1887

Abstract

is the most common bacterial infection of the human reproductive tract globally; however, the mechanisms underlying the adaptation of the organism to its natural target cells, human endocervical epithelial cells, are not clearly understood. To secure its intracellular niche, must modulate the host cellular machinery by secreting virulence factors into the host cytosol to facilitate bacterial growth and survival. Here we used primary human endocervical epithelial cells and HeLa cells infected with to examine the secretion of bacterial proteins during productive growth and persistent growth induced by ampicillin. Specifically, we observed a decrease in secretable chlamydial protease-like activity factor (CPAF) in the cytosol of host epithelial cells exposed to ampicillin with no evident reduction of CPAF product by . In contrast, the expression of CopN and Tarp was downregulated, suggesting that responds to ampicillin exposure by selectively altering the expression of secretable proteins. In addition, we observed a greater accumulation of outer-membrane vesicles from in persistently infected cells. Taken together, these results suggest that the regulation of both gene expression and the secretion of chlamydial virulence proteins is involved in the adaptation of the bacteria to a persistent infection state in human genital epithelial cells.

Funding
This study was supported by the:
  • Louisiana Board of Regents
  • NIH (Award U19AI061972 and AI055869)
Loading

Article metrics loading...

/content/journal/micro/10.1099/mic.0.044917-0
2011-10-01
2024-03-28
Loading full text...

Full text loading...

/deliver/fulltext/micro/157/10/2759.html?itemId=/content/journal/micro/10.1099/mic.0.044917-0&mimeType=html&fmt=ahah

