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Temporal changes in the carbohydrates expressed on BG01 human embryonic stem cells during differentiation as embryoid bodies

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Abstract

Cell surface carbohydrates present on BG01 human embryonic stem cells after 28 days of differentiation were examined using two classes of carbohydrate binding proteins: lectins and antibodies specific for carbohydrate epitopes. Specificity of lectin staining was verified using carbohydrate ligands to block lectin interaction, glycohydrolases to cleave specific sugar residues that are receptors for these proteins, and periodate oxidation to destroy susceptible sugar residues. Specific antibodies were used to identify various tissue types and germ layers present in the 12- and 28-day differentiating embryoid bodies. Results from 12 and 28-day differentiated embryoid bodies were compared to determine changes over time. A slight increase in the sialylation of α-GalNAc was seen between 12 and 28 days of differentiation due to the presence of sialyl Tn and/or other sialylated α-GalNAc residues. Increases were also observed in GalNAc, the T antigen (Gal β1,3 GalNAc), and difucosylated LacNAc residues during this time interval. Additionally, some distinct differences in the pattern of lectin staining between 12 and 28 days were observed. Not unexpectedly, the presence of most differentiated cell-types increased during this time period with the exception of neural progenitors, which decreased. Undifferentiated cells, which were prevalent in the 12-day EBs, were undetectable after 28 days. We conclude that several changes in glycosylation occurred during the differentiation of embryonic stem cells, and that these changes may play a role in embryonic development.

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Abbreviations

ABS:

acetate buffered saline

BSA:

bovine serum albumin

EB:

embryoid body

HESC:

human embryonic stem cell

PBS:

phosphate buffered saline

PFA:

paraformaldehyde

T antigen:

Thomsen–Friedenreich antigen, Gal β1,3 GalNAc α1—Ser/Thr

Tn antigen:

GalNAc α1—Ser/Thr

References

  1. Haltiwanger, R.S., Lowe, J.B.: Role of glycosylation in development. Annu. Rev. Biochem. 73, 491–537 (2004)

    Google Scholar 

  2. Wearne, K.A., Winter, H.C., O’Shea, K., Goldstein, I.J.: Use of lectins for probing differentiated embryonic stem cells for carbohydrates. Glycobiology 16, 981–990 (2006)

    Article  PubMed  CAS  Google Scholar 

  3. Venable, A., Mitalipova, M., Lyons, I., Jones, K., Shin, S., Pierce, M., Stice, S.: Lectin binding profiles of SSEA-4 enriched pluripotent human embryonic stem cell surfaces. BMC Dev. Biol. 5, 5–15 (2005)

    Article  Google Scholar 

  4. Goldstein, I.J., Poretz, R.D.: Isolation, physicochemical characterization, and carbohydrate-binding specificity of lectins. In: Liener, I.E., Sharon, N., Goldstein, I.J. (eds.) The Lectins: Properties, Functions and Applications in Biology and Medicine, pp. 35–247. Academic Press Inc., Orlando (1986)

    Google Scholar 

  5. Kirkeby, S., Winter, H.C., Goldstein, I.J.: Comparison of the binding properties of the mushroom Marasmius oreades lectin and Griffonia simplicifolia I-B4 isolectin to αgalactosyl carbohydrate antigens in the surface phase. Xenotransplantation 11, 254–261 (2004)

    Article  PubMed  Google Scholar 

  6. Winter, H.C., Mostafapour, K., Goldstein, I.J.: The mushroom Marasmius oreades lectin is a blood group type B agglutinin that recognizes the Galα1,3Gal and Galα1,3Galβ1,4GlcNAc porcine xenotransplantation epitopes with high affinity. J. Biol. Chem. 277, 14996–15001 (2002)

    Article  PubMed  CAS  Google Scholar 

  7. Singh, T., Wu, J.H., Peumans, W.J., Rougé, P., Van Damme, E.J.M., Wu, A.M.: Recognition profile of Morus nigra agglutinin (Morniga G) expressed by monomeric ligands, simple clusters and mammalian polyvalent glycotopes. Mol. Immunol. 44, 451–462 (2007)

