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Volume 149, Issue 4

Identification and molecular analysis of cable pilus biosynthesis genes in Free

Abstract

is an opportunistic respiratory pathogen in cystic fibrosis patients. One highly transmissible and virulent clone belonging to genomovar IIIa expresses pili with unique cable morphology, which enable the bacterium to bind cytokeratin 13 in epithelial cells. The gene, encoding the major pilin subunit, is often used as a DNA marker to identify potentially virulent isolates. The authors have now cloned and sequenced four additional genes, , , and , in the pilus gene cluster. This work shows that the products of the first four genes of the operon, , , and , are sufficient for pilus assembly on the bacterial surface. Deletion of abrogated pilus assembly and compromised the stability of the CblA protein in the periplasm. In contrast, deletion of resulted in no pili, but there was no effect on expression and stability of the CblA protein subunit. These results, together with protein sequence homologies, predicted structural analyses, and the presence of typical amino acid motifs, are consistent with the assignment of functional roles for CblB as a chaperone that stabilizes the major pilin subunit in the periplasm, and CblD as the initiator of pilus biogenesis. It is also shown that expression of Cbl pili in is not sufficient to mediate the binding of bacteria to the epithelial cell receptor cytokeratin 13, and that still binds to cytokeratin 13 in the absence of Cbl pili, suggesting that additional bacterial components are required for effective binding.

  • Received:
  • Accepted:
  • Published Online:
Keyword(s): CF, cystic fibrosis
SGM
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2003-04-01
2024-04-16
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References