References

  1. Amano A., Takeuchi H., Furuta N. ( 2010). Outer membrane vesicles function as offensive weapons in host-parasite interactions. Microbes Infect 12:791–798 [View Article][PubMed]
    [Google Scholar]
  2. Bannantine J. P., Stamm W. E., Suchland R. J., Rockey D. D. ( 1998). Chlamydia trachomatis IncA is localized to the inclusion membrane and is recognized by antisera from infected humans and primates. Infect Immun 66:6017–6021[PubMed]
    [Google Scholar]
  3. Beatty W. L. ( 2006). Trafficking from CD63-positive late endocytic multivesicular bodies is essential for intracellular development of Chlamydia trachomatis. J Cell Sci 119:350–359 [View Article][PubMed]
    [Google Scholar]
  4. Beatty W. L., Morrison R. P., Byrne G. I. ( 1994). Persistent chlamydiae: from cell culture to a paradigm for chlamydial pathogenesis. Microbiol Rev 58:686–699[PubMed]
    [Google Scholar]
  5. Belland R. J., Nelson D. E., Virok D., Crane D. D., Hogan D., Sturdevant D., Beatty W. L., Caldwell H. D. ( 2003). Transcriptome analysis of chlamydial growth during IFN-gamma-mediated persistence and reactivation. Proc Natl Acad Sci U S A 100:15971–15976 [View Article][PubMed]
    [Google Scholar]
  6. Betts H. J., Wolf K., Fields K. A. ( 2009). Effector protein modulation of host cells: examples in the Chlamydia spp. arsenal. Curr Opin Microbiol 12:81–87 [View Article][PubMed]
    [Google Scholar]
  7. Brunham R. C., Rey-Ladino J. ( 2005). Immunology of Chlamydia infection: implications for a Chlamydia trachomatis vaccine. Nat Rev Immunol 5:149–161 [View Article][PubMed]
    [Google Scholar]
  8. Chen D., Lei L., Lu C., Flores R., DeLisa M. P., Roberts T. C., Romesberg F. E., Zhong G. ( 2010). Secretion of the chlamydial virulence factor CPAF requires the Sec-dependent pathway. Microbiology 156:3031–3040 [View Article][PubMed]
    [Google Scholar]
  9. Clifton D. R., Fields K. A., Grieshaber S. S., Dooley C. A., Fischer E. R., Mead D. J., Carabeo R. A., Hackstadt T. ( 2004). A chlamydial type III translocated protein is tyrosine-phosphorylated at the site of entry and associated with recruitment of actin. Proc Natl Acad Sci U S A 101:10166–10171 [View Article][PubMed]
    [Google Scholar]
  10. Cocchiaro J. L., Kumar Y., Fischer E. R., Hackstadt T., Valdivia R. H. ( 2008). Cytoplasmic lipid droplets are translocated into the lumen of the Chlamydia trachomatis parasitophorous vacuole. Proc Natl Acad Sci U S A 105:9379–9384 [View Article][PubMed]
    [Google Scholar]
  11. Deka S., Vanover J., Dessus-Babus S., Whittimore J., Howett M. K., Wyrick P. B., Schoborg R. V. ( 2006). Chlamydia trachomatis enters a viable but non-cultivable (persistent) state within herpes simplex virus type 2 (HSV-2) co-infected host cells. Cell Microbiol 8:149–162 [View Article][PubMed]
    [Google Scholar]
  12. Ellis T. N., Kuehn M. J. ( 2010). Virulence and immunomodulatory roles of bacterial outer membrane vesicles. Microbiol Mol Biol Rev 74:81–94 [View Article][PubMed]
    [Google Scholar]
  13. Fichorova R. N., Rheinwald J. G., Anderson D. J. ( 1997). Generation of papillomavirus-immortalized cell lines from normal human ectocervical, endocervical, and vaginal epithelium that maintain expression of tissue-specific differentiation proteins. Biol Reprod 57:847–855 [View Article][PubMed]
    [Google Scholar]
  14. Fields K. A., Hackstadt T. ( 2000). Evidence for the secretion of Chlamydia trachomatis CopN by a type III secretion mechanism. Mol Microbiol 38:1048–1060 [View Article][PubMed]
    [Google Scholar]
  15. Giles D. K., Wyrick P. B. ( 2008). Trafficking of chlamydial antigens to the endoplasmic reticulum of infected epithelial cells. Microbes Infect 10:1494–1503 [View Article][PubMed]
    [Google Scholar]
  16. Giles D. K., Whittimore J. D., LaRue R. W., Raulston J. E., Wyrick P. B. ( 2006). Ultrastructural analysis of chlamydial antigen-containing vesicles everting from the Chlamydia trachomatis inclusion. Microbes Infect 8:1579–1591 [View Article][PubMed]
    [Google Scholar]
  17. Gottlieb R. A., Adachi S. ( 2000). Nitrogen cavitation for cell disruption to obtain mitochondria from cultured cells. Methods Enzymol 322:213–221 [View Article][PubMed]
    [Google Scholar]
  18. Gottlieb S. L., Brunham R. C., Byrne G. I., Martin D. H., Xu F., Berman S. M. ( 2010). Introduction: The natural history and immunobiology of Chlamydia trachomatis genital infection and implications for Chlamydia control. J Infect Dis 201:Suppl. 2S85–S87 [View Article][PubMed]
    [Google Scholar]
  19. Guseva N. V., Dessus-Babus S. C., Whittimore J. D., Moore C. G., Wyrick P. B. ( 2005). Characterization of estrogen-responsive epithelial cell lines and their infectivity by genital Chlamydia trachomatis. Microbes Infect 7:1469–1481 [View Article][PubMed]
    [Google Scholar]
