1887

Abstract

produces fimbria-like structures that are involved with the bacterium's adhesion to cellulose. The subunit protein has been identified in strain 8 (CbpC) and strain 20 (GP25) and both are type IV fimbrial (Pil) proteins. The presence of a locus that is organized similarly in both strains is reported here together with the results of an initial examination of a second Pil protein. Downstream of the / gene (hereafter referred to as ) is a second pilin gene (). Northern blot analysis of and transcripts showed that the transcript is much more abundant in 8, and real-time PCR was used to measure and transcript abundance in 20 and its adhesion-defective mutant D5. Similar to the findings with 8, the relative expression of in the wild-type strain was 73-fold higher than that of following growth with cellobiose, and there were only slight differences between the wild-type and mutant strain in and transcript abundances, indicating that neither nor transcription is adversely affected in the mutant strain. Western immunoblots showed that the PilA2 protein is localized primarily to the membrane fraction, and the anti-PilA2 antiserum does not inhibit bacterial adhesion to cellulose. These results suggest that the PilA2 protein plays a role in the synthesis and assembly of type IV fimbriae-like structures by , but its role is restricted to cell-associated functions, rather than as part of the externalized fimbrial structure.

Loading

Article metrics loading...

/content/journal/micro/10.1099/mic.0.27735-0
2005-04-01
2019-10-22
Loading full text...

Full text loading...

/deliver/fulltext/micro/151/4/mic1511291.html?itemId=/content/journal/micro/10.1099/mic.0.27735-0&mimeType=html&fmt=ahah

