1887

Abstract

SseA, a key virulence determinant, is a small, basic pI protein encoded within the pathogenicity island 2 and serves as a type III secretion system chaperone for SseB and SseD. Both SseA partners are subunits of the surface-localized translocon module that delivers effectors into the host cell; SseB is predicted to compose the translocon sheath and SseD is a putative translocon pore subunit. In this study, SseA molecular interactions with its partners were characterized further. Yeast two-hybrid screens indicate that SseA binding requires a C-terminal domain within both partners. An additional central domain within SseD was found to influence binding. The SseA-binding region within SseB was found to encompass a predicted amphipathic helix of a type participating in coiled-coil interactions that are implicated in the assembly of translocon sheaths. Deletions that impinge upon this putative coiled-coiled domain prevent SseA binding, suggesting that SseA occupies a portion of the coiled-coil. SseA occupancy of this motif is envisioned to be sufficient to prevent premature SseB self-association inside bacteria. Domain mapping on the chaperone was also performed. A deletion of the SseA N-terminus, or site-directed mutations within this region, allowed stabilization of SseB, but its export was disrupted. Therefore, the N-terminus of SseA provides a function that is essential for SseB export, but dispensable for partner binding and stabilization.

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2004-07-01
2020-08-06
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References

  1. Adams A., Gottschling D. E., Kaiser C. A., Stearns T.. 1997; Methods in Yeast Genetics Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
  2. Auvray F., Thomas J., Fraser G. M., Hughes C.. 2001; Flagellin polymerization control by a cytosolic export chaperone. J Mol Biol308:221–229[CrossRef]
    [Google Scholar]
  3. Bennett J. C. Q., Hughes C.. 2000; From flagellum assembly to virulence: the extended family of type III export chaperones. Trends Microbiol8:202–204[CrossRef]
    [Google Scholar]
  4. Beuzón C. R., Banks G., Deiwick J., Hensel M., Holden D. W.. 1999; pH-dependent secretion of SseB, a product of the SPI-2 type III secretion system of Salmonella typhimurium. Mol Microbiol33:806–816[CrossRef]
    [Google Scholar]
  5. Cheng L. W., Schneewind O.. 1999; Yersinia enterocolitica type III secretion. On the role of SycE in targeting YopE into HeLa cells. J Biol Chem274:22102–22108[CrossRef]
    [Google Scholar]
  6. Coombes B. K., Brown N. F., Kujat-Choy S., Vallance B. A., Finlay B. B.. 2003; SseA is required for translocation of Salmonella pathogenicity island-2 effectors into host cells. Microbes Infect5:561–570[CrossRef]
    [Google Scholar]
  7. Cornelis G. R.. 2002; The Yersinia YSC-Yop ‘type III’ weaponry. Nat Rev Mol Cell Biol3:742–752[CrossRef]
    [Google Scholar]
  8. Cornelis G. R., Van Gijsegem F.. 2000; Assembly and function of type III secretory systems. Annu Rev Microbiol54:735–774[CrossRef]
    [Google Scholar]
  9. Creasey E. A., Friedberg D., Shaw R. K., Umanski T., Knutton S., Rosenshine I., Frankel G.. 2003; CesAB is an enteropathogenic Escherichia coli chaperone for the type-III translocator proteins EspA and EspB. Microbiology149:3639–3647[CrossRef]
    [Google Scholar]
  10. Daniell S. J., Takahashi N., Wilson R..7 other authors 2001; The filamentous type III secretion translocon of enteropathogenic Escherichia coli. Cell Microbiol3:865–871[CrossRef]
    [Google Scholar]
  11. Daniell S. J., Kocsis E., Morris E., Knutton S., Booy F. P., Frankel G.. 2003; 3D structure of EspA filaments from enteropathogenic Escherichia coli. Mol Microbiol49:301–308[CrossRef]
    [Google Scholar]
  12. Davis C. R., Richman T. J., Deliduka S. B., Blaisdell J. O., Collins C. C., Johnson D. I.. 1998; Analysis of the mechanisms of action of the Saccharomyces cerevisiae dominant lethal cdc42G12V and dominant negative cdc42D118A mutations. J Biol Chem273:849–858[CrossRef]
