1887

Abstract

yeast cells can enter mammalian cells and may manipulate the host cell environment to favour their own growth and survival. Moreover, fibronectin and several other host extracellular matrix proteins are recognized by various components of the yeast cell extracts. The present study was designed to isolate and characterize a fibronectin-binding protein from . We also compared strain 18, tested before (Pb18a) and after (Pb18b) animal passage, in relation to its adhesion and invasion processes. Extracts from both samples, when cultured on blood agar solid medium, showed higher levels of protein expression than when the same samples were cultured on Fava-Netto solid medium, as demonstrated by two-dimensional electrophoresis and SDS-PAGE. Also, both Pb18a and Pb18b exhibited stronger adhesion to A549 epithelial cells when cultured on blood agar medium than when cultured on Fava-Netto medium. Ligand affinity binding assays revealed a protein of 54 kDa and pI 5.6 in cell-free extracts with the properties of a fibronectin-binding adhesin, which was characterized by tryptic digestion and mass spectroscopy as a homologue of enolase from . Antibody raised against this 54 kDa protein abolished 80 % of adhesion to A549 epithelial cells. Our results demonstrate that produces a fibronectin-binding adhesin, irrespective of the culture medium, and that this activity can be inhibited by a specific antibody and is involved in the adhesion of the fungus to pulmonary epithelial cells.

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2009-06-01
2019-10-15
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References

  1. Andreotti, P. F., Silva, J. L. M., Bailão, A. M., Almeida Soares, C. M., Benard, G., Soares, C. P. & Mendes-Giannini, M. J. S. ( 2005; ). Isolation and partial characterization of a 30 kDa adhesin from Paracoccidioides brasiliensis. Microbes Infect 7, 875–881.[CrossRef]
    [Google Scholar]
  2. Barbosa, M. S., Bao, S. N., Andreotti, P. F., Faria, F. P., Felipe, M. S., Santos Feitosa, L., Mendes-Giannini, M. J. S. & Soares, C. M. ( 2006; ). Glyceraldehyde-3-phosphate dehydrogenase of Paracoccidioides brasiliensis is a cell surface protein involved in fungal adhesion to extracellular matrix proteins and interaction with cells. Infect Immun 74, 382–389.[CrossRef]
    [Google Scholar]
  3. Bergmann, S., Rohde, M., Chhatwal, G. S. & Hammerschmidt, S. ( 2001; ). α-Enolase of Streptococcus pneumoniae is a plasmin(ogen)-binding protein displayed on the bacterial cell surface. Mol Microbiol 40, 1273–1287.[CrossRef]
    [Google Scholar]
  4. Bernal, D., de la Rubia, J. E., Carrasco-Abad, A. M., Toledo, R., Mas-Coma, S. & Marcilla, A. ( 2004; ). Identification of enolase as a plasminogen-binding protein in excretory-secretory products of Fasciola hepatica. FEBS Lett 563, 203–206.[CrossRef]
    [Google Scholar]
  5. Blotta, M. H. S. & Camargo, Z. P. ( 1993; ). Immunological response to cell-free antigens of Paracoccidioides brasiliensis: relationship with clinical forms. J Clin Microbiol 31, 671–676.
    [Google Scholar]
