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Volume 160, Issue 1

Spa13 of has a dual role: chaperone escort and export gate-activator switch of the type III secretion system Free

Abstract

The type III secretion apparatus (T3SA) is used by numerous Gram-negative pathogens to inject virulence factors into eukaryotic cells. The T3SA spans the bacterial envelope and its assembly requires the products of ~20 and genes. Despite progress made in understanding how the T3SA is assembled, the role of several predicted soluble components, such as Spa13, remains elusive. Here, we show that the secretion defect of the mutant is associated with lack of T3SA assembly which is partly due to the instability of the needle component MxiH. In contrast to its counterpart, Spa13 is not a secreted protein. We identified a network of interactions between Spa13 and the ATPase Spa47, the C-ring protein Spa33, and the inner-membrane protein Spa40. Moreover, we revealed a Spa13 interaction with the inner-membrane MxiA and showed that overexpression of the large cytoplasmic domain of MxiA in the WT background shuts off secretion. Lastly, we demonstrated that Spa13 interacts with the cleaved form of Spa40 and with the translocator chaperone IpgC, suggesting that Spa13 intervenes during the secretion hierarchy switch process. Collectively, our results support a dual role of Spa13 as a chaperone escort and as an export gate-activator switch.

  • Received:
  • Accepted:
  • Published Online:
Funding
This study was supported by the:
  • Fonds National de la Recherche Scientifique – Fonds National Belge de la Recherche Scientifique (Award F.3.4556.11)
  • European Community’s Seventh Framework Program FP7/2011-2015 (Award 261742)
  • Fonds National de Recherches Industrielles et Agronomiques
  • Fonds National de Recherches Industrielles et Agronomiques
  • Fonds Defay
  • Alice and David Van Buuren foundation
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2014-01-01
2024-04-25
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References

  1. Abrusci P., Vergara-Irigaray M., Johnson S., Beeby M. D., Hendrixson D. R., Roversi P., Friede M. E., Deane J. E., Jensen G. J. & other authors ( 2013). Architecture of the major component of the type III secretion system export apparatus. Nat Struct Mol Biol 20:99–104 [View Article][PubMed]
     [Google Scholar]
  2. Aizawa S. I. ( 2001). Bacterial flagella and type III secretion systems. FEMS Microbiol Lett 202:157–164 [View Article][PubMed]
     [Google Scholar]
  3. Allaoui A., Mounier J., Prévost M. C., Sansonetti P. J., Parsot C. ( 1992). icsB: a Shigella flexneri virulence gene necessary for the lysis of protrusions during intercellular spread. Mol Microbiol 6:1605–1616 [View Article][PubMed]
     [Google Scholar]
  4. Auvray F., Ozin A. J., Claret L., Hughes C. ( 2002). Intrinsic membrane targeting of the flagellar export ATPase FliI: interaction with acidic phospholipids and FliH. J Mol Biol 318:941–950 [View Article][PubMed]
     [Google Scholar]
  5. Bennett J. C., Hughes C. ( 2000). From flagellum assembly to virulence: the extended family of type III export chaperones. Trends Microbiol 8:202–204 [View Article][PubMed]
     [Google Scholar]
  6. Bergman T., Erickson K., Galyov E., Persson C., Wolf-Watz H. ( 1994). The lcrB (yscN/U) gene cluster of Yersinia pseudotuberculosis is involved in Yop secretion and shows high homology to the spa gene clusters of Shigella flexneri and Salmonella typhimurium . J Bacteriol 176:2619–2626[PubMed]
