Springer Nature is making SARS-CoV-2 and COVID-19 research free. View research | View latest news | Sign up for updates

The lectin-binding pattern of nucleolin and its interaction with endogenous galectin-3

Abstract

Unlike nuclear nucleolin, surface-expressed and cytoplasmic nucleolin exhibit Tn antigen. Here, we show localization-dependent differences in the glycosylation and proteolysis patterns of nucleolin. Our results provide evidence for different paths of nucleolin proteolysis in the nucleus, in the cytoplasm, and on the cell surface. We found that full-length nucleolin and some proteolytic fragments coexist within live cells and are not solely the result of the preparation procedure. Extranuclear nucleolin undergoes N- and O-glycosylation, and unlike cytoplasmic nucleolin, membrane-associated nucleolin is not fucosylated. Here, we show for the first time that nucleolin and endogenous galectin-3 exist in the same complexes in the nucleolus, the cytoplasm, and on the cell surface of melanoma cells. Assessments of the interaction of nucleolin with galectin-3 revealed nucleolar co-localization in interphase, suggesting that galectin-3 may be involved in DNA organization and ribosome biogenesis.

Abbreviations

AAA:

Aleuria aurantia agglutinin

DSA:

Datura stramonium gglutinin

GNA:

Galanthus nivalis agglutinin

H antigen:

Fucα1-2Galβ1-4GlcNAcβ1-3Gal-

LEA:

Lycopersicon esculentum agglutinin

Lex:

Galβ1-4(Fucα1-3)GlcNAcβ-

LTA:

Tetragonolobus purpureus agglutinin

MAA-II:

Maackia amurensis agglutinin

SNA-I:

Sambucus nigra agglutinin

PHA-E:

Phaseolus vulgaris erythroagglutinin

PHAL:

Phaseolus vulgaris leucoagglutinin

PNA:

peanut agglutinin

T antigen:

Galβ1-3GalNAcα1 -Ser/Thr

Tn antigen:

GalNAcα1 -Ser/Thr

UEA-I:

Ulex europaeus agglutinin

VVA:

Vicia villosa agglutinin

References

  1. 1.

    Mehes, G. and Pajor, L. Nucleolin and fibrillarin expression in stimulated lymphocytes and differentiating HL-60 cells. A flow cytometric assay. Cell Prolif. 28 (1995) 329–336.

  2. 2.

    Sirri, V., Roussel, P., Gendron, M.C. and Hernandez-Verdun, D. Amount of the two major Ag-NOR proteins, nucleolin, and protein B23 is cell-cycle dependent. Cytometry 28 (1997) 147–156.

  3. 3.

    Gorczyca, W., Smolewski, P., Grabarek, J., Ardelt, B., Ita, M., Melamed, M.R. and Darzynkiewicz, Z. Morphometry of nucleoli and expression of nucleolin analyzed by laser scanning cytometry in mitogenically stimulated lymphocytes. Cytometry 45 (2001) 206–213.

  4. 4.

    Mongelard, F. and Bouvet, P. Nucleolin: a multiFACeTed protein. Trends Cell Biol. 17 (2007) 80–86.

  5. 5.

    Ginisty, H., Sicard, H., Roger, B. and Bouvet, P. Structure and functions of nucleolin. J. Cell Sci. 112 (1999) 761–772.

  6. 6.

    Bouvet, P., Diaz, J.-J., Kindbeiter, K., Madjar, J.-J. and Amalric, F. Nucleolin interacts with several ribosomal proteins through its RGG domain. J. Biol. Chem. 273 (1998) 19025–19029.

  7. 7.

    Ghisolfis, L., Amalric, G.J.F. and Erard, M. The glycine-rich domain of nucleolin has an unusual super secondary structure responsible for its RNAhelix-destabilizing properties. J. Biol. Chem. 267 (1992) 2955–2959.

  8. 8.

    Ginisty, H., Amalric, F. and Bouvet, P. Nucleolin functions in the first step of ribosomal RNA processing. EMBO J. 17 (1998) 1476–1486.