  1. Bakker D., Willemsen P. T., Simons L. H., van Zijderveld F. G., de Graaf F. K. 1992; Characterization of the antigenic and adhesive properties of FaeG, the major subunit of K88 fimbriae. Mol Microbiol 6:247–255
     [Google Scholar]
  2. Brown M. C. M., Western A., Saunders J. R., Humphreys G. O. 1979; Transformation of E. coli C600 by plasmid DNA at different phases of growth. FEMS Microbiol Lett 5:217–222
     [Google Scholar]
  3. Duthy T. G., Manning P. A., Heuzenroeder M. W. 2001; Characterization of the CsfC and CsfD proteins involved in the biogenesis of CS5 pili from enterotoxigenic Escherichia coli . Microb Pathog 31:115–129
     [Google Scholar]
  4. Folkesson A., Advani A., Sukupolvi S., Pfeifer J. D., Normark S., Lofdahl S. 1999; Multiple insertions of fimbrial operons correlate with the evolution of Salmonella responsible for human disease. Mol Microbiol 33:612–622
     [Google Scholar]
  5. Froehlich B. J., Karakashian A., Melsen L. R., Wakefield J. C., Scott J. R. 1994; CooC and CooD are required for assembly of CS1 pili. Mol Microbiol 12:387–401
     [Google Scholar]
  6. 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]
  7. Holmgren A., Branden C. I. 1989; Crystal structure of chaperone protein PapD reveals an immunoglobulin fold. Nature 342:248–251
     [Google Scholar]
  8. Hultegren S. J., Normark S., Abraham S. N. 1991; Chaperone-assisted assembly and molecular architecture of adhesive pili. Annu Rev Microbiol 45:383–415
     [Google Scholar]
  9. Irvin R. T., Doig P., Lee K. K., Sastry P. A., Paranchych W., Todd T., Hodges R. S. 1989; Characterization of the Pseudomonas aeruginosa pilus adhesin: confirmation that the pilin structural protein subunit contains a human epithelial cell-binding domain. Infect Immun 57:3720–3726
     [Google Scholar]
  10. Jacobs A. A., Simons B. H., de Graaf F. K. 1987; The role of lysine-132 and arginine-136 in the receptor-binding domain of the K99 fibrillar subunit. EMBO J 6:1805–1808
     [Google Scholar]
  11. Kelley L. A., MacCallum R. M., Sternberg M. J. E. 2000; Enhanced genome annotation using structural profiles in the program 3D-PSSM. J Mol Biol 299:499–520
     [Google Scholar]
  12. Krogfelt K. A. 1991; Bacterial adhesion: genetics, biogenesis, and role in pathogenesis of fimbrial adhesins of Escherichia coli . Rev Infect Dis 13:721–735
     [Google Scholar]
  13. Kuehn M. J., Ogg D. J., Kihlberg J., Slonim L. N., Flemmer K., Bergfors T., Hultgren S. J. 1993; Structural basis of pilus subunit recognition by the PapD chaperone. Science 262:1234–1241
     [Google Scholar]
  14. Lefebre M. D., Valvano M. A. 2002; Construction and evaluation of plasmid vectors optimized for constitutive and regulated gene expression in Burkholderia cepacia complex isolates. Appl Environ Microbiol 68:5956–5964
     [Google Scholar]
  15. Lessie T. G., Hendrickson W., Manning B. D., Devereux R. 1996; Genomic complexity and plasticity of Burkholderia cepacia . FEMS Microbiol Lett 144:117–128
     [Google Scholar]
  16. Mahenthiralingam E., Baldwin A., Vandamme P. 2002; Burkholderia cepacia complex infection in patients with cystic fibrosis. J Med Microbiol 51:533–538
     [Google Scholar]
  17. Nielsen H., Engelbrecht J., Brunak S., von Heijne G. 1997; Identification of prokaryotic and eukaryotic signal peptides and prediction of their cleavage sites. Protein Eng 10:1–6
     [Google Scholar]
  18. Nzula S., Vandamme P., Govan J. R. W. 2002; Influence of taxonomic status on the in vitro antimicrobial susceptibility of the Burkholderia cepacia complex. J Antimicrob Chemother 50:265–269
     [Google Scholar]
  19. Pugsley A. P. 1993; The complete general secretory pathway in gram-negative bacteria. Microbiol Rev 57:50–108
     [Google Scholar]
  20. Raivio T. L., Silhavy T. J. 2001; Periplasmic stress and ECF sigma factors. Annu Rev Microbiol 55:591–624
     [Google Scholar]
  21. Sajjan U. S., Forstner J. F. 1993; Role of a 22-kilodalton pilin protein in binding of Pseudomonas cepacia to buccal epithelial cells. Infect Immun 61:3157–3163
     [Google Scholar]
  22. Sajjan U. S., Corey M., Karmali M., Forstner J. F. 1991; Binding of Pseudomonas cepacia to normal human intestinal mucin and respiratory mucin from patients with cystic fibrosis. J Clin Invest 89:648–656
     [Google Scholar]
  23. Sajjan U. S., Sun L., Goldstein R., Forstner J. F. 1995; Cable (Cbl) type II pili of cystic fibrosis-associated Burkholderia ( Pseudomonas ) cepacia : nucleotide sequence of the cbl A major subunit pilin gene and novel morphology of the assembled appendage fibers. J Bacteriol 177:1030–1038
     [Google Scholar]
  24. Sajjan U. S., Sylvester F. A., Forstner J. 2000a; Cable-piliated Burkholderia cepacia bind to cytokeratin 13 of epithelial cells. Infect Immun 68:1787–1795
     [Google Scholar]
  25. Sajjan U. S., Wu Y., Kent G., Forstner J. 2000b; Preferential adherence of cable-piliated Burkholderia cepacia to respiratory epithelia of CF knockout mice and human CF lung explants. J Med Microbiol 49:875–885
     [Google Scholar]
  26. Sajjan U., Liu L., Lu A., Spilker T., Forstner J., LiPuma J. 2002; Lack of cable pili expression by cblA -containing Burkholderia cepacia genomovar I. Microbiology 148:3477–3484
     [Google Scholar]
  27. Sakellaris H., Scott J. R. 1998; New tools in an old trade: CS1 pilus morphogenesis. Mol Microbiol 30:681–687
     [Google Scholar]
  28. Sakellaris H., Munson G. P., Scott J. R. 1999; A conserved residue in the tip proteins of CS1 and CFA/I pili of enterotoxigenic Escherichia coli that is essential for adherence. Proc Natl Acad Sci U S A 96:12828–12832
     [Google Scholar]
  29. Scott J. R., Wakefield J. C., Russell P. W., Orndorff P. E., Froehlich B. J. 1992; CooB is required for assembly but not transport of CS1 pilin. Mol Microbiol 6:293–300
     [Google Scholar]
  30. Sun L., Jiang R. Z., Steinbach S. 7 other authors 1995; The emergence of a highly transmissible lineage of cbl + Pseudomonas ( Burkholderia ) cepacia causing CF centre epidemics in North America and Britain. Nat Med 1:661–666
     [Google Scholar]
  31. Thanassi D. G., Saulino E. T., Lombardo M., Roth R., Heuser J., Hultgren S. J. 1998; The PapC usher forms an oligomeric channel: implications for pilus biogenesis across the outer membrane. Proc Natl Acad Sci U S A 95:3146–3151
     [Google Scholar]
  32. Voegele K., Sakellaris H., Scott J. R. 1997; CooB plays a chaperone-like role for the proteins involved in formation of CS1 pili of enterotoxigenic Escherichia coli . Proc Natl Acad Sci U S A 94:13257–13261
     [Google Scholar]
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Identification and molecular analysis of cable pilus biosynthesis genes in Burkholderia cepacia
Microbiology 149, 961 (2003); https://doi.org/10.1099/mic.0.26176-0
/content/journal/micro/10.1099/mic.0.26176-0
/content/journal/micro/10.1099/mic.0.26176-0
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