    [Google Scholar]
  13. Delahay R. M., Frankel G.. 2002; Coiled-coil proteins associated with type III secretion systems: a versatile domain revisited. Mol Microbiol45:905–916[CrossRef]
    [Google Scholar]
  14. Delahay R. M., Knutton S., Shaw R. K., Hartland E. L., Pallen M. J., Frankel G.. 1999; The coiled-coil domain of EspA is essential for the assembly of the type III secretion translocon on the surface of enteropathogenic Escherichia coli. J Biol Chem274:35969–35974[CrossRef]
    [Google Scholar]
  15. Elliott S. J., Wainwright L. A., McDaniel T. K., Jarvis K. G., Deng Y. K., Lai L. C., McNamara B. P., Donnenberg M. S., Kaper J. B.. 1998; The complete sequence of the locus of enterocyte effacement (LEE) from enteropathogenic Escherichia coli E2348/69. Mol Microbiol28:1–4
    [Google Scholar]
  16. Estojak J., Brent R., Golemis E. A.. 1995; Correlation of two-hybrid affinity data with in vitro measurements. Mol Cell Biol15:5820–5829
    [Google Scholar]
  17. Fashena S. J., Serebriiskii I. G., Golemis E. A.. 2000; LexA-based two-hybrid systems. Methods Enzymol328:14–26
    [Google Scholar]
  18. Foultier B., Troisfontaines P., Muller S., Opperdoes F. R., Cornelis G. R.. 2002; Characterization of the ysa pathogenicity locus in the chromosome of Yersinia entercolitica and phylogeny analysis of type III secretion systems. J Mol Evol55:37–51[CrossRef]
    [Google Scholar]
  19. Francis M. S., Aili M., Wiklund M. L., Wolf-Watz H.. 2000; A study of the YopD-LcrH interaction from Yersinia pseudotuberculosis reveals a role for hydrophobic residues within the amphipathic domain of YopD. Mol Microbiol38:85–102[CrossRef]
    [Google Scholar]
  20. Gauthier A., Finlay B. B.. 2003; Translocated intimin receptor and its chaperone interact with ATPase of the type III secretion apparatus of enteropathogenic Escherichia coli. J Bacteriol185:6747–6755[CrossRef]
    [Google Scholar]
  21. Guy R. L., Gonias L. A., Stein M. A.. 2000; Aggregation of host endosomes by Salmonella requires SPI2 translocation of SseFG and involves SpvR and the fms-aroE intragenic region. Mol Microbiol37:1417–1435[CrossRef]
    [Google Scholar]
  22. 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 Bacteriol177:4121–4131
    [Google Scholar]
  23. Gyuris J., Golemis E., Chertkov H., Brent R.. 1993; Cdi1, a human G1 and S phase protein phosphatase that associates with Cdk2. Cell75:791–803[CrossRef]
    [Google Scholar]
  24. Hartland E. L., Daniell S. J., Delahay R. M., Neves B. C., Wallis T., Shaw R. K., Hale C., Knutton S., Frankel G.. 2000; The type III protein translocation system of enteropathogenic Escherichia coli involves EspA-EspB protein interactions. Mol Microbiol35:1483–1492
    [Google Scholar]
  25. Hensel M., Shea J. E., Waterman S. R.. & 7 other authors. 1998; Genes encoding putative effector proteins of the type III secretion system of Salmonella pathogenicity island 2 are required for bacterial virulence and proliferation in macrophages. Mol Microbiol30:163–174[CrossRef]
    [Google Scholar]
  26. Holden D. W.. 2002; Trafficking of the Salmonella vacuole in macrophages. Traffic 3:161–169[CrossRef]
    [Google Scholar]
  27. Ide T., Laarmann S., Greune L., Schillers H., Oberleithner H., Schmidt M. A.. 2001; Characterization of translocation pores inserted into plasma membranes by type III-secreted Esp proteins of enteropathogenic Escherichia coli. Cell Microbiol3:669–679[CrossRef]
    [Google Scholar]
  28. Klein J. R., Jones B. D.. 2001; Salmonella pathogenicity island 2-encoded proteins SseC and SseD are essential for virulence and are substrates of the type III secretion system. Infect Immun69:737–743[CrossRef]
    [Google Scholar]
  29. Knodler L. A., Steele-Mortimer O.. 2003; Taking possession: biogenesis of the Salmonella-containing vacuole. Traffic4:587–599[CrossRef]