  9. 9.

    Belenguer, P., Baldin, W., Mathieu, C., Prats, H., Bensaid, M., Bouche, G. and Amalric, F. Protein kinase NII and the regulation of rDNA transcription in mammalian cells. Nucleic Acids Res. 17 (1989) 6625–6636.

  10. 10.

    Belenguer, P., Caizergues-Ferrer, M., Labbe, J.-C., Doree, M. and Amalric, F. Mitosis-specific phosphorylation of nucleolin by p34dc2 protein kinase. Mol. Cell. Biol. 10 (1990) 3607–3618.

  11. 11.

    Srivastava, M. and Pollard, H.B. Molecular dissection of nucleolin’s role in growth and cell proliferation: new insights. FASEB J. 13 (1999) 1911–1922.

  12. 12.

    Zhang, J., Tsaprailis, G. and Bowden, G.T. Nucleolin stabilizes Bcl-XL messenger RNA in response to UVA irradiation. Cancer Res. 68 (2008) 1046–1054.

  13. 13.

    Chen, Ch.-Y., Gherzi, R., Andersen, J.S., Gaietta, G., Jurchott, K., Royer, H.-D., Mann, M. and Karin, M. Nucleolin and YB-1 are required for JNKmediated interleukin-2 mRNA stabilization during T-cell activation. Genes Dev. 14 (2000) 1236–1248.

  14. 14.

    Jiang, Y., Xu, X.-S. and Russell, J.E. A nucleolin-binding 3′ untranslated region element stabilizes β-globin mRNA in vivo. Mol. Cell. Biol. 26 (2006) 2419–2429.

  15. 15.

    Lee, P.-T., Liao, P.-C., Chang, W.-C. and Tseng, J.T. Epidermal growth factor increases the interaction between nucleolin and heterogeneous nuclear ribonucleoprotein K/Poly(C) binding protein 1 complex to regulate the gastrin mRNA turnover. Mol. Biol. Cell 18 (2007) 5004–5013.

  16. 16.

    Otake, Y., Soundararajan, S., Sengupta, T.K., Kio, E.A., Smith, J.C., Pineda-Roman, M., Stuart, R.K., Spicer, E, K. and Fernandes, D.J. Overexpression of nucleolin in chronic lymphocytic leukemia cells induces stabilization of bcl2 mRNA. Blood 109 (2007) 3069–3075.

  17. 17.

    Rajagopalan, L.E., Westmark, C.J., Jarzembowski, J.A. and Malter, J.S. hnRNP C increases amyloid precursor protein (APP) production by stabilizing APP mRNA. Nucleic Acids Res. 26 (1998) 3418–3423.

  18. 18.

    Soundararajan, S., Chen, W., Spicer, E.K., Courtenay-Luck, N. and Fernandes, D.J. The Nucleolin targeting aptamer AS1411 destabilizes Bcl-2 messenger RNA in human breast cancer cells. Cancer Res. 68 (2008) 2358–2365.

  19. 19.

    Ishimaru, D., Zuraw, L., Ramalingam, S., Sengupta, T.K., Bandyopadhyay, S., Reuben, A., Fernandes, D.J. and Spicer, E.K. Mechanism of regulation of bcl-2 mRNA by nucleolin and A+U-rich element-binding factor 1 (AUF1). J. Biol. Chem. 285 (2010) 27182–27191.

  20. 20.

    Takagi, M., Absalon, M.J., McLure, K.G. and Kastan, M.B. Regulation of p53 translation and induction after DNA damage by ribosomal protein L26 and nucleolin. Cell 123 (2005) 49–63.

  21. 21.

    Said, E.A., Krust, B., Nisole, S., Svab, J., Briand, J.-P. and Hovanessian, A.G. The anti-HIV cytokine midkine binds the cell surface-expressed nucleolin as a low affinity receptor. J. Biol. Chem. 277 (2002) 37492–37502.

