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. 1996 Mar 15;97(6):1478–1487. doi: 10.1172/JCI118570

Mannose corrects altered N-glycosylation in carbohydrate-deficient glycoprotein syndrome fibroblasts.

K Panneerselvam 1, H H Freeze 1
PMCID: PMC507208  PMID: 8617881

Abstract

Type I carbohydrate-deficient glycoprotein syndrome (CDGS) patients fail to add entire N-linked oligosaccharide chains to some serum glycoproteins. Here we show that four CDGS fibroblast cell lines have two related glycosylation abnormalities. First, they incorporate 3-10-fold less [3H] mannose into proteins, and, second, the size of the lipid-linked oligosaccharide precursor (LLO) is much smaller than in controls. Addition of exogenous mannose, but not glucose, to these CDGS cells corrects both the lowered [3H] mannose incorporation and the size of LLO. These corrections are not permanent, and the defects immediately reappear when mannose is removed. To explore further the basis of mannose correction, we analyzed the amount of 3H-labeled LLO intermediates. Except for dolichol-P-mannose, other precursors, including mannose, mannose-6-phosphate, mannose-1-phosphate, and GDP-mannose, all showed a 3-10-fold decrease in CDGS cells. Thus, there are no obvious lesions in the intracellular conversion of mannose into LLO, and, once inside the cell, [3H]mannose appeared to be metabolized normally. Initial velocities of [3H]mannose uptake were two- to threefold less in CDGS cells compared with controls, and this slower transport may partially explain the reduced [3H]mannose incorporation in CDGS cells. Since we previously showed that the enzymes converting glucose to mannose-6-phosphate appear to be normal, our results suggest that cells may acquire or generate mannose in other ways. Although we have not identified the primary defect in CDGS, these studies show that intracellular mannose is limited and that some patients might benefit from including mannose in their regular diets.

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Selected References

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  1. Anumula K. R., Spiro R. G. Release of glucose-containing polymannose oligosaccharides during glycoprotein biosynthesis. Studies with thyroid microsomal enzymes and slices. J Biol Chem. 1983 Dec 25;258(24):15274–15282. [PubMed] [Google Scholar]
  2. Asano T., Katagiri H., Takata K., Lin J. L., Ishihara H., Inukai K., Tsukuda K., Kikuchi M., Hirano H., Yazaki Y. The role of N-glycosylation of GLUT1 for glucose transport activity. J Biol Chem. 1991 Dec 25;266(36):24632–24636. [PubMed] [Google Scholar]
  3. Asano T., Shibasaki Y., Ohno S., Taira H., Lin J. L., Kasuga M., Kanazawa Y., Akanuma Y., Takaku F., Oka Y. Rabbit brain glucose transporter responds to insulin when expressed in insulin-sensitive Chinese hamster ovary cells. J Biol Chem. 1989 Feb 25;264(6):3416–3420. [PubMed] [Google Scholar]
  4. Baumann H., Jahreis G. P. Glucose starvation leads in rat hepatoma cells to partially N-glycosylated glycoproteins including alpha 1-acid glycoproteins. Identification by endoglycolytic digestions in polyacrylamide gels. J Biol Chem. 1983 Mar 25;258(6):3942–3949. [PubMed] [Google Scholar]