    [Google Scholar]
  30. Kolonin M. G., Zhong J., Finley R. L.. 2000; Interaction mating methods in two-hybrid systems. Methods Enzymol328:26–46
    [Google Scholar]
  31. Lupas A., Van Dyke M., Stock J.. 1991; Predicting coiled coils from protein sequences. Science252:1162–1164[CrossRef]
    [Google Scholar]
  32. Miller J. H.. 1972; Experiments in Molecular Genetics Cold Spring Harbor, NY: Cold Spring Laboratory;
  33. Neyt C., Cornelis G. R.. 1999; Role of SycD, the chaperone of the Yersinia Yop translocators YopB and YopD. Mol Microbiol3:143–156
    [Google Scholar]
  34. Nikolaus T., Deiwick J., Rappl C., Freeman J. A., Schroder W., Miller S. I., Hensel M.. 2001; SseBCD proteins are secreted by the type III secretion system of Salmonella pathogenicity island 2 and function as a translocon. J Bacteriol183:6036–6045[CrossRef]
    [Google Scholar]
  35. Ozin A., Claret L., Uvray F., Hughes C.. 2003; The FliS chaperone selectively binds the disordered flagellin C-termina D0 domain central to polymerization. FEMS Microbiol Lett219:219–224[CrossRef]
    [Google Scholar]
  36. Page A.-L., Fromont-Racine M., Sansonetti P., Legrain P., Parsot C.. 2001; Characterization of the interaction partners of secreted proteins and chaperones of Shigella flexneri. Mol Microbiol42:1133–1145[CrossRef]
    [Google Scholar]
  37. Parsot C., Hamiaux C., Page A.-L.. 2003; The various and varying roles of specific chaperones in type III secretion systems. Curr Opin Microbiol6:7–13[CrossRef]
    [Google Scholar]
  38. Richman T. J., Sawyer M. M., Johnson D. I.. 1999; The Cdc42p GTPase is involved in a G2/M morphogenetic checkpoint regulating the apical-isotropic switch and nuclear division in yeast. J Biol Chem274:16861–16870[CrossRef]
    [Google Scholar]
  39. Ruiz-Albert J., Mundy R., Yu X. J., Holden D. W., Beuzón C. R.. 2003; SseA is a chaperone for the SseB and SseD translocon components of the Salmonella pathogenicity-island-2-encoded type III secretion system. Microbiology149:1103–1111[CrossRef]
    [Google Scholar]
  40. Sekiya K., Ohishi M., Ogino T., Tamano K., Sasakawa C., Abe A.. 2001; Supermolecular structure of the enteropathogenic Escherichia coli type III secretion system and its direct interaction with the EspA-sheath-like structure. Proc Natl Acad Sci U S A98:11638–11643[CrossRef]
    [Google Scholar]
  41. Stein M. A., Leung K. Y., Zwick M., Garcia-del Portillo F., Finlay B. B.. 1996; Identification of a Salmonella virulence gene required for formation of filamentous structures containing lysosomal membrane glycoproteins within epithelial cells. Mol Microbiol20:151–164[CrossRef]
    [Google Scholar]
  42. Suvarnapunya A. E., Stein M. A., Lagassé H. A. D.. 2003; The role of DNA base excision repair in the pathogenesis of Salmonella enterica serovar Typhimurium. Mol Microbiol48:549–559[CrossRef]
    [Google Scholar]
  43. Thomas J., Stafford G. P., Hughes C.. 2004; Docking of cytosolic chaperone-substrate complexes at the membrane ATPase during flagellar type III protein export. Proc Natl Acad Sci U S A101:3945–3950[CrossRef]
    [Google Scholar]
  44. Wainwright L. A., Kaper J. B.. 1998; EspB and EspD require a specific chaperone for proper secretion from enteropathogenic Escherichia coli. Mol Microbiol27:1247–1260[CrossRef]
    [Google Scholar]
  45. Wolf E., Kim P. S., Berger B.. 1997; MultiCoil: a program for predicting two- and three-stranded coiled coils. Protein Sci6:1179–1189[CrossRef]
    [Google Scholar]
  46. Yaffe M. P., Schatz G.. 1984; Two nuclear mutations that block mitochondrial protein import in yeast. Proc Natl Acad Sci U S A81:4819–4823[CrossRef]
    [Google Scholar]
  47. Zurawski D. V., Stein M. A.. 2003; SseA acts as the chaperone for the SseB component of the Salmonella Pathogenicity Island 2 translocon. Mol Microbiol47:1341–1351[CrossRef]
    [Google Scholar]
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