     [Google Scholar]
  7. Blocker A., Jouihri N., Larquet E., Gounon P., Ebel F., Parsot C., Sansonetti P., Allaoui A. ( 2001). Structure and composition of the Shigella flexneri “needle complex”, a part of its type III secreton. Mol Microbiol 39:652–663 [View Article][PubMed]
     [Google Scholar]
  8. Blocker A., Komoriya K., Aizawa S. ( 2003). Type III secretion systems and bacterial flagella: insights into their function from structural similarities. Proc Natl Acad Sci U S A 100:3027–3030 [View Article][PubMed]
     [Google Scholar]
  9. Botteaux A., Sani M., Kayath C. A., Boekema E. J., Allaoui A. ( 2008). Spa32 interaction with the inner-membrane Spa40 component of the type III secretion system of Shigella flexneri is required for the control of the needle length by a molecular tape measure mechanism. Mol Microbiol 70:1515–1528 [View Article][PubMed]
     [Google Scholar]
  10. Botteaux A., Sory M. P., Biskri L., Parsot C., Allaoui A. ( 2009). MxiC is secreted by and controls the substrate specificity of the Shigella flexneri type III secretion apparatus. Mol Microbiol 71:449–460 [View Article][PubMed]
     [Google Scholar]
  11. Botteaux A., Kayath C. A., Page A. L., Jouihri N., Sani M., Boekema E., Biskri L., Parsot C., Allaoui A. ( 2010). The 33 carboxyl-terminal residues of Spa40 orchestrate the multi-step assembly process of the type III secretion needle complex in Shigella flexneri. . Microbiology 156:2807–2817 [View Article][PubMed]
     [Google Scholar]
  12. Buchrieser C., Glaser P., Rusniok C., Nedjari H., D'Hauteville H., Kunst F., Sansonetti P., Parsot C. ( 2000). The virulence plasmid pWR100 and the repertoire of proteins secreted by the type III secretion apparatus of Shigella flexneri . Mol Microbiol 38:760–771 [View Article][PubMed]
     [Google Scholar]
  13. Cherradi Y., Schiavolin L., Moussa S., Meghraoui A., Meksem A., Biskri L., Azarkan M., Allaoui A., Botteaux A. ( 2013). Interplay between predicted inner-rod and gatekeeper in controlling substrate specificity of the type III secretion system. Mol Microbiol 87:1183–1199 [View Article][PubMed]
     [Google Scholar]
  14. Collazo C. M., Zierler M. K., Gatan J. E. ( 1995). Functional analysis of the Salmonella typhimurium invasion genes invl and invJ and identification of a target of the protein secretion apparatus encoded in the inv locus. Mol Microbiol 15:25–38 [View Article][PubMed]
     [Google Scholar]
  15. Cornelis G. R. ( 2006). The type III secretion injectisome. Nat Rev Microbiol 4:811–825 [View Article][PubMed]
     [Google Scholar]
  16. Diepold A., Wiesand U., Amstutz M., Cornelis G. R. ( 2012). Assembly of the Yersinia injectisome: the missing pieces. Mol Microbiol 85:878–892 [View Article][PubMed]
     [Google Scholar]
  17. Erhardt M., Singer H. M., Wee D. H., Keener J. P., Hughes K. T. ( 2011). An infrequent molecular ruler controls flagellar hook length in Salmonella enterica . EMBO J 30:2948–2961 [View Article][PubMed]
     [Google Scholar]
  18. Espina M., Olive A. J., Kenjale R., Moore D. S., Ausar S. F., Kaminski R. W., Oaks E. V., Middaugh C. R., Picking W. D., Picking W. L. ( 2006). IpaD localizes to the tip of the type III secretion system needle of Shigella flexneri . Infect Immun 74:4391–4400 [View Article][PubMed]
     [Google Scholar]
  19. Evans L. D., Hughes C. ( 2009). Selective binding of virulence type III export chaperones by FliJ escort orthologues InvI and YscO. FEMS Microbiol Lett 293:292–297 [View Article][PubMed]