  5. Cacan R., Cecchelli R., Verbert A. Catabolic pathway of oligosaccharide-diphospho-dolichol. Study of the fate of the oligosaccharidic moiety in mouse splenocytes. Eur J Biochem. 1987 Jul 15;166(2):469–474. doi: 10.1111/j.1432-1033.1987.tb13539.x. [DOI] [PubMed] [Google Scholar]
  6. Cacan R., Lepers A., Belard M., Verbert A. Catabolic pathway of oligosaccharide-diphospho-dolichol. Subcellular sites of the degradation of the oligomannoside moiety. Eur J Biochem. 1989 Oct 20;185(1):173–179. doi: 10.1111/j.1432-1033.1989.tb15099.x. [DOI] [PubMed] [Google Scholar]
  7. Cacan R., Villers C., Bélard M., Kaiden A., Krag S. S., Verbert A. Different fates of the oligosaccharide moieties of lipid intermediates. Glycobiology. 1992 Apr;2(2):127–136. doi: 10.1093/glycob/2.2.127. [DOI] [PubMed] [Google Scholar]
  8. Carruthers A. Facilitated diffusion of glucose. Physiol Rev. 1990 Oct;70(4):1135–1176. doi: 10.1152/physrev.1990.70.4.1135. [DOI] [PubMed] [Google Scholar]
  9. Chapman A., Fujimoto K., Kornfeld S. The primary glycosylation defect in class E Thy-1-negative mutant mouse lymphoma cells is an inability to synthesize dolichol-P-mannose. J Biol Chem. 1980 May 25;255(10):4441–4446. [PubMed] [Google Scholar]
  10. Daniel P. F., Winchester B., Warren C. D. Mammalian alpha-mannosidases--multiple forms but a common purpose? Glycobiology. 1994 Oct;4(5):551–566. doi: 10.1093/glycob/4.5.551. [DOI] [PubMed] [Google Scholar]
  11. Datema R., Schwarz R. T. Effect of energy depletion on the glycosylation of a viral glycoprotein. J Biol Chem. 1981 Nov 10;256(21):11191–11198. [PubMed] [Google Scholar]
  12. Davidson S. K., Hunt L. A. Sindbis virus glycoproteins are abnormally glycosylated in Chinese hamster ovary cells deprived of glucose. J Gen Virol. 1985 Jul;66(Pt 7):1457–1468. doi: 10.1099/0022-1317-66-7-1457. [DOI] [PubMed] [Google Scholar]
  13. Freeze H. H., Willies L., Hamilton S., Koza-Taylor P. Two mutants of Dictyostelium discoideum that lack a sulfated carbohydrate antigenic determinant synthesize a truncated lipid-linked precursor of N-linked oligosaccharides. J Biol Chem. 1989 Apr 5;264(10):5653–5659. [PubMed] [Google Scholar]
  14. Freeze H. H., Yeh R., Miller A. L., Kornfeld S. Structural analysis of the asparagine-linked oligosaccharides from three lysosomal enzymes of Dictyostelium discoideum. Evidence for an unusual acid-stable phosphodiester. J Biol Chem. 1983 Dec 25;258(24):14874–14879. [PubMed] [Google Scholar]
  15. Gershman H., Robbins P. W. Transitory effects of glucose starvation on the synthesis of dolichol-linked oligosaccharides in mammalian cells. J Biol Chem. 1981 Aug 10;256(15):7774–7780. [PubMed] [Google Scholar]
  16. Goochee C. F. Bioprocess factors affecting glycoprotein oligosaccharide structure. Dev Biol Stand. 1992;76:95–104. [PubMed] [Google Scholar]
  17. Goochee C. F., Monica T. Environmental effects on protein glycosylation. Biotechnology (N Y) 1990 May;8(5):421–427. doi: 10.1038/nbt0590-421. [DOI] [PubMed] [Google Scholar]
  18. Gould G. W., Thomas H. M., Jess T. J., Bell G. I. Expression of human glucose transporters in Xenopus oocytes: kinetic characterization and substrate specificities of the erythrocyte, liver, and brain isoforms. Biochemistry. 1991 May 28;30(21):5139–5145. doi: 10.1021/bi00235a004. [DOI] [PubMed] [Google Scholar]
