Skip to main content
PMC home page
Molecular and Cellular Biology logoLink to Molecular and Cellular Biology
. 1994 Jul;14(7):4843–4854. doi: 10.1128/mcb.14.7.4843

Tyrosine 569 in the c-Fms juxtamembrane domain is essential for kinase activity and macrophage colony-stimulating factor-dependent internalization.

G M Myles 1, C S Brandt 1, K Carlberg 1, L R Rohrschneider 1
PMCID: PMC358857  PMID: 8007983

Abstract

The receptor (Fms) for macrophage colony-stimulating factor (M-CSF) is a member of the tyrosine kinase class of growth factor receptors. It maintains survival, stimulates growth, and drives differentiation of the macrophage lineage of hematopoietic cells. Fms accumulates on the cell surface and becomes activated for signal transduction after M-CSF binding and is then internalized via endocytosis for eventual degradation in lysosomes. We have investigated the mechanism of endocytosis as part of the overall signaling process of this receptor and have identified an amino acid segment near the cytoplasmic juxtamembrane region surrounding tyrosine 569 that is important for internalization. Mutation of tyrosine 569 to alanine (Y569A) eliminates ligand-induced rapid endocytosis of receptor molecules. The mutant Fms Y569A also lacks tyrosine kinase activity; however, tyrosine kinase activity is not essential for endocytosis because the kinase inactive receptor Fms K614A does undergo ligand-induced endocytosis, albeit at a reduced rate. Mutation of tyrosine 569 to phenylalanine had no effect on the M-CSF-induced endocytosis of Fms, and a four-amino-acid sequence containing Y-569 could support endocytosis when transferred into the cytoplasmic juxtamembrane region of a glycophorin A construct. These results indicate that tyrosine 569 within the juxtamembrane region of Fms is part of a signal recognition sequence for endocytosis that does not require tyrosine phosphorylation at this site and that this domain also influences the kinase activity of the receptor. These results are consistent with a ligand-dependent step in recognition of the potential cryptic internalization signal.

Full text

PDF
4843

Images in this article

4846Image
on p.4846
4846Image
on p.4846
4847Image
on p.4847
4848Image
on p.4848
4849Image
on p.4849
4850Image
on p.4850
4850Image
on p.4850
4851Image
on p.4851
4851Image
on p.4851
4852Image
on p.4852
4853Image
on p.4853

