1887

Abstract

Surface proteins of probiotic microbes, including and , are believed to promote retention in the gut and mediate host–bacterial communications. Sortase, an enzyme that covalently couples a subset of extracellular proteins containing an LPXTG motif to the cell surface, is of particular interest in characterizing bacterial adherence and communication with the mucosal immune system. A sortase gene, , was identified in NCFM (LBA1244) and ATCC 33323 (LGAS_0825). Additionally, eight and six intact sortase-dependent proteins were predicted in and , respectively. Due to the role of sortase in coupling these proteins to the cell wall, Δ deletion mutants of and were created using the -based counterselective gene replacement system. Inactivation of sortase did not cause significant alteration in growth or survival in simulated gastrointestinal juices. Meanwhile, both Δ mutants showed decreased adhesion to porcine mucin Murine dendritic cells exposed to the Δ mutant of or induced lower levels of pro-inflammatory cytokines TNF-α and IL-12, respectively, compared with the parent strains. co-colonization of the Δ mutant and its parent strain in germ-free 129S6/SvEv mice resulted in a significant one-log reduction of the Δ mutant population. Additionally, a similar reduction of the Δ mutant was observed in the caecum. This study shows for the first time that sortase-dependent proteins contribute to gut retention of probiotic microbes in the gastrointestinal tract.

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2015-02-01
2024-12-14
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References

  1. Altermann E., Russell W. M., Azcarate-Peril M. A., Barrangou R., Buck B. L., McAuliffe O., Souther N., Dobson A., Duong T.& other authors ( 2005; Complete genome sequence of the probiotic lactic acid bacterium Lactobacillus acidophilus NCFM. Proc Natl Acad Sci U S A 102:3906–3912 [View Article][PubMed]
    [Google Scholar]
  2. Azcarate-Peril M. A., Altermann E., Goh Y. J., Tallon R., Sanozky-Dawes R. B., Pfeiler E. A., O’Flaherty S., Buck B. L., Dobson A.& other authors ( 2008; Analysis of the genome sequence of Lactobacillus gasseri ATCC 33323 reveals the molecular basis of an autochthonous intestinal organism. Appl Environ Microbiol 74:4610–4625 [View Article][PubMed]
    [Google Scholar]
  3. Barefoot S. F., Klaenhammer T. R. 1983; Detection and activity of lactacin B, a bacteriocin produced by Lactobacillus acidophilus.. Appl Environ Microbiol 45:1808–1815[PubMed]
    [Google Scholar]
  4. Bierne H., Mazmanian S. K., Trost M., Pucciarelli M. G., Liu G., Dehoux P., Jänsch L., Garcia-del Portillo F., Schneewind O., Cossart P.European Listeria Genome Consortium 2002; Inactivation of the srtA gene in Listeria monocytogenes inhibits anchoring of surface proteins and affects virulence. Mol Microbiol 43:869–881 [View Article][PubMed]
    [Google Scholar]
  5. Boot H. J., Kolen C. P., Pot B., Kersters K., Pouwels P. H. 1996; The presence of two S-layer-protein-encoding genes is conserved among species related to Lactobacillus acidophilus.. Microbiology 142:2375–2384 [View Article][PubMed]
    [Google Scholar]
  6. Bron P. A., van Baarlen P., Kleerebezem M. 2012; Emerging molecular insights into the interaction between probiotics and the host intestinal mucosa. Nat Rev Microbiol 10:66–78[PubMed]
    [Google Scholar]
  7. Bu X. D., Li N., Tian X. Q., Huang P. L. 2011; Caco-2 and LS174T cell lines provide different models for studying mucin expression in colon cancer. Tissue Cell 43:201–206 [View Article][PubMed]
    [Google Scholar]
  8. Call E. K., Klaenhammer T. R. 2013; Relevance and application of sortase and sortase-dependent proteins in lactic acid bacteria. Front Microbiol 4:73 [View Article][PubMed]