  6. Brummer, E., Castaneda, E. & Restrepo, A. ( 1993; ). Paracoccidioidomycosis: an update. Clin Microbiol Rev 6, 89–117.
    [Google Scholar]
  7. Carneiro, C. R. W., Postol, E., Nomizo, R., Reis, L. F. & Brentani, R. R. ( 2004; ). Identification of enolase as a laminin-binding protein on the surface of Staphylococcus aureus. Microbes Infect 6, 604–608.[CrossRef]
    [Google Scholar]
  8. Coleman, J. L., Gebbia, J. A., Piesman, J., Dengen, J. L., Bugge, T. H. & Benach, J. L. ( 1997; ). Plasminogen is required for efficient dissemination of B. burgdorferi in ticks and for enhancement of spirochetemia in mice. Cell 89, 1111–1119.[CrossRef]
    [Google Scholar]
  9. Coltri, K. C., Fortunato, A. S. C., Cardoso, M. L. G., Pinzan, C. F., Ruas, L. P., Mariano, V. S., Martinez, J. C. R., Panunto-Castelo, A. & Roque-Barreira, M. C. ( 2006; ). Paracoccin, a GlcNac-binding lectin from Paracoccidioides brasiliensis, binds to laminin and induces TNF-α production by macrophages. Microbes Infect 8, 704–713.[CrossRef]
    [Google Scholar]
  10. Esquenazi, D., de Souza, W., Alviano, C. S. & Rozental, S. ( 2003; ). The role of surface carbohydrates on the interaction of microconidia of Trichophyton mentagrophytes with epithelial cells. FEMS Immunol Med Microbiol 35, 113–123.[CrossRef]
    [Google Scholar]
  11. Fava-Netto, C. ( 1961; ). Contribuição para o estudo imunológico da blastomicose de Lutz. Rev Inst Adolfo Lutz 21, 99–194.
    [Google Scholar]
  12. Gonzalez, A., Gomez, B. L., Diez, S., Hernandez, O., Restrepo, A., Hamilton, A. J. & Cano, L. E. ( 2005; ). Purification and partial characterization of a Paracoccidioides brasiliensis protein with capacity to bind to extracellular matrix proteins. Infect Immun 73, 2486–2495.[CrossRef]
    [Google Scholar]
  13. Hanna, S. A., Monteiro da Silva, J. L. & Mendes-Giannini, M. J. S. ( 2000; ). Adherence and intracellular parasitism of Paracoccidioides brasiliensis in Vero cells. Microbes Infect 2, 877–884.[CrossRef]
    [Google Scholar]
  14. Klotz, S. A., Rutten, M. J., Smith, R. L., Babcock, S. R. & Cunningham, M. D. ( 1993; ). Adherence of Candida albicans to immobilized extracellular matrix proteins is mediated by calcium-dependent surface glycoproteins. Microb Pathog 14, 133–147.[CrossRef]
    [Google Scholar]
  15. Kottom, T. J., Kohler, J. R., Thomas, C. F., Jr, Fink, G. R. & Limper, A. H. ( 2003; ). Lung epithelial cells and extracellular matrix components induce expression of Pneumocystis carinii STE20, a gene complementing the mating and pseudohyphal growth defects of STE20 mutant yeast. Infect Immun 71, 6463–6471.[CrossRef]
    [Google Scholar]
  16. Laemmli, U. K. ( 1970; ). Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227, 680–685.[CrossRef]
    [Google Scholar]
  17. Lähteenmäki, K., Virkola, R., Pouttu, R., Kuusela, P., Kukkonen, M. & Korhonen, T. K. ( 1995; ). Bacterial plasminogen receptors: in vitro evidence for a role in degradation of the mammalian extracellular matrix. Infect Immun 63, 3659–3664.
