Advertisement

LacdiNAcylation of N-glycans in MDA-MB-231 human breast cancer cells results in changes in morphological appearance and adhesive properties of the cells

  • Kiyoko HiranoEmail author
  • Yoshio Takada
  • Kiyoshi Furukawa
Original Paper

Abstract

We demonstrated previously that the expression of the disaccharide, GalNAcβ1 → 4GlcNAc (LacdiNAc), on N-glycans of cell surface glycoproteins in MDA-MB-231 human breast cancer cells suppresses their malignant properties such as tumor formation in nude mice. Here, we report changes in the morphological appearance and adhesive properties of two kinds of clonal cells of MDA-MB-231 cells overexpressing β4-N-acetyl-galactosaminyltransferase 4. The clonal cells exhibited a cobble stone-like shape as compared to a spindle-like shape of the mock-transfected cells and the original MDA-MB-231 cells. This was associated with an increased expression of cell surface E-cadherin, a marker of epithelial cells, and a decreased expression of N-cadherin, vimentin, α-smooth muscle actin and ZEB1, markers of mesenchymal cells. In addition, the clonal cells showed a lower migratory activity compared to the mock-transfected cells by wound-healing assay. These results suggest that mesenchymal–epithelial transition may be occurring in these clonal cells. Furthermore, increased adhesion to extracellular matrix proteins such as fibronectin, collagen type I, collagen type IV, and laminin was observed. The clonal cells spread and enlarged, whereas the mock-transfected cells demonstrated poor spreading on laminin-coated plates in the absence of fetal calf serum, indicating that expression of LacdiNAc on cell surface glycoproteins results in changes in cell adhesive and spreading properties particularly to laminin.

Keywords

LacdiNAc Human breast cancer cells Mesenchymal–epithelial transition Extracellular matrices Adhesiveness and spreading 

Notes

Acknowledgements

We are grateful to Dr. Tomoya Isaji and Dr. Jianquo Gu at Tohoku Medical and Pharmaceutical University, Japan, for their generous gift of anti-β1-integrin antibody.

Compliance with ethical standards

Conflict of interest

The authors declare no conflict of interest.