  22. 22.

    Sinclair, J.F. and O’Brien, A.D. Cell surface-localized nucleolin is a rukaryotic Receptor for the adhesin intimin-γ of enterohemorrhagic Escherichia coli O157:H7. J. Biol. Chem. 277 (2002) 2876–2885.

  23. 23.

    Chen, X., Kube, D.M., Cooper, M.J. and Davis, P.B. Cell surface nucleolin serves as receptor for DNA nanoparticles composed of pegylated polylysine and DNA. Mol. Ther. 16 (2008) 333–342.

  24. 24.

    Christian, S., Pilch, J., Akerman, M.E., Porkka, K., Laakkonen, P. and Ruoslahti, E. Nucleolin expressed at the cell surface is a marker of endothelial cells in angiogenic blood vessels. J. Cell Biol. 163 (2003) 871–878.

  25. 25.

    Harms, G., Kraft, R., Grelle, G., Volz, B., Dernedde, J. and Tauber R. Identification of nucleolin as a new L-selectin ligand. Biochem. J. 360 (2001) 531–538.

  26. 26.

    Joo, E.J., ten Dam, G.B., van Kuppevelt, T.H., Toida, T., Linhardt, R.J. and Kim, Y.S. Nucleolin: acharan sulfate-binding protein on the surface of cancer cells. Glycobiology 15 (2005) 1–9.

  27. 27.

    Legrand, D., Vigie, K., Said, E.A., Elass, E., Masson, M., Slomianny, M.-Ch., Carpentier, M., Briand, J.-P., Mazurier, J. and Hovanessian, A.G. Surface nucleolin participates in both the binding and endocytosis of lactoferrin in target cells. Eur. J. Biochem. 271 (2004) 303–317.

  28. 28.

    Hoja-Łukowicz, D., Przybyło, M., Pocheć, E., Drabik, A., Silberring, J., Kremser, M., Schadendorf, D., Laidler, P. and Lityńska, A. The new face of nucleolin in human melanoma. Cancer Immunol. Immunother. 58 (2009) 1471–1480.

  29. 29.

    Hovanessian, A.G., Puvion-Dutilleul, F., Nisole, S., Svab, J., Perret, E., Deng, J.-S. and Krust, B. The cell-surface-expressed nucleolin is associated with the actin cytoskeleton. Exp. Cell Res. 261 (2000) 312–328.

  30. 30.

    Kusakawa, T., Shimakami, T., Kaneko, S., Yoshioka, K. and Murakami, S. Functional interaction of hepatitis C virus NS5B with nucleolin GAR domain. J. Biochem. 141 (2007) 917–927.

  31. 31.

    Hovanessian, A.G. Midkine, a cytokine that inhibits HIV infection by binding to the cell surface expressed nucleolin. Cell Res. 16 (2006) 174–181.

  32. 32.

    Said, E.A., Courty, J., Svab, J., Delbe, J., Krust, B. and Hovanessian, A.G. Pleiotrophin inhibits HIV infection by binding the cellsurface-expressed nucleolin. FEBS J. 272 (2005) 4646–4659.

  33. 33.

    Reyes-Reyes, E.M. and Akiyama, S.K. Cell-surface nucleolin is a signal transducing P-selectin binding protein for human colon carcinoma cells. Exp. Cell Res. 314 (2008) 2212–2223.

  34. 34.

    Hirano, K., Miki, Y., Hirai, Y., Sato, R., Itoh, T., Hayashi, A., Yamanaka, M., Eda, S. and Beppu, M. A Multifunctional shuttling protein nucleolin is a macrophage receptor for apoptotic cells. J. Biol. Chem. 280 (2005) 39284–39293.

  35. 35.

    Shi, H., Huang, Y., Zhou, H., Song, X., Yuan, S., Fu, Y. and Luo, Y. Nucleolin is a receptor that mediates anti-angiogenic and antitumor activity of endostatin. Blood 110 (2007) 2899–2906.

  36. 36.