    Article  PubMed  CAS  Google Scholar 

  8. Yamamoto, K., Konami, Y., Irimura, T.: Sialic acid-binding motif of Maackia amurensis lectins. J. Biochem. 121, 756–761 (1997)

    PubMed  CAS  Google Scholar 

  9. Endo, T., Ohbayashi, H., Kanazawa, K., Kochibe, N., Kobata, A.: Carbohydrate binding specificity of immobilized Psathyrella velutina lectin. J. Biol. Chem. 267, 707–713 (1992)

    PubMed  CAS  Google Scholar 

  10. Kochibe, N., Matta, K.L.: Purification and Properties of an N-acetylglucosamine-specific lectin from Psathyrella velutina mushroom. J. Biol. Chem. 264, 173–177 (1989)

    PubMed  CAS  Google Scholar 

  11. Ueda, H., Kojima, K., Saitoh, T., Ogawa, H.: Interaction of a lectin from Psathyrella velutina mushroom with N-acetylneuraminic acid. FEBS Lett. 448, 75–80 (1999)

    Article  PubMed  CAS  Google Scholar 

  12. Ueda, H., Matsumoto, H., Takahashi, N., Ogawa, H.: Psathyrella velutina mushroom lectin exhibits high affinity toward sialoglycoproteins possessing terminal N-acetylneuraminic acid α2,3-linked to penultimate galactose residues of trisialyl N-glycans. J. Biol. Chem. 277, 24916–24925 (2002)

    Article  PubMed  CAS  Google Scholar 

  13. Kawashima, H., Sueyoshi, S., Li, H., Yamamoto, K., Osawa, T.: Carbohydrate binding specificities of several poly-N-acetyllactosamine-binding lectins. Glycoconj. J. 7, 323–334 (1990)

    Article  PubMed  CAS  Google Scholar 

  14. Crowley, J.F., Goldstein, I.J., Arnarp, J., Löngren, J.: Carbohydrate binding studies on the lectin from Datura stramonium seeds. Arch. Biochem. Biophys. 231, 524–533 (1984)

    Article  PubMed  CAS  Google Scholar 

  15. Kawagish, H., Mizuno, T.: Purification and properties of a β-galactosyl-specific lectin from the fruiting bodies of Ischnoderma resinosus. FEBS Lett. 227, 99–102 (1988)

    Article  Google Scholar 

  16. Teneberg, S., Ångström, J., Jovall, P-Å., Karlsson, K-A.: Characterization of binding of Galβ4GlcNAc-specific lectins from Erythrina cristagalli and Erythrina corallodendron to glycosphingolipids. J. Biol. Chem. 269, 8554–8563 (1994)

    PubMed  CAS  Google Scholar 

  17. Fukumori, F., Takeuchi, N., Hagiwara, T., Ohbayashi, H., Endo, T., Kochibe, N., Nagata, Y., Kobata, A.: Primary structure of a Fucose-specific lectin obtained from a mushroom, Aleuria aurantia. J. Biochem. 107, 190–196 (1990)

    PubMed  CAS  Google Scholar 

  18. Kochibe, N., Furukawa, K., Purification and properties of a novel fucose-specific hemagglutinin of Aleuria aurantia. Biochemistry 19, 2841–2846 (1980).

    Article  PubMed  CAS  Google Scholar 

  19. Wimmerova, M., Mitchell, E., Sanchez, J.-F., Gautier, C., Imberty, A.: Crystal structure of a fungal lectin, J. Biol. Chem. 278, 27059–27067 (2003)

    Article  PubMed  CAS  Google Scholar 

  20. Yamashita, K., Kochibe, N., Ohkura, T., Ueda, I., Kobata, A.: Fractionation of l-Fucose-containing oligosaccharides on immobilized Aleuria aurantia lectin. J. Biol. Chem. 260, 4688–4693 (1985)

    PubMed  CAS  Google Scholar 

  21. Moreno, F.B.M.B., Martil, D.E., Cavada, B.S., Filguiera de Azevedo, Jr. W.: Crystallization and preliminary X-ray diffraction analysis of an anti-H(O) lectin from Lotus tetragonolobus seeds. Acta Crystallograph. Sect. F. Struct. Biol. Cryst. Commun. 62, 680–683 (2006)