     [Google Scholar]
  20. Evans L. D., Stafford G. P., Ahmed S., Fraser G. M., Hughes C. ( 2006). An escort mechanism for cycling of export chaperones during flagellum assembly. Proc Natl Acad Sci U S A 103:17474–17479 [View Article][PubMed]
     [Google Scholar]
  21. Fraser G. M., González-Pedrajo B., Tame J. R., Macnab R. M. ( 2003). Interactions of FliJ with the Salmonella type III flagellar export apparatus. J Bacteriol 185:5546–5554 [View Article][PubMed]
     [Google Scholar]
  22. Galán J. E., Wolf-Watz H. ( 2006). Protein delivery into eukaryotic cells by type III secretion machines. Nature 444:567–573 [View Article][PubMed]
     [Google Scholar]
  23. González-Pedrajo B., Minamino T., Kihara M., Namba K. ( 2006). Interactions between C ring proteins and export apparatus components: a possible mechanism for facilitating type III protein export. Mol Microbiol 60:984–998 [View Article][PubMed]
     [Google Scholar]
  24. Herrmann M., Schuhmacher A., Mühldorfer I., Melchers K., Prothmann C., Dammeier S. ( 2006). Identification and characterization of secreted effector proteins of Chlamydophila pneumoniae TW183. Res Microbiol 157:513–524 [View Article][PubMed]
     [Google Scholar]
  25. Hueck C. J. ( 1998). Type III protein secretion systems in bacterial pathogens of animals and plants. Microbiol Mol Biol Rev 62:379–433[PubMed]
     [Google Scholar]
  26. Ibuki T., Imada K., Minamino T., Kato T., Miyata T., Namba K. ( 2011). Common architecture of the flagellar type III protein export apparatus and F- and V-type ATPases. Nat Struct Mol Biol 18:277–282 [View Article][PubMed]
     [Google Scholar]
  27. Ibuki T., Uchida Y., Hironaka Y., Namba K., Imada K., Minamino T. ( 2013). Interaction between FliJ and FlhA, components of the bacterial flagellar type III export apparatus. J Bacteriol 195:466–473 [View Article][PubMed]
     [Google Scholar]
  28. Imada K., Minamino T., Kinoshita M., Furukawa Y., Namba K. ( 2010). Structural insight into the regulatory mechanisms of interactions of the flagellar type III chaperone FliT with its binding partners. Proc Natl Acad Sci U S A 107:8812–8817 [View Article][PubMed]
     [Google Scholar]
  29. Jouihri N., Sory M. P., Page A. L., Gounon P., Parsot C., Allaoui A. ( 2003). MxiK and MxiN interact with the Spa47 ATPase and are required for transit of the needle components MxiH and MxiI, but not of Ipa proteins, through the type III secretion apparatus of Shigella flexneri . Mol Microbiol 49:755–767 [View Article][PubMed]
     [Google Scholar]
  30. Journet L., Agrain C., Broz P., Cornelis G. R. ( 2003). The needle length of bacterial injectisomes is determined by a molecular ruler. Science 302:1757–1760 [View Article][PubMed]
     [Google Scholar]
  31. Kim P. S., Berger B., Wolf E. ( 1997). MultiCoil: a program for predicting two- and three-stranded coiled coils. Protein Sci 6:1179–1189 [View Article][PubMed]
     [Google Scholar]
  32. Kotloff K. L., Winickoff J. P., Ivanoff B., Clemens J. D., Swerdlow D. L., Sansonetti P. J., Adak G. K., Levine M. M. ( 1999). Global burden of Shigella infections: implications for vaccine development and implementation of control strategies. Bull World Health Organ 77:651–666[PubMed]
     [Google Scholar]
  33. Kubori T., Matsushima Y., Nakamura D., Uralil J., Lara-Tejero M., Sukhan A., Galán J. E., Aizawa S. I. ( 1998). Supramolecular structure of the Salmonella typhimurium type III protein secretion system. Science 280:602–605 [View Article][PubMed]