    [Google Scholar]
  18. Mendes-Giannini, M. J. S., Taylor, M. L., Bouchara, J. B., Burger, E., Calich, V. L. G., Escalante, E. D., Hanna, S. A., Lenzi, H. L., Machado, M. P. & other authors ( 2000; ). Pathogenesis II: fungal responses to host responses: interaction of host cells with fungi. Med Mycol 38, 113–123.[CrossRef]
    [Google Scholar]
  19. Mendes-Giannini, M. J. S., Soares, C. P., Silva, J. L. M. & Andreotti, P. F. ( 2005; ). Interaction of pathogenic fungi with host cells: molecular and cellular approaches. FEMS Immunol Med Microbiol 45, 383–394.[CrossRef]
    [Google Scholar]
  20. Mendes-Giannini, M. J. S., Andreotti, P. F., Vincenzi, L. R., Silva, J. L. M., Lenzi, H. L., Benard, G., Zancopé-Oliveira, R., Matos Guedes, H. L. & Soares, C. P. ( 2006; ). Binding of extracellular matrix proteins to Paracoccidioides brasiliensis. Microbes Infect 8, 1550–1559.[CrossRef]
    [Google Scholar]
  21. Mundodi, V., Kucknoor, A. S. & Alderete, J. F. ( 2008; ). Immunogenic and plasminogen-binding surface-associated alpha-enolase of Trichomonas vaginalis. Infect Immun 76, 523–531.[CrossRef]
    [Google Scholar]
  22. Neophytou, P. I., Ozegbe, P., Healey, D., Quartey-Papafio, R., Cooke, A. & Hutton, J. C. ( 1996; ). Development of a procedure for the direct cloning of T-cell epitopes using bacterial expression systems. J Immunol Methods 196, 63–72.[CrossRef]
    [Google Scholar]
  23. O'Farrell, P. H. ( 1975; ). High resolution two-dimensional electrophoresis of proteins. J Biol Chem 250, 4007–4021.
    [Google Scholar]
  24. Ofek, I., Kahane, I. & Sharon, N. ( 1996; ). Toward anti-adhesion therapy for microbial diseases. Trends Microbiol 4, 297–299.[CrossRef]
    [Google Scholar]
  25. Pancholi, V. ( 2001; ). Multifunctional alpha-enolase: its role in diseases. Cell Mol Life Sci 58, 902–920.[CrossRef]
    [Google Scholar]
  26. Pearlstein, E., Gold, L. I. & Garcia-Pardo, A. ( 1980; ). Fibronectin: a review of its structure and biological activity. Mol Cell Biochem 29, 103–128.
    [Google Scholar]
  27. Pendrak, M. L., Krutzsch, H. C. & Roberts, D. D. ( 2000; ). Structural requirements for hemoglobin to induce fibronectin receptor expression in Candida albicans. Biochemistry 39, 16110–16118.[CrossRef]
    [Google Scholar]
  28. Pereira, L. A., Báo, S. N., Barbosa, M. S., Silva, J. L., Felipe, M. S., de Santana, J. M., Mendes-Giannini, M. J. & Almeida Soares, C. M. ( 2007; ). Analysis of the Paracoccidioides brasiliensis triosephosphate isomerase suggests the potential for adhesin function. FEMS Yeast Res 7, 1381–1388.[CrossRef]
    [Google Scholar]
  29. Ruoslahti, E. ( 1988; ). Fibronectin and its receptors. Annu Rev Biochem 57, 375–413.[CrossRef]
    [Google Scholar]
  30. Sa-Pereira, P., Duarte, J. & Costa-Ferreira, M. ( 2000; ). Electroelution as a simple and fast protein purification method: isolation of an extracellular xylanase from Bacillus sp. CCMI 966. Enzyme Microb Technol 27, 95–99.[CrossRef]
    [Google Scholar]
  31. Towbin, H., Staehelin, T. & Gordon, J. ( 1979; ). Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A 76, 4350–4352.[CrossRef]
    [Google Scholar]
  32. Vicentini, A. P., Gesztesi, J. L., Franco, M. F., Souza, W., Moraes, J. Z., Travassos, L. R. & Lopes, J. D. ( 1994; ). Binding of Paracoccidioides brasiliensis to laminin through surface glycoprotein gp43 leads to enhancement of fungal pathogenesis. Infect Immun 62, 1465–1469.
    [Google Scholar]
  33. Yan, S., Rodrigues, R. G. & Roberts, D. D. ( 1998; ). Hemoglobin-induced binding of Candida albicans to the cell-binding domain of fibronectin is independent of the Arg-Gly-Asp sequence. Infect Immun 66, 1904–1909.
    [Google Scholar]
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