Skip to main content
NCBI home page
The EMBO Journal logoLink to The EMBO Journal
. 1991 Aug;10(8):1969–1977. doi: 10.1002/j.1460-2075.1991.tb07726.x

Mannosamine, a novel inhibitor of glycosylphosphatidylinositol incorporation into proteins.

M P Lisanti 1, M C Field 1, I W Caras 1, A K Menon 1, E Rodriguez-Boulan 1
PMCID: PMC452876  PMID: 1829673

Abstract

Mannosamine (2-amino-2-deoxy D-mannose) is shown here to block the incorporation of glycosylphosphatidylinositol (GPI) into GPI-anchored proteins. The amino sugar drastically reduced the surface expression of a recombinant GPI-anchored protein in polarized MDCK cells, converted this apical membrane-bound protein to an unpolarized secretory product and blocked the expression of endogenous GPI-anchored proteins. Furthermore, it specifically inhibited the incorporation of [3H]ethanolamine (a GPI component) into mammalian and trypanosomal GPI-anchored proteins and into a well characterized GPI-lipid of Trypanosoma brucei. These results suggest that mannosamine converted an apical GPI-anchored protein to a non-polarized secretory product by depleting transfer competent GPI-precursor lipids. Our inhibitor studies provide new independent evidence for the apical targeting role of GPI in polarized epithelia and open the way towards a greater understanding of the functional role of GPI in membrane trafficking and cell regulation.

Full text

PDF
1969

Images in this article

1970Image
on p.1970
1970Image
on p.1970
1971Image
on p.1971
1972Image
on p.1972
1972Image
on p.1972
1973Image
on p.1973
1973Image
on p.1973
1974Image
on p.1974
1974Image
on p.1974