    [Google Scholar]
  9. Charteris W. P., Kelly P. M., Morelli L., Collins J. K. 1998; Development and application of an in vitro methodology to determine the transit tolerance of potentially probiotic Lactobacillus and Bifidobacterium species in the upper human gastrointestinal tract. J Appl Microbiol 84:759–768 [View Article][PubMed]
    [Google Scholar]
  10. Dieye Y., Oxaran V., Ledue-Clier F., Alkhalaf W., Buist G., Juillard V., Lee C. W., Piard J. C. 2010; Functionality of sortase A in Lactococcus lactis.. Appl Environ Microbiol 76:7332–7337 [View Article][PubMed]
    [Google Scholar]
  11. Frece J., Kos B., Svetec I. K., Zgaga Z., Mrsa V., Susković J. 2005; Importance of S-layer proteins in probiotic activity of Lactobacillus acidophilus M92. J Appl Microbiol 98:285–292 [View Article][PubMed]
    [Google Scholar]
  12. Goh Y. J., Klaenhammer T. R. 2010; Functional roles of aggregation-promoting-like factor in stress tolerance and adherence of Lactobacillus acidophilus NCFM. Appl Environ Microbiol 76:5005–5012 [View Article][PubMed]
    [Google Scholar]
  13. Goh Y. J., Klaenhammer T. R. 2014; Insights into glycogen metabolism in Lactobacillus acidophilus: impact on carbohydrate metabolism, stress tolerance and gut retention. Microb Cell Fact 13:94 [View Article][PubMed]
    [Google Scholar]
  14. Goh Y. J., Azcárate-Peril M. A., O’Flaherty S., Durmaz E., Valence F., Jardin J., Lortal S., Klaenhammer T. R. 2009; Development and application of a upp-based counterselective gene replacement system for the study of the S-layer protein SlpX of Lactobacillus acidophilus NCFM. Appl Environ Microbiol 75:3093–3105 [View Article][PubMed]
    [Google Scholar]
  15. Hanahan D. 1985; Techniques for transformation of E. coli, p. 109–135. In Glover D. M. DNA cloning: a practical approach, vol. 1. IRL Press Ltd.; Oxford, England:
    [Google Scholar]
  16. Hendrickx A. P., Budzik J. M., Oh S. Y., Schneewind O. 2011; Architects at the bacterial surface - sortases and the assembly of pili with isopeptide bonds. Nat Rev Microbiol 9:166–176 [View Article][PubMed]
    [Google Scholar]
  17. Horton R. M., Hunt H. D., Ho S. N., Pullen J. K., Pease L. R. 1989; Engineering hybrid genes without the use of restriction enzymes: gene splicing by overlap extension. Gene 77:61–68 [View Article][PubMed]
    [Google Scholar]
  18. Khazaie K., Zadeh M., Khan M. W., Bere P., Gounari F., Dennis K., Blatner N. R., Owen J. L., Klaenhammer T. R., Mohamadzadeh M. 2012; Abating colon cancer polyposis by Lactobacillus acidophilus deficient in lipoteichoic acid. Proc Natl Acad Sci U S A 109:10462–10467 [View Article][PubMed]
    [Google Scholar]
  19. Kimmel S. A., Roberts R. F. 1998; Development of a growth medium suitable for exopolysaccharide production by Lactobacillus delbrueckii ssp. bulgaricus RR. Int J Food Microbiol 40:87–92 [View Article][PubMed]
    [Google Scholar]
  20. Kleerebezem M., Hols P., Bernard E., Rolain T., Zhou M., Siezen R. J., Bron P. A. 2010; The extracellular biology of the lactobacilli. FEMS Microbiol Rev 34:199–230 [View Article][PubMed]
    [Google Scholar]
  21. Konstantinov S. R., Smidt H., de Vos W. M., Bruijns S. C., Singh S. K., Valence F., Molle D., Lortal S., Altermann E.& other authors ( 2008; S layer protein A of Lactobacillus acidophilus NCFM regulates immature dendritic cell and T cell functions. Proc Natl Acad Sci U S A 105:19474–19479 [View Article][PubMed]
    [Google Scholar]
  22. Law J., Buist G., Haandrikman A., Kok J., Venema G., Leenhouts K. 1995; A system to generate chromosomal mutations in Lactococcus lactis which allows fast analysis of targeted genes. J Bacteriol 177:7011–7018[PubMed]
    [Google Scholar]