Selected References

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

  1. Backer J. M., Shoelson S. E., Weiss M. A., Hua Q. X., Cheatham R. B., Haring E., Cahill D. C., White M. F. The insulin receptor juxtamembrane region contains two independent tyrosine/beta-turn internalization signals. J Cell Biol. 1992 Aug;118(4):831–839. doi: 10.1083/jcb.118.4.831. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bansal A., Gierasch L. M. The NPXY internalization signal of the LDL receptor adopts a reverse-turn conformation. Cell. 1991 Dec 20;67(6):1195–1201. doi: 10.1016/0092-8674(91)90295-a. [DOI] [PubMed] [Google Scholar]
  3. Breitfeld P. P., Casanova J. E., McKinnon W. C., Mostov K. E. Deletions in the cytoplasmic domain of the polymeric immunoglobulin receptor differentially affect endocytotic rate and postendocytotic traffic. J Biol Chem. 1990 Aug 15;265(23):13750–13757. [PubMed] [Google Scholar]
  4. Cama A., Quon M. J., de la Luz Sierra M., Taylor S. I. Substitution of isoleucine for methionine at position 1153 in the beta-subunit of the human insulin receptor. A mutation that impairs receptor tyrosine kinase activity, receptor endocytosis, and insulin action. J Biol Chem. 1992 Apr 25;267(12):8383–8389. [PubMed] [Google Scholar]
  5. Canfield W. M., Johnson K. F., Ye R. D., Gregory W., Kornfeld S. Localization of the signal for rapid internalization of the bovine cation-independent mannose 6-phosphate/insulin-like growth factor-II receptor to amino acids 24-29 of the cytoplasmic tail. J Biol Chem. 1991 Mar 25;266(9):5682–5688. [PubMed] [Google Scholar]
  6. Carlberg K., Tapley P., Haystead C., Rohrschneider L. The role of kinase activity and the kinase insert region in ligand-induced internalization and degradation of the c-fms protein. EMBO J. 1991 Apr;10(4):877–883. doi: 10.1002/j.1460-2075.1991.tb08020.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Chen W. J., Goldstein J. L., Brown M. S. NPXY, a sequence often found in cytoplasmic tails, is required for coated pit-mediated internalization of the low density lipoprotein receptor. J Biol Chem. 1990 Feb 25;265(6):3116–3123. [PubMed] [Google Scholar]
  8. Chen W. S., Lazar C. S., Lund K. A., Welsh J. B., Chang C. P., Walton G. M., Der C. J., Wiley H. S., Gill G. N., Rosenfeld M. G. Functional independence of the epidermal growth factor receptor from a domain required for ligand-induced internalization and calcium regulation. Cell. 1989 Oct 6;59(1):33–43. doi: 10.1016/0092-8674(89)90867-2. [DOI] [PubMed] [Google Scholar]
  9. Claesson-Welsh L., Eriksson A., Westermark B., Heldin C. H. cDNA cloning and expression of the human A-type platelet-derived growth factor (PDGF) receptor establishes structural similarity to the B-type PDGF receptor. Proc Natl Acad Sci U S A. 1989 Jul;86(13):4917–4921. doi: 10.1073/pnas.86.13.4917. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Collawn J. F., Stangel M., Kuhn L. A., Esekogwu V., Jing S. Q., Trowbridge I. S., Tainer J. A. Transferrin receptor internalization sequence YXRF implicates a tight turn as the structural recognition motif for endocytosis. Cell. 1990 Nov 30;63(5):1061–1072. doi: 10.1016/0092-8674(90)90509-d. [DOI] [PubMed] [Google Scholar]
  11. Davis C. G., Lehrman M. A., Russell D. W., Anderson R. G., Brown M. S., Goldstein J. L. The J.D. mutation in familial hypercholesterolemia: amino acid substitution in cytoplasmic domain impedes internalization of LDL receptors. Cell. 1986 Apr 11;45(1):15–24. doi: 10.1016/0092-8674(86)90533-7. [DOI] [PubMed] [Google Scholar]
  12. Davis C. G., van Driel I. R., Russell D. W., Brown M. S., Goldstein J. L. The low density lipoprotein receptor. Identification of amino acids in cytoplasmic domain required for rapid endocytosis. J Biol Chem. 1987 Mar 25;262(9):4075–4082. [PubMed] [Google Scholar]