Selected References

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

  1. Bailey C. A., Gerber L., Howard A. D., Udenfriend S. Processing at the carboxyl terminus of nascent placental alkaline phosphatase in a cell-free system: evidence for specific cleavage of a signal peptide. Proc Natl Acad Sci U S A. 1989 Jan;86(1):22–26. doi: 10.1073/pnas.86.1.22. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bangs J. D., Andrews N. W., Hart G. W., Englund P. T. Posttranslational modification and intracellular transport of a trypanosome variant surface glycoprotein. J Cell Biol. 1986 Jul;103(1):255–263. doi: 10.1083/jcb.103.1.255. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bangs J. D., Hereld D., Krakow J. L., Hart G. W., Englund P. T. Rapid processing of the carboxyl terminus of a trypanosome variant surface glycoprotein. Proc Natl Acad Sci U S A. 1985 May;82(10):3207–3211. doi: 10.1073/pnas.82.10.3207. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Brown D. A., Crise B., Rose J. K. Mechanism of membrane anchoring affects polarized expression of two proteins in MDCK cells. Science. 1989 Sep 29;245(4925):1499–1501. doi: 10.1126/science.2571189. [DOI] [PubMed] [Google Scholar]
  5. Caras I. W., Weddell G. N., Davitz M. A., Nussenzweig V., Martin D. W., Jr Signal for attachment of a phospholipid membrane anchor in decay accelerating factor. Science. 1987 Nov 27;238(4831):1280–1283. doi: 10.1126/science.2446389. [DOI] [PubMed] [Google Scholar]
  6. Caras I. W., Weddell G. N., Williams S. R. Analysis of the signal for attachment of a glycophospholipid membrane anchor. J Cell Biol. 1989 Apr;108(4):1387–1396. doi: 10.1083/jcb.108.4.1387. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Chamberlain J. P. Fluorographic detection of radioactivity in polyacrylamide gels with the water-soluble fluor, sodium salicylate. Anal Biochem. 1979 Sep 15;98(1):132–135. doi: 10.1016/0003-2697(79)90716-4. [DOI] [PubMed] [Google Scholar]
  8. Chapman A. E., Calhoun J. C., 4th Effects of glucose starvation and puromycin treatment on lipid-linked oligosaccharide precursors and biosynthetic enzymes in Chinese hamster ovary cells in vivo and in vitro. Arch Biochem Biophys. 1988 Jan;260(1):320–333. doi: 10.1016/0003-9861(88)90456-0. [DOI] [PubMed] [Google Scholar]
  9. Clayton C. E., Mowatt M. R. The procyclic acidic repetitive proteins of Trypanosoma brucei. Purification and post-translational modification. J Biol Chem. 1989 Sep 5;264(25):15088–15093. [PubMed] [Google Scholar]
  10. Conzelmann A., Spiazzi A., Bron C. Glycolipid anchors are attached to Thy-1 glycoprotein rapidly after translation. Biochem J. 1987 Sep 15;246(3):605–610. doi: 10.1042/bj2460605. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Conzelmann A., Spiazzi A., Bron C., Hyman R. No glycolipid anchors are added to Thy-1 glycoprotein in Thy-1-negative mutant thymoma cells of four different complementation classes. Mol Cell Biol. 1988 Feb;8(2):674–678. doi: 10.1128/mcb.8.2.674. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Conzelmann A., Spiazzi A., Hyman R., Bron C. Anchoring of membrane proteins via phosphatidylinositol is deficient in two classes of Thy-1 negative mutant lymphoma cells. EMBO J. 1986 Dec 1;5(12):3291–3296. doi: 10.1002/j.1460-2075.1986.tb04642.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Cross G. A. Glycolipid anchoring of plasma membrane proteins. Annu Rev Cell Biol. 1990;6:1–39. doi: 10.1146/annurev.cb.06.110190.000245. [DOI] [PubMed] [Google Scholar]
  14. DeGasperi R., Thomas L. J., Sugiyama E., Chang H. M., Beck P. J., Orlean P., Albright C., Waneck G., Sambrook J. F., Warren C. D. Correction of a defect in mammalian GPI anchor biosynthesis by a transfected yeast gene. Science. 1990 Nov 16;250(4983):988–991. doi: 10.1126/science.1978413. [DOI] [PubMed] [Google Scholar]
  15. Doering T. L., Masterson W. J., Englund P. T., Hart G. W. Biosynthesis of the glycosyl phosphatidylinositol membrane anchor of the trypanosome variant surface glycoprotein. Origin of the non-acetylated glucosamine. J Biol Chem. 1989 Jul 5;264(19):11168–11173. [PubMed] [Google Scholar]
  16. Doering T. L., Masterson W. J., Hart G. W., Englund P. T. Biosynthesis of glycosyl phosphatidylinositol membrane anchors. J Biol Chem. 1990 Jan 15;265(2):611–614. [PubMed] [Google Scholar]