    Carpentier, M., Morelle, W., Coddeville, B., Pons, A., Masson, M., Mazurier, J. and Legrand, D. Nucleolin undergoes partial N- and O- glycosylations in the extranuclear cell compartment. Biochemistry 44 (2005) 5804–5815.

  37. 37.

    Aldi, S., Giovampaola, C.D., Focarelli, R., Armini, A., Ziche, M., Finetti, F. and Rosati, F. A fucose-containing O-glycoepitope on bovine and human nucleolin. Glycobiology 19 (2009) 337–343.

  38. 38.

    Hoja-Łukowicz, D., Lityńska, A., Pocheć, E., Przybyło, M., Kremser, E., Ciołczyk-Wierzbicka, D. and Laidler, P. Identification of PNA-positive proteins in the primary uveal melanoma cell line by mass spectrometry. Acta Biol. Cracov. Seria Zool. 47 (2006) 27–33.

  39. 39.

    Watanabe, T., Tsuge, H., Imagawa, T., Kise, D., Hirano, K., Beppu, M., Takahashi, A., Yamaguchi, K., Fujiki, H. and Suganuma M. Nucleolin as cell surface receptor for tumor necrosis factor-alpha inducing protein: a carcinogenic factor of Helicobacter pylori. J. Cancer Res. Clin. Oncol. 136 (2010) 911–921.

  40. 40.

    Görelik, E., Galili, U. and Raz, A. On the role of cell surface carbohydrates and their binding proteins (lectins) in tumor metastasis. Cancer Metastasis Rev. 20 (2001) 245–277.

  41. 41.

    Voss, P.G., Haudek, K.C., Patterson, R.J. and Wang, J.L. Inhibition of cellfree splicing by saccharides that bind galectins and SR proteins. J. Carbohydr. Chem. 31 (2012) 519–534.

  42. 42.

    Chen, Ch.-M., Chiang, S.-Y. and Yeh, N.-H. Increased stability of nucleolin in proliferating cells by inhibition of its self-cleaving activity. J. Biol. Chem. 266 (1991) 7754–7758.

  43. 43.

    Fang, S.H. and Yeh, N.H. The self-cleaving activity of nucleolin determines its molecular dynamics in relation to cell proliferation. Exp. Cell Res. 208 (1993) 48–53.

  44. 44.

    Lee, N., Wang, W.-Ch. and Fukuda, M. Granulocytic differentiation of HL-60 cells is associated with increase of poly-N-acetyllactosamine in Asn-linked oligosaccharides attached to human lysosomal membrane glycoproteins. J. Biol. Chem. 265 (1990) 20476–20487.

  45. 45.

    Yan, L., Wilkins, P.P., Alvarez-Manilla, G., Do, S.-I., Smith, D.F. and Cummings, R.D. Immobilized Lotus tetragonolobus agglutinin binds oligosaccharides containing the Lex determinant. Glycoconj. J. 14 (1997) 45–55.

  46. 46.

    Hoja-Łukowicz, D., Link-Lenczowski, P., Carpentieri, A., Amoresano, A., Pocheć, E., Artemenko, K.A., Bergquist, J. and Lityńska, A. L1CAM from human melanoma carries a novel type of N-glycan with Galβ1-4Galβ1-motif. Involvement of N-linked glycans in migratory and invasive behaviour of melanoma cells. Glycoconj. J. 30 (2013) 205–225.

  47. 47.

    Caizergues-Ferrer, M., Belenguer, P., Lapeyre, B., Amalric, F., Wallace, M.O. and Olson, M.O.J. Phosphorylation of nucleolin by a nucleolar type NII protein kinase. Biochemistry 26 (1987) 7876–7883.

  48. 48.

    Tediose, T., Kolev, M., Sivasankar, B., Brennan, P., Morgan, B.P. and Donev, R. Interplay between REST and nucleolin transcription factors: a key mechanism in the overexpression of genes upon increased phosphorylation. Nucleic Acids Res. 38 (2010) 2799–2812.

