1887

Abstract

is a human diarrhoeal pathogen that is a major cause of gastrointestinal disease and death worldwide. Pathogenic strains are characterized by the presence of a pathogenicity island (VPI) that encodes virulence factors, including the toxin co-regulated pilus (TCP). TagA is encoded within the VPI and is positively co-regulated with cholera toxin and TCP. TagA is a sequelogue of the StcE mucinase of O157 : H7. We investigated whether this sequence homology reflected a conserved enzymic substrate profile. TagA exhibited metalloprotease activity toward crude purified mucins, salivary mucin and LS174T goblet cell surface mucin. Like StcE, TagA did not cleave general protease substrates, but unlike StcE, TagA did not cleave the mucin-like serpin C1 esterase inhibitor. Both proteins cleaved the immune cell surface mucin CD43, but TagA demonstrated reduced enzymic efficiency relative to StcE. TagA was expressed and secreted by under ToxR-dependent conditions. A -deficient strain showed no defect in a model of attachment to the HEp-2 cell line; however, overexpression of a proteolytically inactive mutant, TagA(E433D), caused a significant increase in attachment. The increased attachment was reduced by pretreatment of epithelial monolayers with active TagA. Our results indicate that TagA is a mucinase and suggest that TagA may directly modify host cell surface molecules during infection.

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2011-02-01
2019-10-20
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References

  1. Austin, B., Austin, D., Sutherland, R., Thompson, F. & Swings, J. ( 2005; ). Pathogenicity of vibrios to rainbow trout (Oncorhynchus mykiss, Walbaum) and Artemia nauplii. Environ Microbiol 7, 1488–1495.[CrossRef]
    [Google Scholar]
  2. Beaz Hidalgo, R., Cleenwerck, I., Balboa, S., De Wachter, M., Thompson, F. L., Swings, J., De Vos, P. & Romalde, J. L. ( 2008; ). Diversity of Vibrios associated with reared clams in Galicia (NW Spain). Syst Appl Microbiol 31, 215–222.[CrossRef]
    [Google Scholar]
  3. Bina, J., Zhu, J., Dziejman, M., Faruque, S., Calderwood, S. & Mekalanos, J. ( 2003; ). ToxR regulon of Vibrio cholerae and its expression in vibrios shed by cholera patients. Proc Natl Acad Sci U S A 100, 2801–2806.[CrossRef]
    [Google Scholar]
  4. Booth, B. A., Boesman-Finkelstein, M. & Finkelstein, R. A. ( 1983; ). Vibrio cholerae soluble hemagglutinin/protease is a metalloenzyme. Infect Immun 42, 639–644.
    [Google Scholar]
  5. Booth, B. A., Boesman-Finkelstein, M. & Finkelstein, R. A. ( 1984; ). Vibrio cholerae hemagglutinin/protease nicks cholera enterotoxin. Infect Immun 45, 558–560.
    [Google Scholar]
  6. Bos, M. P., Robert, V. & Tommassen, J. ( 2007; ). Biogenesis of the Gram-negative bacterial outer membrane. Annu Rev Microbiol 61, 191–214.[CrossRef]
    [Google Scholar]
  7. Burnet, F. M. ( 1949; ). Ovomucin as a substrate for the mucinolytic enzymes of V cholerae filtrates. Aust J Exp Biol Med Sci 27, 245–252.[CrossRef]
    [Google Scholar]
  8. Champion, G. A., Neely, M. N., Brennan, M. A. & DiRita, V. J. ( 1997; ). A branch in the ToxR regulatory cascade of Vibrio cholerae revealed by characterization of toxT mutant strains. Mol Microbiol 23, 323–331.[CrossRef]
    [Google Scholar]
  9. d'Enfert, C., Chapon, C. & Pugsley, A. P. ( 1987; ). Export and secretion of the lipoprotein pullulanase by Klebsiella pneumoniae. Mol Microbiol 1, 107–116.[CrossRef]
    [Google Scholar]
  10. Finkelstein, R. A., Boesman-Finkelstein, M. & Holt, P. ( 1983; ). Vibrio cholerae hemagglutinin/lectin/protease hydrolyzes fibronectin and ovomucin: F.M. Burnet revisited. Proc Natl Acad Sci U S A 80, 1092–1095.[CrossRef]
    [Google Scholar]
  11. Finkelstein, R. A., Boesman-Finkelstein, M., Chang, Y. & Häse, C. C. ( 1992; ). Vibrio cholerae hemagglutinin/protease, colonial variation, virulence, and detachment. Infect Immun 60, 472–478.
    [Google Scholar]
  12. Grys, T. E., Siegel, M. B., Lathem, W. W. & Welch, R. A. ( 2005; ). The StcE protease contributes to intimate adherence of enterohemorrhagic Escherichia coli O157:H7 to host cells. Infect Immun 73, 1295–1303.[CrossRef]
    [Google Scholar]
  13. Grys, T. E., Walters, L. L. & Welch, R. A. ( 2006; ). Characterization of the StcE protease activity of Escherichia coli O157:H7. J Bacteriol 188, 4646–4653.[CrossRef]
    [Google Scholar]
  14. Guzman, L. M., Belin, D., Carson, M. J. & Beckwith, J. ( 1995; ). Tight regulation, modulation, and high-level expression by vectors containing the arabinose PBAD promoter. J Bacteriol 177, 4121–4130.