  20. Harper A., Pogson C. I., Jones M. L., Pearce J. H. ( 2000). Chlamydial development is adversely affected by minor changes in amino acid supply, blood plasma amino acid levels, and glucose deprivation. Infect Immun 68:1457–1464 [View Article][PubMed]
    [Google Scholar]
  21. Hatch T. P. ( 1999). Developmental biology. Chlamydia. Intracellular Biology, Pathogenesis, and Immunity Stephens R. S. 29–67 Washington, DC: American Society for Microbiology;
    [Google Scholar]
  22. Herbst-Kralovetz M. M., Quayle A. J., Ficarra M., Greene S., Rose W. A. II, Chesson R., Spagnuolo R. A., Pyles R. B. ( 2008). Quantification and comparison of Toll-like receptor expression and responsiveness in primary and immortalized human female lower genital tract epithelia. Am J Reprod Immunol 59:212–224 [View Article][PubMed]
    [Google Scholar]
  23. Heuer D., Brinkmann V., Meyer T. F., Szczepek A. J. ( 2003). Expression and translocation of chlamydial protease during acute and persistent infection of the epithelial HEp-2 cells with Chlamydophila (Chlamydia) pneumoniae. Cell Microbiol 5:315–322 [View Article][PubMed]
    [Google Scholar]
  24. Hogan R. J., Mathews S. A., Mukhopadhyay S., Summersgill J. T., Timms P. ( 2004). Chlamydial persistence: beyond the biphasic paradigm. Infect Immun 72:1843–1855 [View Article][PubMed]
    [Google Scholar]
  25. Hower S., Wolf K., Fields K. A. ( 2009). Evidence that CT694 is a novel Chlamydia trachomatis T3S substrate capable of functioning during invasion or early cycle development. Mol Microbiol 72:1423–1437 [View Article][PubMed]
    [Google Scholar]
  26. Hua Z., Rao X., Feng X., Luo X., Liang Y., Shen L. ( 2009). Mutagenesis of region 4 of sigma 28 from Chlamydia trachomatis defines determinants for protein-protein and protein-DNA interactions. J Bacteriol 191:651–660 [View Article][PubMed]
    [Google Scholar]
  27. Huang J., Lesser C. F., Lory S. ( 2008). The essential role of the CopN protein in Chlamydia pneumoniae intracellular growth. Nature 456:112–115 [View Article][PubMed]
    [Google Scholar]
  28. Hybiske K., Stephens R. S. ( 2007). Mechanisms of host cell exit by the intracellular bacterium Chlamydia. Proc Natl Acad Sci U S A 104:11430–11435 [View Article][PubMed]
    [Google Scholar]
  29. Kadurugamuwa J. L., Beveridge T. J. ( 1995). Virulence factors are released from Pseudomonas aeruginosa in association with membrane vesicles during normal growth and exposure to gentamicin: a novel mechanism of enzyme secretion. J Bacteriol 177:3998–4008[PubMed]
    [Google Scholar]
  30. Kawana K., Quayle A. J., Ficarra M., Ibana J. A., Shen L., Kawana Y., Yang H., Marrero L., Yavagal S. et al. ( 2007). CD1d degradation in Chlamydia trachomatis-infected epithelial cells is the result of both cellular and chlamydial proteasomal activity. J Biol Chem 282:7368–7375 [View Article][PubMed]
    [Google Scholar]
  31. Klos A., Thalmann J., Peters J., Gérard H. C., Hudson A. P. ( 2009). The transcript profile of persistent Chlamydophila (Chlamydia) pneumoniae in vitro depends on the means by which persistence is induced. FEMS Microbiol Lett 291:120–126 [View Article][PubMed]
    [Google Scholar]
  32. Lambden P. R., Pickett M. A., Clarke I. N. ( 2006). The effect of penicillin on Chlamydia trachomatis DNA replication. Microbiology 152:2573–2578 [View Article][PubMed]
    [Google Scholar]
  33. Loeb M. R. ( 1974). Bacteriophage T4-mediated release of envelope components from Escherichia coli. J Virol 13:631–641[PubMed]
    [Google Scholar]
  34. Matsumoto A., Manire G. P. ( 1970). Electron microscopic observations on the effects of penicillin on the morphology of Chlamydia psittaci. J Bacteriol 101:278–285[PubMed]
    [Google Scholar]
  35. McBroom A. J., Johnson A. P., Vemulapalli S., Kuehn M. J. ( 2006). Outer membrane vesicle production by Escherichia coli is independent of membrane instability. J Bacteriol 188:5385–5392 [View Article][PubMed]
    [Google Scholar]
  36. Molano M., Meijer C. J., Weiderpass E., Arslan A., Posso H., Franceschi S., Ronderos M., Muñoz N., van den Brule A. J. ( 2005). The natural course of Chlamydia trachomatis infection in asymptomatic Colombian women: a 5-year follow-up study. J Infect Dis 191:907–916 [View Article][PubMed]
    [Google Scholar]
  37. Moorman D. R., Sixbey J. W., Wyrick P. B. ( 1986). Interaction of Chlamydia trachomatis with human genital epithelium in culture. J Gen Microbiol 132:1055–1067[PubMed]
    [Google Scholar]
  38. Moulder J. W. ( 1991). Interaction of chlamydiae and host cells in vitro. Microbiol Rev 55:143–190[PubMed]
    [Google Scholar]
  39. Ouellette S. P., Hatch T. P., AbdelRahman Y. M., Rose L. A., Belland R. J., Byrne G. I. ( 2006). Global transcriptional upregulation in the absence of increased translation in Chlamydia during IFNγ-mediated host cell tryptophan starvation. Mol Microbiol 62:1387–1401 [View Article][PubMed]