  49. 49.

    Garcia, M.C., Williams, J., Johnson, K., Olden, K. and Roberts, J.D. Arachidonic acid stimulates formation of a novel complex containing nucleolin and RhoA. FEBS Lett. 585 (2011) 618–622.

  50. 50.

    Warrener, P. and Petryshyn, R. Phosphorylation and proteolytic degradation of nucleolin from 3T3-F442A cells. Biochem. Biophys. Res. Commun. 180 (1991) 716–723.

  51. 51.

    Bourbon, H., Bugler, B., Caizergues-Ferrer, M. and Amalric, F. Role of phosphorylation on the maturation pathways of a 100 kDa nucleolar protein. FEBS Lett. 155 (1983) 218–222.

  52. 52.

    Semba, S., Mizuuchi, E. and Yokozaki, H. Requirement of phosphatase of regenerating liver-3 for the nucleolar localization of nucleolin during the progression of colorectal carcinoma. Cancer Sci. 1012 (2010) 254–226.

  53. 53.

    Losfeld, M.-E., Khoury, D.E., Mariot, P., Carpentier, M., Krust, B., Briand, J.-P., Mazurier, J., Hovanessian, A.G. and Legrand, D. The cell surface expressed nucleolin is a glycoprotein that triggers calcium entry into mammalian cells. Exp. Cell Res. 315 (2009) 357–369.

  54. 54.

    Losfeld, M.-E., Leroy, A., Coddeville, B., Carpentier, M., Mazurier, J. and Legrand, D. N-glycosylation influences the structure and self-association abilities of recombinant nucleolin. FEBS J. 278 (2011) 2552–2564.

  55. 55.

    Agrwal, N., Wang, J.L. and Voss, P.G. Carbohydrate-binding Protein 35. J. Biol. Chem. 264 (1989) 17236–17242.

  56. 56.

    Paces-Fessy, M., Boucher, D., Petit, E., Paute-Briand, S. and Blanchet-Tournier, M.-F. The negative regulator of Gli, Suppressor of fused (Sufu), interacts with SAP18, Galectin3 and other nuclear proteins. Biochem. J. 378 (2004) 353–362.

  57. 57.

    Haudek, K.C., Spronk, K.J., Voss, P.G., Patterson, R.J., Wang, J.L. and Arnoys, E.J. Dynamics of galectin-3 in the nucleus and cytoplasm. Biochim. Biophys. Acta 1800 (2010) 181–189.

  58. 58.

    Mehul, B., Bawumia, S. and Hughes, R.C. Cross-linking of galectin 3, a galactose-binding protein of mammalian cells, by tissue-type transglutaminase. FEBS Lett. 360 (1995) 160–164.

  59. 59.

    Ahmad, N., Gabius, H.J., Andre, S., Kaltner, H., Sabesan, S., Roy, R., Liu, B., Macaluso, F. and Brewer, C.F. Galectin-3 precipitates as a pentamer with synthetic multivalent carbohydrates and forms heterogeneous cross-linked complexes. J. Biol. Chem. 279 (2004) 10841–10847.

  60. 60.

    Lajoie, P., Goetz, J.G., Dennis, J.W. and Nabi, I.R. Lattices, rafts, and scaffolds: domain regulation of receptor signaling at the plasma membrane. J. Cell Biol. 185 (2009) 381–385.

Download references

Author information

Correspondence to Dorota Hoja-Łukowicz.

Electronic supplementary material

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Hoja-Łukowicz, D., Kedracka-Krok, S., Duda, W. et al. The lectin-binding pattern of nucleolin and its interaction with endogenous galectin-3. Cell Mol Biol Lett 19, 461–482 (2014). https://doi.org/10.2478/s11658-014-0206-4

Download citation

Keywords

  • Glycosylation of nucleolin
  • Galectin-3
  • Melanoma
  • Mass spectrometry
  • Confocal microscopy
  • Lectin assay
  • Co-immunoprecipitation