  23. Maresso A. W., Schneewind O. 2008; Sortase as a target of anti-infective therapy. Pharmacol Rev 60:128–141 [View Article][PubMed]
    [Google Scholar]
  24. Marraffini L. A., Dedent A. C., Schneewind O. 2006; Sortases and the art of anchoring proteins to the envelopes of gram-positive bacteria. Microbiol Mol Biol Rev 70:192–221 [View Article][PubMed]
    [Google Scholar]
  25. Mazmanian S. K., Liu G., Ton-That H., Schneewind O. 1999; Staphylococcus aureus sortase, an enzyme that anchors surface proteins to the cell wall. Science 285:760–763 [View Article][PubMed]
    [Google Scholar]
  26. Muñoz-Provencio D., Rodríguez-Díaz J., Collado M. C., Langella P., Bermúdez-Humarán L. G., Monedero V. 2012; Functional analysis of the Lactobacillus casei BL23 sortases. Appl Environ Microbiol 78:8684–8693 [View Article][PubMed]
    [Google Scholar]
  27. O’Callaghan J., Buttó L. F., MacSharry J., Nally K., O’Toole P. W. 2012; Influence of adhesion and bacteriocin production by Lactobacillus salivarius on the intestinal epithelial cell transcriptional response. Appl Environ Microbiol 78:5196–5203 [View Article][PubMed]
    [Google Scholar]
  28. O'Flaherty S., Klaenhammer T. R. 2010; The role and potential of probiotic bacteria in the gut, and the communication between gut microflora and gut/host. Int Dairy J 20:262–268 [View Article]
    [Google Scholar]
  29. Paterson G. K., Mitchell T. J. 2006; The role of Streptococcus pneumoniae sortase A in colonisation and pathogenesis. Microbes Infect 8:145–153 [View Article][PubMed]
    [Google Scholar]
  30. Petersen T. N., Brunak S., von Heijne G., Nielsen H. 2011; SignalP 4.0: discriminating signal peptides from transmembrane regions. Nat Methods 8:785–786 [View Article][PubMed]
    [Google Scholar]
  31. Remus D. M., Bongers R. S., Meijerink M., Fusetti F., Poolman B., de Vos P., Wells J. M., Kleerebezem M., Bron P. A. 2013; Impact of Lactobacillus plantarum sortase on target protein sorting, gastrointestinal persistence, and host immune response modulation. J Bacteriol 195:502–509 [View Article][PubMed]
    [Google Scholar]
  32. Russell W. M., Klaenhammer T. R. 2001; Efficient system for directed integration into the Lactobacillus acidophilus and Lactobacillus gasseri chromosomes via homologous recombination. Appl Environ Microbiol 67:4361–4364 [View Article][PubMed]
    [Google Scholar]
  33. Rutherford K., Parkhill J., Crook J., Horsnell T., Rice P., Rajandream M. A., Barrell B. 2000; Artemis: sequence visualization and annotation. Bioinformatics 16:944–945 [View Article][PubMed]
    [Google Scholar]
  34. Selle K., Goh Y. J., O’Flaherty S., Klaenhammer T. R. 2014; Development of an integration mutagenesis system in Lactobacillus gasseri.. Gut Microbes 5:326–332 [View Article][PubMed]
    [Google Scholar]
  35. Spirig T., Weiner E. M., Clubb R. T. 2011; Sortase enzymes in Gram-positive bacteria. Mol Microbiol 82:1044–1059 [View Article][PubMed]
    [Google Scholar]
  36. van Pijkeren J. P., Canchaya C., Ryan K. A., Li Y., Claesson M. J., Sheil B., Steidler L., O’Mahony L., Fitzgerald G. F.& other authors ( 2006; Comparative and functional analysis of sortase-dependent proteins in the predicted secretome of Lactobacillus salivarius UCC118. Appl Environ Microbiol 72:4143–4153 [View Article][PubMed]
    [Google Scholar]
  37. Walker D. C., Aoyama K., Klaenhammer T. R. 1996; Electrotransformation of lactobacillus acidophilus group A1. FEMS Microbiol Lett 138:233–237 [View Article][PubMed]
    [Google Scholar]
  38. Zhou M., Theunissen D., Wels M., Siezen R. J. 2010; LAB-Secretome: a genome-scale comparative analysis of the predicted extracellular and surface-associated proteins of Lactic Acid Bacteria. BMC Genomics 11:651 [View Article][PubMed]
    [Google Scholar]
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