  19. Jaeken J., Carchon H., Stibler H. The carbohydrate-deficient glycoprotein syndromes: pre-Golgi and Golgi disorders? Glycobiology. 1993 Oct;3(5):423–428. doi: 10.1093/glycob/3.5.423. [DOI] [PubMed] [Google Scholar]
  20. Jaeken J., Eggermont E., Stibler H. An apparent homozygous X-linked disorder with carbohydrate-deficient serum glycoproteins. Lancet. 1987 Dec 12;2(8572):1398–1398. doi: 10.1016/s0140-6736(87)91287-6. [DOI] [PubMed] [Google Scholar]
  21. Jaeken J., Schachter H., Carchon H., De Cock P., Coddeville B., Spik G. Carbohydrate deficient glycoprotein syndrome type II: a deficiency in Golgi localised N-acetyl-glucosaminyltransferase II. Arch Dis Child. 1994 Aug;71(2):123–127. doi: 10.1136/adc.71.2.123. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Jaeken J., Stibler H., Hagberg B. The carbohydrate-deficient glycoprotein syndrome. A new inherited multisystemic disease with severe nervous system involvement. Acta Paediatr Scand Suppl. 1991;375:1–71. [PubMed] [Google Scholar]
  23. Jaeken J., van Eijk H. G., van der Heul C., Corbeel L., Eeckels R., Eggermont E. Sialic acid-deficient serum and cerebrospinal fluid transferrin in a newly recognized genetic syndrome. Clin Chim Acta. 1984 Dec 29;144(2-3):245–247. doi: 10.1016/0009-8981(84)90059-7. [DOI] [PubMed] [Google Scholar]
  24. Kmiécik D., Herman V., Stroop C. J., Michalski J. C., Mir A. M., Labiau O., Verbert A., Cacan R. Catabolism of glycan moieties of lipid intermediates leads to a single Man5GlcNAc oligosaccharide isomer: a study with permeabilized CHO cells. Glycobiology. 1995 Jul;5(5):483–494. doi: 10.1093/glycob/5.5.483. [DOI] [PubMed] [Google Scholar]
  25. Knauer R., Lehle L., Hanefeld F., von Figura K. Normal N-oligosaccharyltransferase activity in fibroblasts from patients with carbohydrate-deficient glycoprotein syndrome. J Inherit Metab Dis. 1994;17(5):541–544. doi: 10.1007/BF00711588. [DOI] [PubMed] [Google Scholar]
  26. Kornfeld R., Kornfeld S. Assembly of asparagine-linked oligosaccharides. Annu Rev Biochem. 1985;54:631–664. doi: 10.1146/annurev.bi.54.070185.003215. [DOI] [PubMed] [Google Scholar]
  27. Krasnewich D. M., Holt G. D., Brantly M., Skovby F., Redwine J., Gahl W. A. Abnormal synthesis of dolichol-linked oligosaccharides in carbohydrate-deficient glycoprotein syndrome. Glycobiology. 1995 Jul;5(5):503–510. doi: 10.1093/glycob/5.5.503. [DOI] [PubMed] [Google Scholar]
  28. Martinsson T., Bjursell C., Stibler H., Kristiansson B., Skovby F., Jaeken J., Blennow G., Strömme P., Hanefeld F., Wahlström J. Linkage of a locus for carbohydrate-deficient glycoprotein syndrome type I (CDG1) to chromosome 16p, and linkage disequilibrium to microsatellite marker D16S406. Hum Mol Genet. 1994 Nov;3(11):2037–2042. [PubMed] [Google Scholar]
  29. Mayor P., Maianu L., Garvey W. T. Glucose and insulin chronically regulate insulin action via different mechanisms in BC3H1 myocytes. Effects on glucose transporter gene expression. Diabetes. 1992 Mar;41(3):274–285. doi: 10.2337/diab.41.3.274. [DOI] [PubMed] [Google Scholar]