  17. Dotti C. G., Parton R. G., Simons K. Polarized sorting of glypiated proteins in hippocampal neurons. Nature. 1991 Jan 10;349(6305):158–161. doi: 10.1038/349158a0. [DOI] [PubMed] [Google Scholar]
  18. Elbein A. D. Glycosylation inhibitors for N-linked glycoproteins. Methods Enzymol. 1987;138:661–709. doi: 10.1016/0076-6879(87)38060-7. [DOI] [PubMed] [Google Scholar]
  19. Fatemi S. H., Tartakoff A. M. Hydrophilic anchor-deficient Thy-1 is secreted by a class E mutant T lymphoma. Cell. 1986 Aug 29;46(5):653–657. doi: 10.1016/0092-8674(86)90340-5. [DOI] [PubMed] [Google Scholar]
  20. Fatemi S. H., Tartakoff A. M. The phenotype of five classes of T lymphoma mutants. Defective glycophospholipid anchoring, rapid degradation, and secretion of Thy-1 glycoprotein. J Biol Chem. 1988 Jan 25;263(3):1288–1294. [PubMed] [Google Scholar]
  21. Ferguson M. A., Duszenko M., Lamont G. S., Overath P., Cross G. A. Biosynthesis of Trypanosoma brucei variant surface glycoproteins. N-glycosylation and addition of a phosphatidylinositol membrane anchor. J Biol Chem. 1986 Jan 5;261(1):356–362. [PubMed] [Google Scholar]
  22. Ferguson M. A., Homans S. W., Dwek R. A., Rademacher T. W. Glycosyl-phosphatidylinositol moiety that anchors Trypanosoma brucei variant surface glycoprotein to the membrane. Science. 1988 Feb 12;239(4841 Pt 1):753–759. doi: 10.1126/science.3340856. [DOI] [PubMed] [Google Scholar]
  23. Ferguson M. A., Williams A. F. Cell-surface anchoring of proteins via glycosyl-phosphatidylinositol structures. Annu Rev Biochem. 1988;57:285–320. doi: 10.1146/annurev.bi.57.070188.001441. [DOI] [PubMed] [Google Scholar]
  24. Frahm R., Schwarz M., Geishauser M. Seltene Hypersupinationsluxation im distalen radioulnaren Gelenk. Rofo. 1989 Jun;150(6):746–748. doi: 10.1055/s-2008-1047119. [DOI] [PubMed] [Google Scholar]
  25. Gorman C. M., Howard B. H., Reeves R. Expression of recombinant plasmids in mammalian cells is enhanced by sodium butyrate. Nucleic Acids Res. 1983 Nov 11;11(21):7631–7648. doi: 10.1093/nar/11.21.7631. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Gottlieb T. A., Beaudry G., Rizzolo L., Colman A., Rindler M., Adesnik M., Sabatini D. D. Secretion of endogenous and exogenous proteins from polarized MDCK cell monolayers. Proc Natl Acad Sci U S A. 1986 Apr;83(7):2100–2104. doi: 10.1073/pnas.83.7.2100. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Green R. F., Meiss H. K., Rodriguez-Boulan E. Glycosylation does not determine segregation of viral envelope proteins in the plasma membrane of epithelial cells. J Cell Biol. 1981 May;89(2):230–239. doi: 10.1083/jcb.89.2.230. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Homans S. W., Ferguson M. A., Dwek R. A., Rademacher T. W., Anand R., Williams A. F. Complete structure of the glycosyl phosphatidylinositol membrane anchor of rat brain Thy-1 glycoprotein. Nature. 1988 May 19;333(6170):269–272. doi: 10.1038/333269a0. [DOI] [PubMed] [Google Scholar]
  29. Hooper N. M., Turner A. J. Ectoenzymes of the kidney microvillar membrane. Differential solubilization by detergents can predict a glycosyl-phosphatidylinositol membrane anchor. Biochem J. 1988 Mar 15;250(3):865–869. doi: 10.1042/bj2500865. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Karnieli E., Armoni M., Cohen P., Kanter Y., Rafaeloff R. Reversal of insulin resistance in diabetic rat adipocytes by insulin therapy. Restoration of pool of glucose transporters and enhancement of glucose-transport activity. Diabetes. 1987 Aug;36(8):925–931. doi: 10.2337/diab.36.8.925. [DOI] [PubMed] [Google Scholar]
  31. Kornfeld S. Lysosomal enzyme targeting. Biochem Soc Trans. 1990 Jun;18(3):367–374. doi: 10.1042/bst0180367. [DOI] [PubMed] [Google Scholar]