  13. Eberle W., Sander C., Klaus W., Schmidt B., von Figura K., Peters C. The essential tyrosine of the internalization signal in lysosomal acid phosphatase is part of a beta turn. Cell. 1991 Dec 20;67(6):1203–1209. doi: 10.1016/0092-8674(91)90296-b. [DOI] [PubMed] [Google Scholar]
  14. Glickman J. N., Conibear E., Pearse B. M. Specificity of binding of clathrin adaptors to signals on the mannose-6-phosphate/insulin-like growth factor II receptor. EMBO J. 1989 Apr;8(4):1041–1047. doi: 10.1002/j.1460-2075.1989.tb03471.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Goldstein J. L., Brown M. S., Anderson R. G., Russell D. W., Schneider W. J. Receptor-mediated endocytosis: concepts emerging from the LDL receptor system. Annu Rev Cell Biol. 1985;1:1–39. doi: 10.1146/annurev.cb.01.110185.000245. [DOI] [PubMed] [Google Scholar]
  16. Hanks S. K., Quinn A. M., Hunter T. The protein kinase family: conserved features and deduced phylogeny of the catalytic domains. Science. 1988 Jul 1;241(4861):42–52. doi: 10.1126/science.3291115. [DOI] [PubMed] [Google Scholar]
  17. Helin K., Beguinot L. Internalization and down-regulation of the human epidermal growth factor receptor are regulated by the carboxyl-terminal tyrosines. J Biol Chem. 1991 May 5;266(13):8363–8368. [PubMed] [Google Scholar]
  18. Jing S. Q., Spencer T., Miller K., Hopkins C., Trowbridge I. S. Role of the human transferrin receptor cytoplasmic domain in endocytosis: localization of a specific signal sequence for internalization. J Cell Biol. 1990 Feb;110(2):283–294. doi: 10.1083/jcb.110.2.283. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Keating M. T., Escobedo J. A., Williams L. T. Ligand activation causes a phosphorylation-dependent change in platelet-derived growth factor receptor conformation. J Biol Chem. 1988 Sep 15;263(26):12805–12808. [PubMed] [Google Scholar]
  20. Ktistakis N. T., Thomas D., Roth M. G. Characteristics of the tyrosine recognition signal for internalization of transmembrane surface glycoproteins. J Cell Biol. 1990 Oct;111(4):1393–1407. doi: 10.1083/jcb.111.4.1393. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Kudo S., Fukuda M. Structural organization of glycophorin A and B genes: glycophorin B gene evolved by homologous recombination at Alu repeat sequences. Proc Natl Acad Sci U S A. 1989 Jun;86(12):4619–4623. doi: 10.1073/pnas.86.12.4619. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Kunkel T. A., Roberts J. D., Zakour R. A. Rapid and efficient site-specific mutagenesis without phenotypic selection. Methods Enzymol. 1987;154:367–382. doi: 10.1016/0076-6879(87)54085-x. [DOI] [PubMed] [Google Scholar]
  23. 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]
  24. Lazarovits J., Roth M. A single amino acid change in the cytoplasmic domain allows the influenza virus hemagglutinin to be endocytosed through coated pits. Cell. 1988 Jun 3;53(5):743–752. doi: 10.1016/0092-8674(88)90092-x. [DOI] [PubMed] [Google Scholar]
  25. Lobel P., Fujimoto K., Ye R. D., Griffiths G., Kornfeld S. Mutations in the cytoplasmic domain of the 275 kd mannose 6-phosphate receptor differentially alter lysosomal enzyme sorting and endocytosis. Cell. 1989 Jun 2;57(5):787–796. doi: 10.1016/0092-8674(89)90793-9. [DOI] [PubMed] [Google Scholar]
  26. Lyman S. D., Rohrschneider L. R. The kinase activity of the v-fms encoded protein has a low pH optimum. Oncogene Res. 1989;4(2):149–155. [PubMed] [Google Scholar]
  27. Manger R., Najita L., Nichols E. J., Hakomori S., Rohrschneider L. Cell surface expression of the McDonough strain of feline sarcoma virus fms gene product (gp 140fms). Cell. 1984 Dec;39(2 Pt 1):327–337. doi: 10.1016/0092-8674(84)90011-4. [DOI] [PubMed] [Google Scholar]