  30. Miyamoto K., Hase K., Takagi T., Fujii T., Taketani Y., Minami H., Oka T., Nakabou Y. Differential responses of intestinal glucose transporter mRNA transcripts to levels of dietary sugars. Biochem J. 1993 Oct 1;295(Pt 1):211–215. doi: 10.1042/bj2950211. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Panneerselvam K., Freeze H. H. Enzymes involved in the synthesis of mannose-6-phosphate from glucose are normal in carbohydrate deficient glycoprotein syndrome fibroblasts. Biochem Biophys Res Commun. 1995 Mar 17;208(2):517–522. doi: 10.1006/bbrc.1995.1369. [DOI] [PubMed] [Google Scholar]
  32. Payton M. A., Rheinnecker M., Klig L. S., DeTiani M., Bowden E. A novel Saccharomyces cerevisiae secretory mutant possesses a thermolabile phosphomannose isomerase. J Bacteriol. 1991 Mar;173(6):2006–2010. doi: 10.1128/jb.173.6.2006-2010.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Powell L. D., Paneerselvam K., Vij R., Diaz S., Manzi A., Buist N., Freeze H., Varki A. Carbohydrate-deficient glycoprotein syndrome: not an N-linked oligosaccharide processing defect, but an abnormality in lipid-linked oligosaccharide biosynthesis? J Clin Invest. 1994 Nov;94(5):1901–1909. doi: 10.1172/JCI117540. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Rearick J. I., Chapman A., Kornfeld S. Glucose starvation alters lipid-linked oligosaccharide biosynthesis in Chinese hamster ovary cells. J Biol Chem. 1981 Jun 25;256(12):6255–6261. [PubMed] [Google Scholar]
  35. Rosner M. R., Hubbard S. C., Ivatt R. J., Robbins P. W. N-asparagine-linked oligosaccharides: biosynthesis of the lipid-linked oligosaccharides. Methods Enzymol. 1982;83:399–408. doi: 10.1016/0076-6879(82)83037-1. [DOI] [PubMed] [Google Scholar]
  36. Roux L., Holojda S., Sundblad G., Freeze H. H., Varki A. Sulfated N-linked oligosaccharides in mammalian cells. I. Complex-type chains with sialic acids and O-sulfate esters. J Biol Chem. 1988 Jun 25;263(18):8879–8889. [PubMed] [Google Scholar]
  37. Sampath D., Varki A., Freeze H. H. The spectrum of incomplete N-linked oligosaccharides synthesized by endothelial cells in the presence of brefeldin A. J Biol Chem. 1992 Mar 5;267(7):4440–4455. [PubMed] [Google Scholar]
  38. Seeholzer S. H. Phosphoglucose isomerase: a ketol isomerase with aldol C2-epimerase activity. Proc Natl Acad Sci U S A. 1993 Feb 15;90(4):1237–1241. doi: 10.1073/pnas.90.4.1237. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Soyama K. Enzymatic determination of D-mannose in serum. Clin Chem. 1984 Feb;30(2):293–294. [PubMed] [Google Scholar]
  40. Spiro M. J., Spiro R. G. Potential regulation of N-glycosylation precursor through oligosaccharide-lipid hydrolase action and glucosyltransferase-glucosidase shuttle. J Biol Chem. 1991 Mar 15;266(8):5311–5317. [PubMed] [Google Scholar]
  41. Spiro R. G., Spiro M. J., Bhoyroo V. D. Studies on the regulation of the biosynthesis of glucose-containing oligosaccharide-lipids. Effect of energy deprivation. J Biol Chem. 1983 Aug 10;258(15):9469–9476. [PubMed] [Google Scholar]
  42. Stark N. J., Heath E. C. Glucose-dependent glycosylation of secretory glycoprotein in mouse myeloma cells. Arch Biochem Biophys. 1979 Feb;192(2):599–609. doi: 10.1016/0003-9861(79)90131-0. [DOI] [PubMed] [Google Scholar]
  43. Stoll J., Cacan R., Verbert A., Krag S. S. Lec15 cells transfer glucosylated oligosaccharides to protein. Arch Biochem Biophys. 1992 Dec;299(2):225–231. doi: 10.1016/0003-9861(92)90268-2. [DOI] [PubMed] [Google Scholar]