  32. Lisanti M. P., Caras I. W., Davitz M. A., Rodriguez-Boulan E. A glycophospholipid membrane anchor acts as an apical targeting signal in polarized epithelial cells. J Cell Biol. 1989 Nov;109(5):2145–2156. doi: 10.1083/jcb.109.5.2145. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Lisanti M. P., Caras I. W., Gilbert T., Hanzel D., Rodriguez-Boulan E. Vectorial apical delivery and slow endocytosis of a glycolipid-anchored fusion protein in transfected MDCK cells. Proc Natl Acad Sci U S A. 1990 Oct;87(19):7419–7423. doi: 10.1073/pnas.87.19.7419. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Lisanti M. P., Le Bivic A., Saltiel A. R., Rodriguez-Boulan E. Preferred apical distribution of glycosyl-phosphatidylinositol (GPI) anchored proteins: a highly conserved feature of the polarized epithelial cell phenotype. J Membr Biol. 1990 Feb;113(2):155–167. doi: 10.1007/BF01872889. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Lisanti M. P., Rodriguez-Boulan E. Glycophospholipid membrane anchoring provides clues to the mechanism of protein sorting in polarized epithelial cells. Trends Biochem Sci. 1990 Mar;15(3):113–118. doi: 10.1016/0968-0004(90)90195-h. [DOI] [PubMed] [Google Scholar]
  36. Lisanti M. P., Rodriguez-Boulan E., Saltiel A. R. Emerging functional roles for the glycosyl-phosphatidylinositol membrane protein anchor. J Membr Biol. 1990 Jul;117(1):1–10. doi: 10.1007/BF01871561. [DOI] [PubMed] [Google Scholar]
  37. Lisanti M. P., Sargiacomo M., Graeve L., Saltiel A. R., Rodriguez-Boulan E. Polarized apical distribution of glycosyl-phosphatidylinositol-anchored proteins in a renal epithelial cell line. Proc Natl Acad Sci U S A. 1988 Dec;85(24):9557–9561. doi: 10.1073/pnas.85.24.9557. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Low M. G., Saltiel A. R. Structural and functional roles of glycosyl-phosphatidylinositol in membranes. Science. 1988 Jan 15;239(4837):268–275. doi: 10.1126/science.3276003. [DOI] [PubMed] [Google Scholar]
  39. Low M. G. The glycosyl-phosphatidylinositol anchor of membrane proteins. Biochim Biophys Acta. 1989 Dec 6;988(3):427–454. doi: 10.1016/0304-4157(89)90014-2. [DOI] [PubMed] [Google Scholar]
  40. Masterson W. J., Doering T. L., Hart G. W., Englund P. T. A novel pathway for glycan assembly: biosynthesis of the glycosyl-phosphatidylinositol anchor of the trypanosome variant surface glycoprotein. Cell. 1989 Mar 10;56(5):793–800. doi: 10.1016/0092-8674(89)90684-3. [DOI] [PubMed] [Google Scholar]
  41. Mayor S., Menon A. K., Cross G. A., Ferguson M. A., Dwek R. A., Rademacher T. W. Glycolipid precursors for the membrane anchor of Trypanosoma brucei variant surface glycoproteins. I. Can structure of the phosphatidylinositol-specific phospholipase C sensitive and resistant glycolipids. J Biol Chem. 1990 Apr 15;265(11):6164–6173. [PubMed] [Google Scholar]
  42. Mayor S., Menon A. K., Cross G. A. Glycolipid precursors for the membrane anchor of Trypanosoma brucei variant surface glycoproteins. II. Lipid structures of phosphatidylinositol-specific phospholipase C sensitive and resistant glycolipids. J Biol Chem. 1990 Apr 15;265(11):6174–6181. [PubMed] [Google Scholar]
  43. Meiss H. K., Green R. F., Rodriguez-Boulan E. J. Lectin-resistant mutants of polarized epithelial cells. Mol Cell Biol. 1982 Oct;2(10):1287–1294. doi: 10.1128/mcb.2.10.1287. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Menon A. K., Mayor S., Ferguson M. A., Duszenko M., Cross G. A. Candidate glycophospholipid precursor for the glycosylphosphatidylinositol membrane anchor of Trypanosoma brucei variant surface glycoproteins. J Biol Chem. 1988 Feb 5;263(4):1970–1977. [PubMed] [Google Scholar]
  45. Menon A. K., Mayor S., Schwarz R. T. Biosynthesis of glycosyl-phosphatidylinositol lipids in Trypanosoma brucei: involvement of mannosyl-phosphoryldolichol as the mannose donor. EMBO J. 1990 Dec;9(13):4249–4258. doi: 10.1002/j.1460-2075.1990.tb07873.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Menon A. K., Schwarz R. T., Mayor S., Cross G. A. Cell-free synthesis of glycosyl-phosphatidylinositol precursors for the glycolipid membrane anchor of Trypanosoma brucei variant surface glycoproteins. Structural characterization of putative biosynthetic intermediates. J Biol Chem. 1990 Jun 5;265(16):9033–9042. [PubMed] [Google Scholar]
  47. Moran P., Raab H., Kohr W. J., Caras I. W. Glycophospholipid membrane anchor attachment. Molecular analysis of the cleavage/attachment site. J Biol Chem. 1991 Jan 15;266(2):1250–1257. [PubMed] [Google Scholar]