    [Google Scholar]
  40. Rao X., Deighan P., Hua Z., Hu X., Wang J., Luo M., Wang J., Liang Y., Zhong G. et al. ( 2009). A regulator from Chlamydia trachomatis modulates the activity of RNA polymerase through direct interaction with the beta subunit and the primary sigma subunit. Genes Dev 23:1818–1829 [View Article][PubMed]
    [Google Scholar]
  41. Rasmussen S. J., Eckmann L., Quayle A. J., Shen L., Zhang Y. X., Anderson D. J., Fierer J., Stephens R. S., Kagnoff M. F. ( 1997). Secretion of proinflammatory cytokines by epithelial cells in response to Chlamydia infection suggests a central role for epithelial cells in chlamydial pathogenesis. J Clin Invest 99:77–87 [View Article][PubMed]
    [Google Scholar]
  42. Raulston J. E. ( 1997). Response of Chlamydia trachomatis serovar E to iron restriction in vitro and evidence for iron-regulated chlamydial proteins. Infect Immun 65:4539–4547[PubMed]
    [Google Scholar]
  43. Reveneau N., Crane D. D., Fischer E., Caldwell H. D. ( 2005). Bactericidal activity of first-choice antibiotics against gamma interferon-induced persistent infection of human epithelial cells by Chlamydia trachomatis. Antimicrob Agents Chemother 49:1787–1793 [View Article][PubMed]
    [Google Scholar]
  44. Rockey D. D., Scidmore M. A., Bannantine J. P., Brown W. J. ( 2002). Proteins in the chlamydial inclusion membrane. Microbes Infect 4:333–340 [View Article][PubMed]
    [Google Scholar]
  45. Roth A., König P., van Zandbergen G., Klinger M., Hellwig-Bürgel T., Däubener W., Bohlmann M. K., Rupp J. ( 2010). Hypoxia abrogates antichlamydial properties of IFN-γ in human fallopian tube cells in vitro and ex vivo. Proc Natl Acad Sci U S A 107:19502–19507 [View Article][PubMed]
    [Google Scholar]
  46. Shaw A. C., Vandahl B. B., Larsen M. R., Roepstorff P., Gevaert K., Vandekerckhove J., Christiansen G., Birkelund S. ( 2002). Characterization of a secreted Chlamydia protease. Cell Microbiol 4:411–424 [View Article][PubMed]
    [Google Scholar]
  47. Shen L., Shi Y., Douglas A. L., Hatch T. P., O’Connell C. M. C., Chen J.-M., Zhang Y.-X. ( 2000). Identification and characterization of promoters regulating tuf expression in Chlamydia trachomatis serovar F. Arch Biochem Biophys 379:46–56 [View Article][PubMed]
    [Google Scholar]
  48. Shen L., Li M., Zhang Y. X. ( 2004). Chlamydia trachomatis sigma28 recognizes the fliC promoter of Escherichia coli and responds to heat shock in chlamydiae. Microbiology 150:205–215 [View Article][PubMed]
    [Google Scholar]
  49. Stirling P., Richmond S. J. ( 1980). Production of outer membrane blebs during chlamydial replication. FEMS Microbiol Lett 9:103–105 [View Article]
    [Google Scholar]
  50. Ünal C. M., Schaar V., Riesbeck K. ( 2010). Bacterial outer membrane vesicles in disease and preventive medicine. Semin Immunopathol1–14[PubMed]
    [Google Scholar]
  51. Wang Y., Berg E. A., Feng X., Shen L., Smith T., Costello C. E., Zhang Y.-X. ( 2006). Identification of surface-exposed components of MOMP of Chlamydia trachomatis serovar F. Protein Sci 15:122–134 [View Article][PubMed]
    [Google Scholar]
  52. Wolf K., Fischer E., Hackstadt T. ( 2000). Ultrastructural analysis of developmental events in Chlamydia pneumoniae-infected cells. Infect Immun 68:2379–2385 [View Article][PubMed]
    [Google Scholar]
  53. Wyrick P. ( 2006). Polarized epithelial cell culture for Chlamydia trachomatis Wymondham, Norfolk, UK: Horizon Bioscience;
    [Google Scholar]
  54. Wyrick P. B. ( 2010). Chlamydia trachomatis persistence in vitro – an overview. J Infect Dis 201:Suppl. 2S88–S95 [View Article]
    [Google Scholar]
  55. Wyrick P. B., Knight S. T. ( 2004). Pre-exposure of infected human endometrial epithelial cells to penicillin in vitro renders Chlamydia trachomatis refractory to azithromycin. J Antimicrob Chemother 54:79–85 [View Article][PubMed]
    [Google Scholar]
  56. Zhong G., Fan T., Liu L. ( 1999). Chlamydia inhibits interferon gamma-inducible major histocompatibility complex class II expression by degradation of upstream stimulatory factor 1. J Exp Med 189:1931–1938 [View Article][PubMed]
    [Google Scholar]
  57. Zhong G., Liu L., Fan T., Fan P., Ji H. ( 2000). Degradation of transcription factor RFX5 during the inhibition of both constitutive and interferon gamma-inducible major histocompatibility complex class I expression in Chlamydia-infected cells. J Exp Med 191:1525–1534 [View Article][PubMed]
    [Google Scholar]
  58. Zhong G., Fan P., Ji H., Dong F., Huang Y. ( 2001). Identification of a chlamydial protease-like activity factor responsible for the degradation of host transcription factors. J Exp Med 193:935–942 [View Article][PubMed]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/mic.0.044917-0
Loading
/content/journal/micro/10.1099/mic.0.044917-0
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