    [Google Scholar]
  15. Harrison, L. M., Rallabhandi, P., Michalski, J., Zhou, X., Steyert, S. R., Vogel, S. N. & Kaper, J. B. ( 2008; ). Vibrio cholerae flagellins induce Toll-like receptor 5-mediated interleukin-8 production through mitogen-activated protein kinase and NF-kappaB activation. Infect Immun 76, 5524–5534.[CrossRef]
    [Google Scholar]
  16. Häse, C. C. & Finkelstein, R. A. ( 1990; ). Comparison of the Vibrio cholerae hemagglutinin/protease and the Pseudomonas aeruginosa elastase. Infect Immun 58, 4011–4015.
    [Google Scholar]
  17. Häse, C. C. & Finkelstein, R. A. ( 1991; ). Cloning and nucleotide sequence of the Vibrio cholerae hemagglutinin/protease (HA/protease) gene and construction of an HA/protease-negative strain. J Bacteriol 173, 3311–3317.
    [Google Scholar]
  18. Herrero, M., de Lorenzo, V. & Timmis, K. N. ( 1990; ). Transposon vectors containing non-antibiotic resistance selection markers for cloning and stable chromosomal insertion of foreign genes in Gram-negative bacteria. J Bacteriol 172, 6557–6567.
    [Google Scholar]
  19. Herrington, D. A., Hall, R. H., Losonsky, G., Mekalanos, J. J., Taylor, R. K. & Levine, M. M. ( 1988; ). Toxin, toxin-coregulated pili, and the toxR regulon are essential for Vibrio cholerae pathogenesis in humans. J Exp Med 168, 1487–1492.[CrossRef]
    [Google Scholar]
  20. Ho, T. D., Davis, B. M., Ritchie, J. M. & Waldor, M. K. ( 2008; ). Type 2 secretion promotes enterohemorrhagic Escherichia coli adherence and intestinal colonization. Infect Immun 76, 1858–1865.[CrossRef]
    [Google Scholar]
  21. Kaper, J. B., Morris, J. G. & Levine, M. M. ( 1995; ). Cholera. Clin Microbiol Rev 8, 48–86.
    [Google Scholar]
  22. Karaolis, D. K., Johnson, J. A., Bailey, C. C., Boedeker, E. C., Kaper, J. B. & Reeves, P. R. ( 1998; ). A Vibrio cholerae pathogenicity island associated with epidemic and pandemic strains. Proc Natl Acad Sci U S A 95, 3134–3139.[CrossRef]
    [Google Scholar]
  23. Karaolis, D. K., Lan, R., Kaper, J. B. & Reeves, P. R. ( 2001; ). Comparison of Vibrio cholerae pathogenicity islands in sixth and seventh pandemic strains. Infect Immun 69, 1947–1952.[CrossRef]
    [Google Scholar]
  24. Lathem, W. W., Grys, T. E., Witowski, S. E., Torres, A. G., Kaper, J. B., Tarr, P. I. & Welch, R. A. ( 2002; ). StcE, a metalloprotease secreted by Escherichia coli O157:H7, specifically cleaves C1 esterase inhibitor. Mol Microbiol 45, 277–288.[CrossRef]
    [Google Scholar]
  25. Lathem, W. W., Bergsbaken, T. & Welch, R. A. ( 2004; ). Potentiation of C1 esterase inhibitor by StcE, a metalloprotease secreted by Escherichia coli O157:H7. J Exp Med 199, 1077–1087.[CrossRef]
    [Google Scholar]
  26. Levine, M. M., Kaper, J. B., Black, R. E. & Clements, M. L. ( 1983; ). New knowledge on pathogenesis of bacterial enteric infections as applied to vaccine development. Microbiol Rev 47, 510–550.
    [Google Scholar]
  27. Matson, J. S., Withey, J. H. & DiRita, V. J. ( 2007; ). Regulatory networks controlling Vibrio cholerae virulence gene expression. Infect Immun 75, 5542–5549.[CrossRef]
    [Google Scholar]
  28. Miller, V. L. & Mekalanos, J. J. ( 1988; ). A novel suicide vector and its use in construction of insertion mutations: osmoregulation of outer membrane proteins and virulence determinants in Vibrio cholerae requires toxR. J Bacteriol 170, 2575–2583.