  48. Musil L. S., Baenziger J. U. Cleavage of membrane secretory component to soluble secretory component occurs on the cell surface of rat hepatocyte monolayers. J Cell Biol. 1987 Jun;104(6):1725–1733. doi: 10.1083/jcb.104.6.1725. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Pan Y. T., Elbein A. D. The effect of mannosamine on the formation of lipid-linked oligosaccharides and glycoproteins in canine kidney cells. Arch Biochem Biophys. 1985 Nov 1;242(2):447–456. doi: 10.1016/0003-9861(85)90229-2. [DOI] [PubMed] [Google Scholar]
  50. Pan Y. T., Elbein A. D. The formation of lipid-linked oligosaccharides in Madin-Darby canine kidney cells. Changes in oligosaccharide profiles induced by glucosamine. J Biol Chem. 1982 Mar 25;257(6):2795–2801. [PubMed] [Google Scholar]
  51. Powell S. K., Lisanti M. P., Rodriguez-Boulan E. J. Thy-1 expresses two signals for apical localization in epithelial cells. Am J Physiol. 1991 Apr;260(4 Pt 1):C715–C720. doi: 10.1152/ajpcell.1991.260.4.C715. [DOI] [PubMed] [Google Scholar]
  52. Reggio H., Webster P., Louvard D. Use of immunocytochemical techniques in studying the biogenesis of cell surfaces in polarized epithelia. Methods Enzymol. 1983;98:379–395. doi: 10.1016/0076-6879(83)98166-1. [DOI] [PubMed] [Google Scholar]
  53. Rodriguez-Boulan E., Nelson W. J. Morphogenesis of the polarized epithelial cell phenotype. Science. 1989 Aug 18;245(4919):718–725. doi: 10.1126/science.2672330. [DOI] [PubMed] [Google Scholar]
  54. Rodriguez-Boulan E. Polarized assembly of enveloped viruses from cultured epithelial cells. Methods Enzymol. 1983;98:486–501. doi: 10.1016/0076-6879(83)98176-4. [DOI] [PubMed] [Google Scholar]
  55. Sargiacomo M., Lisanti M., Graeve L., Le Bivic A., Rodriguez-Boulan E. Integral and peripheral protein composition of the apical and basolateral membrane domains in MDCK cells. J Membr Biol. 1989 Mar;107(3):277–286. doi: 10.1007/BF01871942. [DOI] [PubMed] [Google Scholar]
  56. Schneider P., Ferguson M. A., McConville M. J., Mehlert A., Homans S. W., Bordier C. Structure of the glycosyl-phosphatidylinositol membrane anchor of the Leishmania major promastigote surface protease. J Biol Chem. 1990 Oct 5;265(28):16955–16964. [PubMed] [Google Scholar]
  57. Sharma C., Dalferes E. R., Jr, Radhakrishnamurthy B., DePaolo C. J., Berenson G. S. Hepatic glycoprotein synthesis in streptozotocin diabetic rats. Biochem Int. 1987 Aug;15(2):395–401. [PubMed] [Google Scholar]
  58. Skillen A. W., Hawthorne G. C., Turner G. A. Serum alkaline phosphatase in rats with streptozotocin-induced diabetes. Horm Metab Res. 1987 Oct;19(10):505–506. doi: 10.1055/s-2007-1011865. [DOI] [PubMed] [Google Scholar]
  59. Stroynowski I., Soloski M., Low M. G., Hood L. A single gene encodes soluble and membrane-bound forms of the major histocompatibility Qa-2 antigen: anchoring of the product by a phospholipid tail. Cell. 1987 Aug 28;50(5):759–768. doi: 10.1016/0092-8674(87)90334-5. [DOI] [PubMed] [Google Scholar]
  60. Takami N., Ogata S., Oda K., Misumi Y., Ikehara Y. Biosynthesis of placental alkaline phosphatase and its post-translational modification by glycophospholipid for membrane-anchoring. J Biol Chem. 1988 Feb 25;263(6):3016–3021. [PubMed] [Google Scholar]
  61. Tanaka K., Nakamura T., Ichihara A. A unique trypsin-like protease associated with plasma membranes of rat liver. J Biol Chem. 1986 Feb 25;261(6):2610–2615. [PubMed] [Google Scholar]
  62. Thomas J. R., Dwek R. A., Rademacher T. W. Structure, biosynthesis, and function of glycosylphosphatidylinositols. Biochemistry. 1990 Jun 12;29(23):5413–5422. doi: 10.1021/bi00475a001. [DOI] [PubMed] [Google Scholar]
  63. Urban J., Parczyk K., Leutz A., Kayne M., Kondor-Koch C. Constitutive apical secretion of an 80-kD sulfated glycoprotein complex in the polarized epithelial Madin-Darby canine kidney cell line. J Cell Biol. 1987 Dec;105(6 Pt 1):2735–2743. doi: 10.1083/jcb.105.6.2735. [DOI] [PMC free article] [PubMed] [Google Scholar]
  64. Wilson J. M., Fasel N., Kraehenbuhl J. P. Polarity of endogenous and exogenous glycosyl-phosphatidylinositol-anchored membrane proteins in Madin-Darby canine kidney cells. J Cell Sci. 1990 May;96(Pt 1):143–149. doi: 10.1242/jcs.96.1.143. [DOI] [PubMed] [Google Scholar]

Articles from The EMBO Journal are provided here courtesy of Nature Publishing Group

RESOURCES