    Article  PubMed  Google Scholar 

  22. Petryniak, J., Duś, D., Podwińska, J.: Agglutination of murine and guinea pig peritoneal cells by α-l-fucose-binding lectin: Evonymus europaea. Eur. J. Immunol. 13, 459–464 (1983)

    Article  PubMed  CAS  Google Scholar 

  23. Petryniak, J., Goldstein, I.J.: Immunochemical studies on the interaction between synthetic glycoconjugates and α-l-fucosyl binding lectins. Biochemistry 25, 2829–2838 (1986)

    Article  PubMed  CAS  Google Scholar 

  24. Pereira, M.E.A., Kabat, E.A.: Specificity of Purified Hemagglutinin (Lectin) from Lotus tetragonolobus. Biochemistry 13, 3184–3192 (1974)

    Article  PubMed  CAS  Google Scholar 

  25. Iyer, P.N.S., Wilkinson, K.D., Goldstein, I.J.: An N-acetyl-d-glucosamine binding lectin from Bandeiraea simplicifolia seeds. Arch. Biochem. Biophys. 177, 330–333 (1976)

    Article  CAS  Google Scholar 

  26. Nakamura-Tsuruta, S., Kominami, J., Kuno, A., Hirabayashi, J.: Evidence that Agaricus bisporus agglutinin (ABA) has dual sugar-binding specificity. Biochem. Biophys. Res. Commun. 347, 215–220 (2006)

    Article  PubMed  CAS  Google Scholar 

  27. Wu, A.M., Wu, J.H., Herp, A., Liu, J.-H.: Effect of polyvalencies of glycotopes on the binding of a lectin from the edible mushroom, Agaricus bisporus. Biochem. J. 371, 311–320 (2003)

    Article  PubMed  CAS  Google Scholar 

  28. Boland, C.R., Chen, Y.-F., Rinderle, S.J., Resau, J.H., Luk, G.D., Lynch, H.T., Goldstein, I.J.: Use of the lectin from Amaranthus caudatus as a histochemical probe of proliferating colonic epithelial cells. Cancer Res. 51, 657–665 (1991)

    PubMed  CAS  Google Scholar 

  29. Rinderle, S.J., Goldstein, I.J., Matta, K.L., Ratcliffe, R.M.: Isolation and characterization of amaranthin, a lectin present in the seeds of Amaranthus caudatus, that recognizes the T- (or cryptic T)-antigen. J. Biol. Chem. 264, 16123–16131 (1989)

    PubMed  CAS  Google Scholar 

  30. Grahn, E., Askarieh, G., Rocklöv, Å.H, Tateno, H., Winter, H.C., Goldstein, I.J., Krengel, U.: Crystal structures of the Marasmius oreades mushroom lectin in complex with a xenotransplantation epitope. J. Mol. Biol. 369, 710–721 (2007)

    Article  PubMed  CAS  Google Scholar 

  31. Shibuya, N., Goldstein, I.J., Van Damme, E.J.M., Peumans, W.J.: Binding properties of a mannose-specific lectin from the snowdrop (Galanthus nivalis) bulb. J. Biol. Chem. 263, 728–734 (1988)

    PubMed  CAS  Google Scholar 

  32. Mo, H., Winter, H.C., Goldstein, I.J.: Purification and characterization of a Neu5Acα2-6Galβ1,4Glc/GlcNAc-specific lectin from the fruiting body of the polypore mushroom Polyporus squamosus. J. Biol. Chem. 275, 10623–10629 (2000)

    Article  PubMed  CAS  Google Scholar 

  33. Shibuya, N., Goldstein, I.J., Broekaert, W.F., Nsimba-Lubaki, M., Peeters, B., Peumans, W.J.: The elderberry (Sambucus nigra L.) bark lectin recognizes the Neu5Ac(α2-6)Gal/GalNAc sequence. J. Biol. Chem. 262, 1596–1601 (1987)

