates the activity of the peptide as a modulator of G-protein function.

Full text

PDF
2229

Images in this article

2232Fig. 4
on p.2232

Selected References

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

  1. Arshavsky VYu, Bownds M. D. Regulation of deactivation of photoreceptor G protein by its target enzyme and cGMP. Nature. 1992 Jun 4;357(6377):416–417. doi: 10.1038/357416a0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Beguinot L., Hanover J. A., Ito S., Richert N. D., Willingham M. C., Pastan I. Phorbol esters induce transient internalization without degradation of unoccupied epidermal growth factor receptors. Proc Natl Acad Sci U S A. 1985 May;82(9):2774–2778. doi: 10.1073/pnas.82.9.2774. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bosch F., Bouscarel B., Slaton J., Blackmore P. F., Exton J. H. Epidermal growth factor mimics insulin effects in rat hepatocytes. Biochem J. 1986 Nov 1;239(3):523–530. doi: 10.1042/bj2390523. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Brandt D. R., Asano T., Pedersen S. E., Ross E. M. Reconstitution of catecholamine-stimulated guanosinetriphosphatase activity. Biochemistry. 1983 Sep 13;22(19):4357–4362. doi: 10.1021/bi00288a002. [DOI] [PubMed] [Google Scholar]
  5. Cadena D. L., Chan C. L., Gill G. N. The intracellular tyrosine kinase domain of the epidermal growth factor receptor undergoes a conformational change upon autophosphorylation. J Biol Chem. 1994 Jan 7;269(1):260–265. [PubMed] [Google Scholar]
  6. Carpenter G., Cohen S. Epidermal growth factor. J Biol Chem. 1990 May 15;265(14):7709–7712. [PubMed] [Google Scholar]
  7. Carpenter G. Receptors for epidermal growth factor and other polypeptide mitogens. Annu Rev Biochem. 1987;56:881–914. doi: 10.1146/annurev.bi.56.070187.004313. [DOI] [PubMed] [Google Scholar]
  8. Downward J., Waterfield M. D., Parker P. J. Autophosphorylation and protein kinase C phosphorylation of the epidermal growth factor receptor. Effect on tyrosine kinase activity and ligand binding affinity. J Biol Chem. 1985 Nov 25;260(27):14538–14546. [PubMed] [Google Scholar]
  9. Graziano M. P., Freissmuth M., Gilman A. G. Purification of recombinant Gs alpha. Methods Enzymol. 1991;195:192–202. doi: 10.1016/0076-6879(91)95166-h. [DOI] [PubMed] [Google Scholar]
  10. Hunter T., Ling N., Cooper J. A. Protein kinase C phosphorylation of the EGF receptor at a threonine residue close to the cytoplasmic face of the plasma membrane. Nature. 1984 Oct 4;311(5985):480–483. doi: 10.1038/311480a0. [DOI] [PubMed] [Google Scholar]
  11. Iyengar R., Bhat M. K., Riser M. E., Birnbaumer L. Receptor-specific desensitization of the S49 lymphoma cell adenylyl cyclase. Unaltered behavior of the regulatory component. J Biol Chem. 1981 May 25;256(10):4810–4815. [PubMed] [Google Scholar]
  12. Koch C. A., Anderson D., Moran M. F., Ellis C., Pawson T. SH2 and SH3 domains: elements that control interactions of cytoplasmic signaling proteins. Science. 1991 May 3;252(5006):668–674. doi: 10.1126/science.1708916. [DOI] [PubMed] [Google Scholar]
  13. Markby D. W., Onrust R., Bourne H. R. Separate GTP binding and GTPase activating domains of a G alpha subunit. Science. 1993 Dec 17;262(5141):1895–1901. doi: 10.1126/science.8266082. [DOI] [PubMed] [Google Scholar]
  14. Moolenaar W. H., Aerts R. J., Tertoolen L. G., de Laat S. W. The epidermal growth factor-induced calcium signal in A431 cells. J Biol Chem. 1986 Jan 5;261(1):279–284. [PubMed] [Google Scholar]
  15. Moule S. K., McGivan J. D. Epidermal growth factor, like glucagon, exerts a short-term stimulation of alanine transport in rat hepatocytes. Biochem J. 1987 Oct 1;247(1):233–235. doi: 10.1042/bj2470233. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Mumby S., Pang I. H., Gilman A. G., Sternweis P. C. Chromatographic resolution and immunologic identification of the alpha 40 and alpha 41 subunits of guanine nucleotide-binding regulatory proteins from bovine brain. J Biol Chem. 1988 Feb 5;263(4):2020–2026. [PubMed] [Google Scholar]