    [Google Scholar]
  29. Pillai, L., Sha, J., Erova, T. E., Fadl, A. A., Khajanchi, B. K. & Chopra, A. K. ( 2006; ). Molecular and functional characterization of a ToxR-regulated lipoprotein from a clinical isolate of Aeromonas hydrophila. Infect Immun 74, 3742–3755.[CrossRef]
    [Google Scholar]
  30. Rawlings, N. D., Barrett, A. J. & Bateman, A. ( 2010; ). merops: the peptidase database. Nucleic Acids Res 38, D227–D233.[CrossRef]
    [Google Scholar]
  31. Rui, H., Ritchie, J. M., Bronson, R. T., Mekalanos, J. J., Zhang, Y. & Waldor, M. K. ( 2010; ). Reactogenicity of live-attenuated Vibrio cholerae vaccines is dependent on flagellins. Proc Natl Acad Sci U S A 107, 4359–4364.[CrossRef]
    [Google Scholar]
  32. Sack, D. A., Sack, R. B., Nair, G. B. & Siddique, A. K. ( 2004; ). Cholera. Lancet 363, 223–233.[CrossRef]
    [Google Scholar]
  33. Schneider, D. R. & Parker, C. D. ( 1982; ). Purification and characterization of the mucinase of Vibrio cholerae. J Infect Dis 145, 474–482.[CrossRef]
    [Google Scholar]
  34. Silva, T. M., Schleupner, M. A., Tacket, C. O., Steiner, T. S., Kaper, J. B., Edelman, R. & Guerrant, R. ( 1996; ). New evidence for an inflammatory component in diarrhea caused by selected new, live attenuated cholera vaccines and by El Tor and Q139 Vibrio cholerae. Infect Immun 64, 2362–2364.
    [Google Scholar]
  35. Silva, A. J., Pham, K. & Benitez, J. A. ( 2003; ). Haemagglutinin/protease expression and mucin gel penetration in El Tor biotype Vibrio cholerae. Microbiology 149, 1883–1891.[CrossRef]
    [Google Scholar]
  36. Silva, A. J., Leitch, G. J., Camilli, A. & Benitez, J. A. ( 2006; ). Contribution of hemagglutinin/protease and motility to the pathogenesis of El Tor biotype cholera. Infect Immun 74, 2072–2079.[CrossRef]
    [Google Scholar]
  37. Skorupski, K. & Taylor, R. K. ( 1996; ). Positive selection vectors for allelic exchange. Gene 169, 47–52.[CrossRef]
    [Google Scholar]
  38. Sperandio, V., Girón, J. A., Silveira, W. D. & Kaper, J. B. ( 1995; ). The OmpU outer membrane protein, a potential adherence factor of Vibrio cholerae. Infect Immun 63, 4433–4438.
    [Google Scholar]
  39. Stathopoulos, C., Hendrixson, D. R., Thanassi, D. G., Hultgren, S. J., St Geme, J. W., III & Curtiss, R., III ( 2000; ). Secretion of virulence determinants by the general secretory pathway in Gram-negative pathogens: an evolving story. Microbes Infect 2, 1061–1072.[CrossRef]
    [Google Scholar]
  40. Stewart-Tull, D. E., Ollar, R. A. & Scobie, T. S. ( 1986; ). Studies on the Vibrio cholerae mucinase complex. I. Enzymic activities associated with the complex. J Med Microbiol 22, 325–333.[CrossRef]
    [Google Scholar]
  41. Szabady, R. L., Lokuta, M. A., Walters, K. B., Huttenlocher, A. & Welch, R. A. ( 2009; ). Modulation of neutrophil function by a secreted mucinase of Escherichia coli O157:H7. PLoS Pathog 5, e1000320.[CrossRef]
    [Google Scholar]
  42. Taylor, R. K., Miller, V. L., Furlong, D. B. & Mekalanos, J. J. ( 1987; ). Use of phoA gene fusions to identify a pilus colonization factor coordinately regulated with cholera toxin. Proc Natl Acad Sci U S A 84, 2833–2837.[CrossRef]
    [Google Scholar]
  43. Thompson, F. L., Iida, T. & Swings, J. ( 2004; ). Biodiversity of vibrios. Microbiol Mol Biol Rev 68, 403–431.[CrossRef]
    [Google Scholar]
  44. Thompson, F. L. & Klose, K. E., AVIB Group ( 2006; ). Vibrio2005: the First International Conference on the Biology of Vibrios. J Bacteriol 188, 4592–4596.[CrossRef]
    [Google Scholar]
  45. Thornton, D. J. & Sheehan, J. K. ( 2004; ). From mucins to mucus: toward a more coherent understanding of this essential barrier. Proc Am Thorac Soc 1, 54–61.[CrossRef]
    [Google Scholar]
  46. Withey, J. H. & Dirita, V. J. ( 2005; ). Vibrio cholerae ToxT independently activates the divergently transcribed aldA and tagA genes. J Bacteriol 187, 7890–7900.[CrossRef]
    [Google Scholar]
  47. Yamamoto, T., Kamano, T., Uchimura, M., Iwanaga, M. & Yokota, T. ( 1988; ). Vibrio cholerae O1 adherence to villi and lymphoid follicle epithelium: in vitro model using formalin-treated human small intestine and correlation between adherence and cell-associated hemagglutinin levels. Infect Immun 56, 3241–3250.
    [Google Scholar]
  48. Zhu, J., Miller, M. B., Vance, R. E., Dziejman, M., Bassler, B. L. & Mekalanos, J. J. ( 2002; ). Quorum-sensing regulators control virulence gene expression in Vibrio cholerae. Proc Natl Acad Sci U S A 99, 3129–3134.[CrossRef]
    [Google Scholar]
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