  44. Tachibana H., Taniguchi K., Ushio Y., Teruya K., Osada K., Murakami H. Changes of monosaccharide availability of human hybridoma lead to alteration of biological properties of human monoclonal antibody. Cytotechnology. 1994;16(3):151–157. doi: 10.1007/BF00749902. [DOI] [PubMed] [Google Scholar]
  45. Turco S. J. Modification of oligosaccharide-lipid synthesis and protein glycosylation in glucose-deprived cells. Arch Biochem Biophys. 1980 Dec;205(2):330–339. doi: 10.1016/0003-9861(80)90115-0. [DOI] [PubMed] [Google Scholar]
  46. Turco S. J., Pickard J. L. Altered G-protein glycosylation in vesicular stomatitis virus-infected glucose-deprived baby hamster kidney cells. J Biol Chem. 1982 Aug 10;257(15):8674–8679. [PubMed] [Google Scholar]
  47. Varki A. Metabolic radiolabeling of glycoconjugates. Methods Enzymol. 1994;230:16–32. doi: 10.1016/0076-6879(94)30004-6. [DOI] [PubMed] [Google Scholar]
  48. Varki A. Radioactive tracer techniques in the sequencing of glycoprotein oligosaccharides. FASEB J. 1991 Feb;5(2):226–235. doi: 10.1096/fasebj.5.2.2004668. [DOI] [PubMed] [Google Scholar]
  49. Villers C., Cacan R., Mir A. M., Labiau O., Verbert A. Release of oligomannoside-type glycans as a marker of the degradation of newly synthesized glycoproteins. Biochem J. 1994 Feb 15;298(Pt 1):135–142. doi: 10.1042/bj2980135. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. WOOD F. C., Jr, CAHILL G. F., Jr MANNOSE UTILIZATION IN MAN. J Clin Invest. 1963 Aug;42:1300–1312. doi: 10.1172/JCI104814. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Wada Y., Gu J., Okamoto N., Inui K. Diagnosis of carbohydrate-deficient glycoprotein syndrome by matrix-assisted laser desorption time-of-flight mass spectrometry. Biol Mass Spectrom. 1994 Feb;23(2):108–109. doi: 10.1002/bms.1200230211. [DOI] [PubMed] [Google Scholar]
  52. Wada Y., Nishikawa A., Okamoto N., Inui K., Tsukamoto H., Okada S., Taniguchi N. Structure of serum transferrin in carbohydrate-deficient glycoprotein syndrome. Biochem Biophys Res Commun. 1992 Dec 15;189(2):832–836. doi: 10.1016/0006-291x(92)92278-6. [DOI] [PubMed] [Google Scholar]
  53. Yamashita K., Ideo H., Ohkura T., Fukushima K., Yuasa I., Ohno K., Takeshita K. Sugar chains of serum transferrin from patients with carbohydrate deficient glycoprotein syndrome. Evidence of asparagine-N-linked oligosaccharide transfer deficiency. J Biol Chem. 1993 Mar 15;268(8):5783–5789. [PubMed] [Google Scholar]
  54. Yamashita K., Ohkura T., Ideo H., Ohno K., Kanai M. Electrospray ionization-mass spectrometric analysis of serum transferrin isoforms in patients with carbohydrate-deficient glycoprotein syndrome. J Biochem. 1993 Dec;114(6):766–769. doi: 10.1093/oxfordjournals.jbchem.a124253. [DOI] [PubMed] [Google Scholar]
  55. Yasugi E., Nakasuji M., Dohi T., Oshima M. Major defect of carbohydrate-deficient-glycoprotein syndrome is not found in the synthesis of dolichyl phosphate or N-acetylglucosaminyl-pyrophosphoryl-dolichol. Biochem Biophys Res Commun. 1994 Apr 29;200(2):816–820. doi: 10.1006/bbrc.1994.1524. [DOI] [PubMed] [Google Scholar]

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