     [Google Scholar]
  34. Lorenzini E., Singer A., Singh B., Lam R., Skarina T., Chirgadze N. Y., Savchenko A., Gupta R. S. ( 2010). Structure and protein–protein interaction studies on Chlamydia trachomatis protein CT670 (YscO homolog). J Bacteriol 192:2746–2756 [View Article][PubMed]
     [Google Scholar]
  35. Macnab R. M. ( 1999). The bacterial flagellum: reversible rotary propellor and type III export apparatus. J Bacteriol 181:7149–7153[PubMed]
     [Google Scholar]
  36. Macnab R. M. ( 2004). Type III flagellar protein export and flagellar assembly. Biochim Biophys Acta 1694:207–217 [View Article][PubMed]
     [Google Scholar]
  37. Magdalena J., Hachani A., Chamekh M., Jouihri N., Gounon P., Blocker A., Allaoui A. ( 2002). Spa32 regulates a switch in substrate specificity of the type III secreton of Shigella flexneri from needle components to Ipa proteins. J Bacteriol 184:3433–3441 [View Article][PubMed]
     [Google Scholar]
  38. McDonnell A. V., Jiang T., Keating A. E., Berger B. ( 2006). Paircoil2: improved prediction of coiled coils from sequence. Bioinformatics 22:356–358 [View Article][PubMed]
     [Google Scholar]
  39. Meitert T., Pencu E., Ciudin L., Tonciu M., Mihai I., Nicolescu S. ( 1991). Correlation between Congo red binding as virulence marker in Shigella species and Sereny test. Roum Arch Microbiol Immunol 50:45–52[PubMed]
     [Google Scholar]
  40. 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[PubMed]
     [Google Scholar]
  41. Minamino T., Macnab R. M. ( 2000). Interactions among components of the Salmonella flagellar export apparatus and its substrates. Mol Microbiol 35:1052–1064 [View Article][PubMed]
     [Google Scholar]
  42. Minamino T., Chu R., Yamaguchi S., Macnab R. M. ( 2000). Role of FliJ in flagellar protein export in Salmonella . J Bacteriol 182:4207–4215 [View Article][PubMed]
     [Google Scholar]
  43. Minamino T., Imada K., Namba K. ( 2008). Mechanisms of type III protein export for bacterial flagellar assembly. Mol Biosyst 4:1105–1115 [View Article][PubMed]
     [Google Scholar]
  44. Minamino T., Morimoto Y. V., Hara N., Namba K. ( 2011). An energy transduction mechanism used in bacterial flagellar type III protein export. Nat Commun 2:475 [View Article][PubMed]
     [Google Scholar]
  45. Morita-Ishihara T., Ogawa M., Sagara H., Yoshida M., Katayama E., Sasakawa C. ( 2006). Shigella Spa33 is an essential C-ring component of type III secretion machinery. J Biol Chem 281:599–607 [View Article][PubMed]
     [Google Scholar]
  46. Mukerjea R., Ghosh P. ( 2013). Functionally essential interaction between Yersinia YscO and the T3S4 domain of YscP. J Bacteriol 195:4631–4638 [View Article][PubMed]
     [Google Scholar]
  47. Parsot C. ( 2009). Shigella type III secretion effectors: how, where, when, for what purposes. Curr Opin Microbiol 12:110–116 [View Article][PubMed]
     [Google Scholar]
  48. Payne P. L., Straley S. C. ( 1998). YscO of Yersinia pestis is a mobile core component of the Yop secretion system. J Bacteriol 180:3882–3890[PubMed]
     [Google Scholar]
  49. Payne P. L., Straley S. C. ( 1999). YscP of Yersinia pestis is a secreted component of the Yop secretion system. J Bacteriol 181:2852–2862[PubMed]
     [Google Scholar]
  50. Penno C., Parsot C. ( 2006). Transcriptional slippage in mxiE controls transcription and translation of the downstream mxiD gene, which encodes a component of the Shigella flexneri type III secretion apparatus. J Bacteriol 188:1191–1198[PubMed] [CrossRef]