References

  1. Adamczyk B, Jin C, Polom K, Muñoz P, Rojas-Macias MA, Zeeberg D, Borén M, Roviello F, Karlsson NG (2018) Sample handling of gastric tissue and O-glycan alterations in paired gastric cancer and non-tumorigenic tissues. Sci Rep 8:242CrossRefGoogle Scholar
  2. Alam N, Goel HL, Zarif MJ, Butterfield JE, Perkins HM, Sansoucy BG, Sawyer TK, Languino LR (2007) The integrin-growth factor receptor duet. J Cell Physiol 213:649–653CrossRefGoogle Scholar
  3. Anugraham M, Jacob F, Everest-Dass AV, Schoetzau A, Nixdorf S, Hacker NF, Fink D, Heinzelmann-Schwarz V, Packer NH (2017) Tissue glycomics distinguish tumour sites in women with advanced serous adenocarcinoma. Mol Oncol 11:1595–1615CrossRefGoogle Scholar
  4. Asada M, Furukawa K, Segawa K, Endo T, Kobata A (1997) Increased expression of highly branched N-glycans at cell surface is correlated with the malignant phenotypes of mouse tumor cells. Cancer Res 57:1073–1080PubMedGoogle Scholar
  5. Austin P, Freeman SA, Gray CA, Gold MR, Vogl AW, Andersen RJ, Roberge M, Roskelley CD (2013) The invasion inhibitor sarasinoside A1 reverses mesenchymal tumor transformation in an E-cadherin-independent manner. Mol Cancer Res 11:530–540CrossRefGoogle Scholar
  6. Blick T, Widodo E, Hugo H, Waltham M, Lenburg ME, Neve RM, Tompson EW (2008) Epithelial mesenchymal transition traits in human breast cancer cell lines. Clin Exp Metastasis 25:629–642CrossRefGoogle Scholar
  7. Bray F, Ferlay J, Soerjomataram I, Siegel RL, Torre LA, Jemal A (2018) Global cancer statistics 2018: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries. CA Caner J Clin 68:394–424CrossRefGoogle Scholar
  8. Castillo LF, Tascón R, Lago Huvelle MR, Novack G, Llorens MC, Dos Santos AF, Shortrede J, Cabanillas AM, de Kier Bal, Joffé E, Labriola L, Peters MG (2016) Glypican-3 induces a mesenchymal to epithelial transition in human breast cancer cells. Oncotarget 7:60133–60154CrossRefGoogle Scholar
  9. Che MI, Huang J, Hung JS, Lin YC, Huang MJ, Lai HS, Hsu WM, Liang JT, Huang MC (2014) β1,4-N-acetylgalactosaminyltransferase III modulates cancer stemness through EGFR signaling pathway in colon cancer cells. Oncotarget 15:3673–3684Google Scholar
  10. Colak S, Dijke P (2017) Targeting TGF-β-signaling in cancer. Trend Cancer 3:56–71CrossRefGoogle Scholar
  11. Derynck R, Zhang YE (2003) Smad-dependent and Smad-independent pathways in TGF-β family signalling. Nature 425:577–584CrossRefGoogle Scholar
  12. Fukushima K, Satoh T, Baba S, Yamashita K (2010) α1,2-Fucosylated and β-N-acetylgalactosaminylated prostate-specific antigen as an efficient marker of prostatic cancer. Glycobiology 20:452–460CrossRefGoogle Scholar
  13. Furukawa K, Kitamura N, Sato T, Hiraizumi S (2001) Differentiation-associated expression of β-N-acetylgalactosaminylated N-linked oligosaccharides in mammary epithelial cells. Adv Exp Med Biol 491:313–323CrossRefGoogle Scholar
  14. Gu J, Taniguchi N (2008) Potential of N-glycan in cell adhesion and migration as either a positive or negative regulator. Cell Adh Migr 2:243–245CrossRefGoogle Scholar
  15. Gu J, Isaji T, Xu Q, Kariya Y, Gu W, Fukuda T, Du Y (2012) Potential roles of N-glycosylation in cell adhesion. Glycoconj J 29:599–607CrossRefGoogle Scholar
  16. Gupta I, Sareyeldin RM, Al-Hashimi I, Al-Thawadi HA, Al Farsi H, Vranic S, Al Moustafa AE (2019) Triple negative breast cancer profile, from gene to microRNA, in relation to ethnicity. Cancers (Basel) 11:363CrossRefGoogle Scholar
  17. Hirano K, Zuber C, Roth J, Ziak M (2003) The proteasome is involved in the degradation of different aquaporin-2 mutants causing nephrogenic diabetes insibidus. Am J Path 163:111–120CrossRefGoogle Scholar
  18. Hirano K, Matsuda A, Shirai T, Furukawa K (2014) Expression of LacdiNAc groups on N-glycans among human tumor is complex. BioMed Res Int.  https://doi.org/10.1155/2014/981627 CrossRefGoogle Scholar
  19. Hirano K, Matsuda A, Kuji R, Nakandakari S, Shirai T, Furukawa K (2015) Enhanced expression of the β4-N-acetylgalactosaminyltransferase 4 gene impairs tumor growth of human breast cancer cells. Biochem Biophys Res Commun 461:80–85CrossRefGoogle Scholar
  20. Hou S, Isaji T, Hang Q, Im S, Fukuda T, Gu J (2016) Distinct effects of β1 integrin on cell proliferation and cellular signaling in MDA-MB-231 breast cancer cells. Sci Rep 6:18430CrossRefGoogle Scholar
  21. Hsu WM, Che MI, Liao YF, Chang HH, Chen CH, Huang YM, Jeng YM, Huang J, Quon MJ, Lee H, Huang HC, Huang MC (2011) B4GALNT3 expression predicts a favorable prognosis and suppresses cell migration and invasion via β1-integrin signaling in neuroblastoma. Am J Path 179:1394–1404CrossRefGoogle Scholar
  22. Huang J, Liang JT, Huang HC, Shen TL, Chen HY, Lin NY, Che MI, Lin WC, Huang MC (2007) β1,4-N-acetylgalactosaminyltransferase III enhances malignant phenotypes of colon cancer cells. Mol Cancer Res 5:543–552CrossRefGoogle Scholar
  23. Hwang SY, Park S, Kwon Y (2019) Recent therapeutic trends and promising targets in triple negative breast cancer. Pharmacol Ther 199:30–57CrossRefGoogle Scholar