References

  1. Alm, R. A. & Mattick, J. S. ( 1997; ). Genes involved in the biogenesis and function of type-4 fimbriae in Pseudomonas aeruginosa. Gene 192, 89–98.[CrossRef]
    [Google Scholar]
  2. Altschul, S. F., Gish, W., Miller, W., Myers, E. W. & Lipman, D. J. ( 1990; ). Basic local alignment search tool. J Mol Biol 215, 403–410.[CrossRef]
    [Google Scholar]
  3. Bhaya, D., Bianco, N. R., Bryant, D. & Grossman, A. ( 2000; ). Type IV pilus biogenesis and motility in the cyanobacterium Synechocystis sp. PCC6803. Mol Microbiol 37, 941–951.[CrossRef]
    [Google Scholar]
  4. Bolhuis, A., Broekhuizen, C. P., Sorokin, A., van Roosmalen, M. L., Venema, G., Bron, S., Quax, W. J. & van Dijl, J. M. ( 1998; ). SecDF of Bacillus subtilis, a molecular Siamese twin required for the efficient secretion of proteins. J Biol Chem 273, 21217–21224.[CrossRef]
    [Google Scholar]
  5. Dalrymple, B. & Mattick, J. S. ( 1987; ). An analysis of the organization and evolution of type 4 fimbrial (MePhe) subunit proteins. J Mol Evol 25, 261–269.[CrossRef]
    [Google Scholar]
  6. Devillard, E., Goodheart, D. B., Karnati, S. K., Bayer, E. A., Lamed, R., Miron, J., Nelson, K. E. & Morrison, M. ( 2004; ). Ruminococcus albus 8 mutants defective in cellulose degradation are deficient in two processive endocellulases, Cel48A and Cel9B, both of which possess a novel modular architecture. J Bacteriol 186, 136–145.[CrossRef]
    [Google Scholar]
  7. Ding, S. Y., Rincon, M. T., Lamed, R., Martin, J. C., McCrae, S. I., Aurilia, V., Shoham, Y., Bayer, E. A. & Flint, H. J. ( 2001; ). Cellulosomal scaffoldin-like proteins from Ruminococcus flavefaciens. J Bacteriol 183, 1945–1953.[CrossRef]
    [Google Scholar]
  8. Gaboriaud, C., Bissery, V., Benchetrit, T. & Mornon, J. P. ( 1987; ). Hydrophobic cluster analysis: an efficient new way to compare and analyse amino acid sequences. FEBS Lett 224, 149–155.[CrossRef]
    [Google Scholar]
  9. Gong, V., Egbosimba, E. E. & Forsberg, C. W. ( 1996; ). Cellulose-binding proteins of Fibrobacter succinogenes and the possible role of a 180-kDa cellulose-binding glycoprotein in adhesion to cellulose. Can J Microbiol 42, 453–460.[CrossRef]
    [Google Scholar]
  10. Graupner, S. & Wackernagel, W. ( 2001; ). Pseudomonas stutzeri has two closely related pilA genes (type IV pilus structural protein) with opposite influences on natural genetic transformation. J Bacteriol 183, 2359–2366.[CrossRef]
    [Google Scholar]
  11. Hames, B. D. ( 1981; ). Recovery of separated proteins. In Gel Electrophoresis of Proteins: a Practical Approach, pp. 61–64. Edited by B. D. Hames & D. Rickwood. Oxford: IRL Press.
  12. Hazes, B., Sastry, P. A., Hayakawa, K., Read, R. J. & Irvin, R. T. ( 2000; ). Crystal structure of Pseudomonas aeruginosa PAK pilin suggests a main-chain-dominated mode of receptor binding. J Mol Biol 299, 1005–1017.[CrossRef]
    [Google Scholar]
  13. Hobbs, M. & Mattick, J. S. ( 1993; ). Common components in the assembly of type 4 fimbriae, DNA transfer systems, filamentous phage and protein-secretion apparatus: a general system for the formation of surface-associated protein complexes. Mol Microbiol 10, 233–243.[CrossRef]
    [Google Scholar]
  14. Hobbs, M., Collie, E. S., Free, P. D., Livingston, S. P. & Mattick, J. S. ( 1993; ). PilS and PilR, a two-component transcriptional regulatory system controlling expression of type 4 fimbriae in Pseudomonas aeruginosa. Mol Microbiol 7, 669–682.[CrossRef]
    [Google Scholar]
  15. Malburg, S. R., Malburg, L. M., Jr, Liu, T., Iyo, A. H. & Forsberg, C. W. ( 1997; ). Catalytic properties of the cellulose-binding endoglucanase F from Fibrobacter succinogenes S85. Appl Environ Microbiol 63, 2449–2453.
    [Google Scholar]
  16. Mattick, J. S. ( 2002; ). Type IV pili and twitching motility. Annu Rev Microbiol 56, 289–314.[CrossRef]
    [Google Scholar]
  17. Miron, J. & Forsberg, C. W. ( 1999; ). Characterisation of cellulose-binding proteins that are involved in the adhesion mechanism of Fibrobacter intestinalis DR7. Appl Microbiol Biotechnol 51, 491–497.[CrossRef]
    [Google Scholar]
  18. Miron, J., Ben-Ghedalia, D. & Morrison, M. ( 2001; ). Invited review: adhesion mechanisms of rumen cellulolytic bacteria. J Dairy Sci 84, 1294–1309.[CrossRef]
    [Google Scholar]
  19. Mitsumori, M. & Minato, H. ( 2000; ). Identification of the cellulose-binding domain of Fibrobacter succinogenes endoglucanase F. FEMS Microbiol Lett 183, 99–103.[CrossRef]
    [Google Scholar]
  20. Morrison, M. & Miron, J. ( 2000; ). Adhesion to cellulose by Ruminococcus albus: a combination of cellulosomes and Pil-proteins? FEMS Microbiol Lett 185, 109–115.[CrossRef]
    [Google Scholar]
  21. Mosoni, P. & Gaillard-Martinie, B. ( 2001; ). Characterization of a spontaneous adhesion-defective mutant of Ruminococcus albus strain 20. Arch Microbiol 176, 52–61.[CrossRef]
    [Google Scholar]
  22. Nunn, D., Bergman, S. & Lory, S. ( 1990; ). Products of three accessory genes, pilB, pilC, and pilD, are required for biogenesis of Pseudomonas aeruginosa pili. J Bacteriol 172, 2911–2919.
    [Google Scholar]
  23. Pegden, R. S., Larson, M. A., Grant, R. J. & Morrison, M. ( 1998; ). Adherence of the gram-positive bacterium Ruminococcus albus to cellulose and identification of a novel form of cellulose-binding protein which belongs to the Pil family of proteins. J Bacteriol 180, 5921–5927.
    [Google Scholar]
  24. Pugsley, A. P. ( 1993; ). The complete general secretory pathway in gram-negative bacteria. Microbiol Rev 57, 50–108.
    [Google Scholar]
  25. Rakotoarivonina, H., Jubelin, G., Hebraud, M., Gaillard-Martinie, B., Forano, E. & Mosoni, P. ( 2002; ). Adhesion to cellulose of the Gram-positive bacterium Ruminococcus albus involves type IV pili. Microbiology 148, 1871–1880.
    [Google Scholar]
  26. Reveneau, C., Adams, S. E., Cotta, M. A. & Morrison, M. ( 2003; ). Phenylacetic and phenylpropionic acids do not affect xylan degradation by Ruminococcus albus. Appl Environ Microbiol 69, 6954–6958.[CrossRef]
    [Google Scholar]
  27. Rincon, M. T., McCrae, S. I., Kirby, J., Scott, K. P. & Flint, H. J. ( 2001; ). EndB, a multidomain family 44 cellulase from Ruminococcus flavefaciens 17, binds to cellulose via a novel cellulose-binding module and to another R. flavefaciens protein via a dockerin domain. Appl Environ Microbiol 67, 4426–4431.[CrossRef]
    [Google Scholar]
  28. Rincon, M. T., Ding, S. Y., McCrae, S. I., Martin, J. C., Aurilia, V., Lamed, R., Shoham, Y., Bayer, E. A. & Flint, H. J. ( 2003; ). Novel organization and divergent dockerin specificities in the cellulosome system of Ruminococcus flavefaciens. J Bacteriol 185, 703–713.[CrossRef]
    [Google Scholar]
  29. Sambrook, J., Fritsch, E. F. & Maniatis, T. ( 1989; ). Molecular Cloning: a Laboratory Manual, 2nd edn. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory.
  30. Villar, M. T., Helber, J. T., Hood, B., Schaefer, M. R. & Hirschberg, R. L. ( 1999; ). Eikenella corrodens phase variation involves a posttranslational event in pilus formation. J Bacteriol 181, 4154–4160.
    [Google Scholar]
  31. Villar, M. T., Hirschberg, R. L. & Schaefer, M. R. ( 2001; ). Role of the Eikenella corrodens pilA locus in pilus function and phase variation. J Bacteriol 183, 55–62.[CrossRef]
    [Google Scholar]
  32. Wolfgang, M., van Putten, J. P., Hayes, S. F., Dorward, D. & Koomey, M. ( 2000; ). Components and dynamics of fiber formation define a ubiquitous biogenesis pathway for bacterial pili. EMBO J 19, 6408–6418.[CrossRef]
    [Google Scholar]
  33. Zheng, D., Alm, E. W., Stahl, D. A. & Raskin, L. ( 1996; ). Characterization of universal small-subunit rRNA hybridization probes for quantitative molecular microbial ecology studies. Appl Environ Microbiol 62, 4504–4513.
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/mic.0.27735-0
Loading
/content/journal/micro/10.1099/mic.0.27735-0
Loading

Data & Media loading...

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error