     [Google Scholar]
  51. Penno C., Hachani A., Biskri L., Sansonetti P., Allaoui A., Parsot C. ( 2006). Transcriptional slippage controls production of type III secretion apparatus components in Shigella flexneri . Mol Microbiol 62:1460–1468 [View Article][PubMed]
     [Google Scholar]
  52. Riordan K. E., Schneewind O. ( 2008). YscU cleavage and the assembly of Yersinia type III secretion machine complexes. Mol Microbiol 68:1485–1501 [View Article][PubMed]
     [Google Scholar]
  53. Riordan K. E., Sorg J. A., Berube B. J., Schneewind O. ( 2008). Impassable YscP substrates and their impact on the Yersinia enterocolitica type III secretion pathway. J Bacteriol 190:6204–6216 [View Article][PubMed]
     [Google Scholar]
  54. Saijo-Hamano Y., Imada K., Minamino T., Kihara M., Shimada M., Kitao A., Namba K. ( 2010). Structure of the cytoplasmic domain of FlhA and implication for flagellar type III protein export. Mol Microbiol 76:260–268 [View Article][PubMed]
     [Google Scholar]
  55. Samudrala R., Heffron F., McDermott J. E. ( 2009). Accurate prediction of secreted substrates and identification of a conserved putative secretion signal for type III secretion systems. PLoS Pathog 5:e1000375 [View Article][PubMed]
     [Google Scholar]
  56. Sani M., Botteaux A., Parsot C., Sansonetti P., Boekema E. J., Allaoui A. ( 2007). IpaD is localized at the tip of the Shigella flexneri type III secretion apparatus. Biochim Biophys Acta 1770:307–311 [View Article][PubMed]
     [Google Scholar]
  57. Sasakawa C., Komatsu K., Tobe T., Suzuki T., Yoshikawa M. ( 1993). Eight genes in region 5 that form an operon are essential for invasion of epithelial cells by Shigella flexneri 2a. J Bacteriol 175:2334–2346[PubMed]
     [Google Scholar]
  58. Schroeder G. N., Hilbi H. ( 2008). Molecular pathogenesis of Shigella spp.: controlling host cell signaling, invasion, and death by type III secretion. Clin Microbiol Rev 21:134–156 [View Article][PubMed]
     [Google Scholar]
  59. Schuch R., Maurelli A. T. ( 2001). Spa33, a cell surface-associated subunit of the Mxi-Spa type III secretory pathway of Shigella flexneri, regulates Ipa protein traffic. Infect Immun 69:2180–2189 [View Article][PubMed]
     [Google Scholar]
  60. Shibata S., Takahashi N., Chevance F. F., Karlinsey J. E., Hughes K. T., Aizawa S. ( 2007). FliK regulates flagellar hook length as an internal ruler. Mol Microbiol 64:1404–1415 [View Article][PubMed]
     [Google Scholar]
  61. Stone C. B., Johnson D. L., Bulir D. C., Gilchrist J. D., Mahony J. B. ( 2008). Characterization of the putative type III secretion ATPase CdsN (Cpn0707) of Chlamydophila pneumoniae . J Bacteriol 190:6580–6588 [View Article][PubMed]
     [Google Scholar]
  62. Wagner S., Stenta M., Metzger L. C., Dal Peraro M., Cornelis G. R. ( 2010). Length control of the injectisome needle requires only one molecule of Yop secretion protein P (YscP). Proc Natl Acad Sci U S A 107:13860–13865 [View Article][PubMed]
     [Google Scholar]
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Spa13 of Shigella flexneri has a dual role: chaperone escort and export gate-activator switch of the type III secretion system
Microbiology 160, 130 (2014); https://doi.org/10.1099/mic.0.071712-0
/content/journal/micro/10.1099/mic.0.071712-0
/content/journal/micro/10.1099/mic.0.071712-0
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