  24. Isaji T, Gu J, Nishiuchi R, Zhao Y, Takahashi M, Miyoshi E, Honke K, Sekiguchi K, Taniguchi N (2004) Introduction of bisecting GlcNAc into integrin α5β1 reduces ligand binding and down-regulates cell adhesion and cell migration. J Biol Chem 279:19747–19754CrossRefGoogle Scholar
  25. Isaji T, Sato Y, Zhao Y, Miyoshi E, Wada Y, Taniguchi N, Gu J (2006) N-glycosylation of the β-propeller domain of the integrin α5 subunit is essential for α5β1 heterodimerization, expression on the cell surface, and its biological function. J Biol Chem 281:33258–33267CrossRefGoogle Scholar
  26. Kitamura N, Guo S, Sato T, Hiraizumi S, Taka J, Ikekita M, Sawada S, Fujisawa H, Furukawa K (2003) Prognostic significance of reduced expression of β-N-acetyl- galactosaminylated N-linked oligosaccharides in human breast cancer. Int J Cancer 105:533–541CrossRefGoogle Scholar
  27. Lamouille S, Xu J, Derynck R (2014) Molecular mechanisms of epithelial–mesenchymal transition. Nat Rev Mol Cell Biol 15:178–196CrossRefGoogle Scholar
  28. Li X, Wang X, Tan Z, Chen S, Guan F (2016) Role of glycans in cancer cells undergoing epithelial–mesenchymal transition. Front Oncol 6:33CrossRefGoogle Scholar
  29. Lin D, Kuang G, Wan J, Zhang X, Li H, Gong X, Li H (2017) Luteolin suppresses the metastasis of triple-negative breast cancer by reversing epithelial-to-mesenchymal transition via downregulation of β-catenin expression. Oncol Rep 37:895–902CrossRefGoogle Scholar
  30. Lu J, Isaji T, Im S, Fukuda T, Hashii N, Takakura D, Kawasaki N, Gu J (2014) β-Galactoside α2,6-sialyltranferase 1 promotes transforming growth factor-β-mediated epithelial-mesenchymal transition. J Biol Chem 289:34627–34641CrossRefGoogle Scholar
  31. Machado E, Kandzia S, Carilho R, Altevogt P, Conradt HS, Costa J (2011) N-Glycosylation of total cellular glycoproteins from the human ovarian carcinoma SKOV3 cell line and of recombinantly expressed human erythropoietin. Glycobiology 21:376–386CrossRefGoogle Scholar
  32. Miranti CK, Brugge JS (2002) Sensing the environment: a historical perspective on integrin signal transduction. Nat Cell Biol 4:E83–E90CrossRefGoogle Scholar
  33. Nakata N, Furukawa K, Greenwalt DE, Sato T, Kobata A (1993) Structural study of the sugar chains of CD36 purified from bovine mammary epithelial cells: occurrence of novel hybrid-type sugar chains containing the Neu5Acα2 → 6GalNAcβ1 → 4GlcNAc and the Manα1 → 2Manα1 → 3Manα1 → 6Man groups. Biochemistry 32:4369–4383CrossRefGoogle Scholar
  34. Sarrió D, Rodriguez-Pinilla SM, Hardisson D, Cano A, Moreno-Bueno G, Palacios J (2008) Epithelial-mesenchymal transition in breast cancer relates to the basal-like phenotype. Cancer Res 68:989–997CrossRefGoogle Scholar
  35. Sasaki N, Shinomi M, Hirano K, Ui-Tei K, Nishihara S (2011) LacdiNAc (GalNAcβ1-4GlcNAc) contributes to self-renewal of mouse embryonic stem cells by regulating leukemia inhibitory factor/STAT3 signaling. Stem Cell 29:641–650CrossRefGoogle Scholar
  36. Sato T, Taka J, Aoki N, Matsuda T, Furukawa K (1997) Expression β-N-acetylgalactosaminylated N-linked sugar chain is associated with functional differentiation of bovine mammary gland. J Biochem 122:1068–1073CrossRefGoogle Scholar
  37. Sato T, Gotoh M, Kiyohara K, Kameyama A, Kubota T, Kikuchi N, Ishizuka Y, Iwasaki H, Togayachi A, Kudo T, Ohkura T, Nakanishi H, Narimatsu H (2003) Molecular cloning and characterization of a novel human β1,4-N-acetylgalactosaminyl-transferase, β4GalNAc-T3, responsible for the synthesis of N, N′-diacetyl-lactosediamine, GalNAcβ1-4GlcNAc. J Biol Chem 278:47534–47544CrossRefGoogle Scholar
  38. Thiery JP, Sleeman JP (2006) Complex networks orchestrate epithelial-mesenchymal transitions. Nat Rev Mol Cell Biol 7:131–142CrossRefGoogle Scholar
  39. van ’t Veer LJ, Dai H, van de Vijver MJ, He YD, Hart AA, Mao M, Peterse HL, van der Kooy K, Marton MJ, Witteveen AT, Schreiber GJ, Kerkhoven RM, Roberts C, Linsley PS, Bernards R, Friend SH (2002) Gene expression profiling predicts clinical outcome of breast cancer. Nature 415:530–536CrossRefGoogle Scholar
  40. Varki A (1993) Biological roles of oligosaccharides: all of the theories are correct. Glycobology 3:97–130CrossRefGoogle Scholar
  41. Wells A, Yates C, Shepard CR (2008) E-cadherin as an indicator of mesenchymal to epithelial reverting transitions during the metastatic seeding of disseminated carcinomas. Clin Exp Metastasis 25:621–628CrossRefGoogle Scholar
  42. Wu Y, Sarkissyan M, Vadgama JV (2016) Epithelial–mesenchymal transition and breast cancer. J Clin Med 5(2):13.  https://doi.org/10.3390/jcm5020013 CrossRefGoogle Scholar
  43. Xu Q, Isaji T, Lu Y, Gu W, Kondo M, Fukuda T, Du Y, Gu J (2012) Roles of N-acetylglucosaminyltransferase III in epithelial-to-mesenchymal transition induced by transforming growth factor β1 (TGF-β1) in epithelial cell lines. J Biol Chem 287:16563–16574CrossRefGoogle Scholar
  44. Xu Q, Niu X, Wang W, Yang W, Du Y, Gu J, Song L (2017) Specific N-glycan alterations are coupled in EMT induced by different density cultivation of MCF 10A epithelial cells. Glycoconj J 34:219–227CrossRefGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Laboratory of GlycobiologyThe Noguchi InstituteTokyoJapan
  2. 2.Laboratory of Glycobiology, Department of BioengineeringNagaoka University of TechnologyNagaokaJapan

Personalised recommendations