  28. Matthews W., Jordan C. T., Wiegand G. W., Pardoll D., Lemischka I. R. A receptor tyrosine kinase specific to hematopoietic stem and progenitor cell-enriched populations. Cell. 1991 Jun 28;65(7):1143–1152. doi: 10.1016/0092-8674(91)90010-v. [DOI] [PubMed] [Google Scholar]
  29. McClain D. A. Mechanism and role of insulin receptor endocytosis. Am J Med Sci. 1992 Sep;304(3):192–201. doi: 10.1097/00000441-199209000-00009. [DOI] [PubMed] [Google Scholar]
  30. McGraw T. E., Maxfield F. R. Human transferrin receptor internalization is partially dependent upon an aromatic amino acid on the cytoplasmic domain. Cell Regul. 1990 Mar;1(4):369–377. doi: 10.1091/mbc.1.4.369. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Mooibroek M. J., Cheng H. C., Wang J. H. Differential in vitro phosphorylation of clathrin light chains by the epidermal growth factor receptor-associated protein tyrosine kinase and a pp60c-src-related spleen tyrosine kinase. Arch Biochem Biophys. 1992 Feb 1;292(2):448–455. doi: 10.1016/0003-9861(92)90015-o. [DOI] [PubMed] [Google Scholar]
  32. Ohtsuka M., Roussel M. F., Sherr C. J., Downing J. R. Ligand-induced phosphorylation of the colony-stimulating factor 1 receptor can occur through an intermolecular reaction that triggers receptor down modulation. Mol Cell Biol. 1990 Apr;10(4):1664–1671. doi: 10.1128/mcb.10.4.1664. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Pearse B. M. Receptors compete for adaptors found in plasma membrane coated pits. EMBO J. 1988 Nov;7(11):3331–3336. doi: 10.1002/j.1460-2075.1988.tb03204.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Qiu F. H., Ray P., Brown K., Barker P. E., Jhanwar S., Ruddle F. H., Besmer P. Primary structure of c-kit: relationship with the CSF-1/PDGF receptor kinase family--oncogenic activation of v-kit involves deletion of extracellular domain and C terminus. EMBO J. 1988 Apr;7(4):1003–1011. doi: 10.1002/j.1460-2075.1988.tb02907.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Reedijk M., Liu X., van der Geer P., Letwin K., Waterfield M. D., Hunter T., Pawson T. Tyr721 regulates specific binding of the CSF-1 receptor kinase insert to PI 3'-kinase SH2 domains: a model for SH2-mediated receptor-target interactions. EMBO J. 1992 Apr;11(4):1365–1372. doi: 10.1002/j.1460-2075.1992.tb05181.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Rettenmier C. W., Roussel M. F. Differential processing of colony-stimulating factor 1 precursors encoded by two human cDNAs. Mol Cell Biol. 1988 Nov;8(11):5026–5034. doi: 10.1128/mcb.8.11.5026. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Rohrschneider L. R., Metcalf D. Induction of macrophage colony-stimulating factor-dependent growth and differentiation after introduction of the murine c-fms gene into FDC-P1 cells. Mol Cell Biol. 1989 Nov;9(11):5081–5092. doi: 10.1128/mcb.9.11.5081. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Rohrschneider L. R., Rothwell V. M., Nicola N. A. Transformation of murine fibroblasts by a retrovirus encoding the murine c-fms proto-oncogene. Oncogene. 1989 Aug;4(8):1015–1022. [PubMed] [Google Scholar]
  39. Rosnet O., Marchetto S., deLapeyriere O., Birnbaum D. Murine Flt3, a gene encoding a novel tyrosine kinase receptor of the PDGFR/CSF1R family. Oncogene. 1991 Sep;6(9):1641–1650. [PubMed] [Google Scholar]
  40. Roussel M. F., Dull T. J., Rettenmier C. W., Ralph P., Ullrich A., Sherr C. J. Transforming potential of the c-fms proto-oncogene (CSF-1 receptor). Nature. 1987 Feb 5;325(6104):549–552. doi: 10.1038/325549a0. [DOI] [PubMed] [Google Scholar]
  41. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Sbraccia P., Wong K. Y., Brunetti A., Rafaeloff R., Trischitta V., Hawley D. M., Goldfine I. D. Insulin down-regulates insulin receptor number and up-regulates insulin receptor affinity in cells expressing a tyrosine kinase-defective insulin receptor. J Biol Chem. 1990 Mar 25;265(9):4902–4907. [PubMed] [Google Scholar]