  17. Nair B. G., Parikh B., Milligan G., Patel T. B. Gs alpha mediates epidermal growth factor-elicited stimulation of rat cardiac adenylate cyclase. J Biol Chem. 1990 Dec 5;265(34):21317–21322. [PubMed] [Google Scholar]
  18. Nair B. G., Patel T. B. Regulation of cardiac adenylyl cyclase by epidermal growth factor (EGF). Role of EGF receptor protein tyrosine kinase activity. Biochem Pharmacol. 1993 Oct 5;46(7):1239–1245. doi: 10.1016/0006-2952(93)90473-a. [DOI] [PubMed] [Google Scholar]
  19. Nair B. G., Rashed H. M., Patel T. B. Epidermal growth factor produces inotropic and chronotropic effects in rat hearts by increasing cyclic AMP accumulation. Growth Factors. 1993;8(1):41–48. doi: 10.3109/08977199309029133. [DOI] [PubMed] [Google Scholar]
  20. Nair B. G., Rashed H. M., Patel T. B. Epidermal growth factor stimulates rat cardiac adenylate cyclase through a GTP-binding regulatory protein. Biochem J. 1989 Dec 1;264(2):563–571. doi: 10.1042/bj2640563. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Neer E. J., Lok J. M., Wolf L. G. Purification and properties of the inhibitory guanine nucleotide regulatory unit of brain adenylate cyclase. J Biol Chem. 1984 Nov 25;259(22):14222–14229. [PubMed] [Google Scholar]
  22. Northup J. K., Smigel M. D., Gilman A. G. The guanine nucleotide activating site of the regulatory component of adenylate cyclase. Identification by ligand binding. J Biol Chem. 1982 Oct 10;257(19):11416–11423. [PubMed] [Google Scholar]
  23. Okamoto T., Katada T., Murayama Y., Ui M., Ogata E., Nishimoto I. A simple structure encodes G protein-activating function of the IGF-II/mannose 6-phosphate receptor. Cell. 1990 Aug 24;62(4):709–717. doi: 10.1016/0092-8674(90)90116-v. [DOI] [PubMed] [Google Scholar]
  24. Okamoto T., Murayama Y., Hayashi Y., Inagaki M., Ogata E., Nishimoto I. Identification of a Gs activator region of the beta 2-adrenergic receptor that is autoregulated via protein kinase A-dependent phosphorylation. Cell. 1991 Nov 15;67(4):723–730. doi: 10.1016/0092-8674(91)90067-9. [DOI] [PubMed] [Google Scholar]
  25. Parker E. M., Ross E. M. Truncation of the extended carboxyl-terminal domain increases the expression and regulatory activity of the avian beta-adrenergic receptor. J Biol Chem. 1991 May 25;266(15):9987–9996. [PubMed] [Google Scholar]
  26. Pike L. J., Eakes A. T. Epidermal growth factor stimulates the production of phosphatidylinositol monophosphate and the breakdown of polyphosphoinositides in A431 cells. J Biol Chem. 1987 Feb 5;262(4):1644–1651. [PubMed] [Google Scholar]
  27. Rashed S. M., Patel T. B. Regulation of hepatic energy metabolism by epidermal growth factor. Eur J Biochem. 1991 May 8;197(3):805–813. doi: 10.1111/j.1432-1033.1991.tb15975.x. [DOI] [PubMed] [Google Scholar]
  28. Sternweis P. C., Gilman A. G. Reconstitution of catecholamine-sensitive adenylate cyclase. Reconstitution of the uncoupled variant of the S40 lymphoma cell. J Biol Chem. 1979 May 10;254(9):3333–3340. [PubMed] [Google Scholar]
  29. Ullrich A., Coussens L., Hayflick J. S., Dull T. J., Gray A., Tam A. W., Lee J., Yarden Y., Libermann T. A., Schlessinger J. Human epidermal growth factor receptor cDNA sequence and aberrant expression of the amplified gene in A431 epidermoid carcinoma cells. 1984 May 31-Jun 6Nature. 309(5967):418–425. doi: 10.1038/309418a0. [DOI] [PubMed] [Google Scholar]
  30. Wahl M., Carpenter G. Regulation of epidermal growth factor-stimulated formation of inositol phosphates in A-431 cells by calcium and protein kinase C. J Biol Chem. 1988 Jun 5;263(16):7581–7590. [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