    PubMed  CAS  Google Scholar 

  34. Mo, H., Winter, H.C., Van Damme, E.J.M., Peumans, W.J., Misaki, A., Goldstein, I.J.: Carbohydrate binding properties of banana (Musa acuminata) lectin. Eur. J. Biochem. 268, 2609–2615 (2001)

    Article  PubMed  CAS  Google Scholar 

  35. Hammarström, S., Murphy, L.A., Goldstein, I.J., Etzler, M.E., Carbohydrate binding specificity of four N-acetyl-d-galactosamine-“specific” lectins: Helix pomatia A hemagglutinin, soy bean agglutinin, lima bean lectin, and Dolichos biflorus lectin. Biochemistry 16, 2750–2755 (1977)

    Article  PubMed  Google Scholar 

  36. Singh, T., Wu, J.H., Peumans, W.J., Rougé, P., Van Damme, E.J.M., Avarez, R.A., Blixt, O.: Carbohydrate specificity of an insecticidal lectin isolated from the leaves of Glechoma hederacea (ground ivy) towards mammalian glycoconjugates. Biochem. J. 393, 331–341 (2006)

    Article  PubMed  CAS  Google Scholar 

  37. Ishiyama, I., Uhlenbruck, G., Hermann, G.: Isolation of an anti-A-agglutinin from Helix aspersa. Blut 24, 178–179 (1972)

    Article  PubMed  CAS  Google Scholar 

  38. Grubhoffer, L., Ticha, M., Kocourek, J.: Isolation and properties of a lectin from the seeds of hairy vetch (Vicia villosa Roth). Biochem. J. 195, 623–626 (1981)

    PubMed  CAS  Google Scholar 

  39. Knibbs, R.N., Osborne, S.E., Glick, G.D., Goldstein, I.J.: Binding determinants of the sialic acid-specific lectin from the slug Limax flavus. J. Biol. Chem. 268, 18524–18531 (1993)

    PubMed  CAS  Google Scholar 

  40. Muresan, V., Iwanij, V., Smith, Z.D.J., Jamieson, J.D.: Purification and use of limulin: a sialic acid-specific lectin. J. Histochem. Cytochem. 30, 938–946 (1982)

    PubMed  CAS  Google Scholar 

  41. Ciopraga, J., Ångström, J., Bergström, J., Larsson, T., Karlsson, N., Motas, C., Gozia, O., Teneberg, S.: Isolectins from Solanum tuberosum with different detailed carbohydrate binding specificities: unexpected recognition of lactosylceramide by N-acetyllactosamine-binding lectins. J. Biochem. 128, 855–867 (2000)

    PubMed  CAS  Google Scholar 

  42. Matsumoto, I., Jimbo, A., Mizuno, Y., Seno, N., Jeanloz, R.W.: Purification and characterization of potato lectin. J. Biol. Chem. 258, 2886–2891 (1983)

    PubMed  CAS  Google Scholar 

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Acknowledgements

We thank Sue O’Shea for helpful advice and critical reading of this manuscript and Crystal Pacut for growing the BG01 hESCs and supplying us with cryostat sections. We also thank the Consortium for Functional Glycomics for the use of their glycan array on our fucose-binding lectins. Finally, we thank the NIH for funding this project with grants GM-29470 and GM-069985 (This research is supported (in part) by the National Institutes of Health through the University of Michigan’s Cancer Center Support Grant (5 P30 CA46592)).

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Authors and Affiliations

  1. Department of Biological Chemistry, University of Michigan, 1150 West Medical Center Drive, 3220E MSRBIII, Ann Arbor, MI, 48109, USA

    Kimberly A. Wearne, Harry C. Winter & Irwin J. Goldstein

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  1. Kimberly A. Wearne
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  2. Harry C. Winter
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  3. Irwin J. Goldstein
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Correspondence to Irwin J. Goldstein.

Additional information

Lectin abbreviations can be found in Table 1.

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Wearne, K.A., Winter, H.C. & Goldstein, I.J. Temporal changes in the carbohydrates expressed on BG01 human embryonic stem cells during differentiation as embryoid bodies. Glycoconj J 25, 121–136 (2008). https://doi.org/10.1007/s10719-007-9064-x

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