  43. Sherr C. J., Rettenmier C. W., Sacca R., Roussel M. F., Look A. T., Stanley E. R. The c-fms proto-oncogene product is related to the receptor for the mononuclear phagocyte growth factor, CSF-1. Cell. 1985 Jul;41(3):665–676. doi: 10.1016/s0092-8674(85)80047-7. [DOI] [PubMed] [Google Scholar]
  44. Sorkin A., Carpenter G. Interaction of activated EGF receptors with coated pit adaptins. Science. 1993 Jul 30;261(5121):612–615. doi: 10.1126/science.8342026. [DOI] [PubMed] [Google Scholar]
  45. Sorkin A., Waters C., Overholser K. A., Carpenter G. Multiple autophosphorylation site mutations of the epidermal growth factor receptor. Analysis of kinase activity and endocytosis. J Biol Chem. 1991 May 5;266(13):8355–8362. [PubMed] [Google Scholar]
  46. Sorkin A., Westermark B., Heldin C. H., Claesson-Welsh L. Effect of receptor kinase inactivation on the rate of internalization and degradation of PDGF and the PDGF beta-receptor. J Cell Biol. 1991 Feb;112(3):469–478. doi: 10.1083/jcb.112.3.469. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Tapley P., Kazlauskas A., Cooper J. A., Rohrschneider L. R. Macrophage colony-stimulating factor-induced tyrosine phosphorylation of c-fms proteins expressed in FDC-P1 and BALB/c 3T3 cells. Mol Cell Biol. 1990 Jun;10(6):2528–2538. doi: 10.1128/mcb.10.6.2528. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Trowbridge I. S. Endocytosis and signals for internalization. Curr Opin Cell Biol. 1991 Aug;3(4):634–641. doi: 10.1016/0955-0674(91)90034-v. [DOI] [PubMed] [Google Scholar]
  49. Ullrich A., Schlessinger J. Signal transduction by receptors with tyrosine kinase activity. Cell. 1990 Apr 20;61(2):203–212. doi: 10.1016/0092-8674(90)90801-k. [DOI] [PubMed] [Google Scholar]
  50. Wang Z. E., Myles G. M., Brandt C. S., Lioubin M. N., Rohrschneider L. Identification of the ligand-binding regions in the macrophage colony-stimulating factor receptor extracellular domain. Mol Cell Biol. 1993 Sep;13(9):5348–5359. doi: 10.1128/mcb.13.9.5348. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Woolford J., McAuliffe A., Rohrschneider L. R. Activation of the feline c-fms proto-oncogene: multiple alterations are required to generate a fully transformed phenotype. Cell. 1988 Dec 23;55(6):965–977. doi: 10.1016/0092-8674(88)90242-5. [DOI] [PubMed] [Google Scholar]
  52. Yarden Y., Escobedo J. A., Kuang W. J., Yang-Feng T. L., Daniel T. O., Tremble P. M., Chen E. Y., Ando M. E., Harkins R. N., Francke U. Structure of the receptor for platelet-derived growth factor helps define a family of closely related growth factor receptors. Nature. 1986 Sep 18;323(6085):226–232. doi: 10.1038/323226a0. [DOI] [PubMed] [Google Scholar]
  53. Yeung Y. G., Berg K. L., Pixley F. J., Angeletti R. H., Stanley E. R. Protein tyrosine phosphatase-1C is rapidly phosphorylated in tyrosine in macrophages in response to colony stimulating factor-1. J Biol Chem. 1992 Nov 25;267(33):23447–23450. [PubMed] [Google Scholar]
  54. Zoller M. J., Smith M. Oligonucleotide-directed mutagenesis of DNA fragments cloned into M13 vectors. Methods Enzymol. 1983;100:468–500. doi: 10.1016/0076-6879(83)00074-9. [DOI] [PubMed] [Google Scholar]
  55. van der Geer P., Hunter T. Identification of tyrosine 706 in the kinase insert as the major colony-stimulating factor 1 (CSF-1)-stimulated autophosphorylation site in the CSF-1 receptor in a murine macrophage cell line. Mol Cell Biol. 1990 Jun;10(6):2991–3002. doi: 10.1128/mcb.10.6.2991. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Molecular and Cellular Biology are provided here courtesy of Taylor & Francis

RESOURCES