1887

Abstract

The localization and membrane topology of the YscJ lipoprotein, an essential component of the type III secretion apparatus, was investigated. YscJ was demonstrated to be an inner membrane (IM) lipoprotein that is anchored to the periplasmic face of the IM via an N-terminal lipid moiety and via a C-terminal transmembrane (TM) domain. Localization of the N-terminal lipid moiety to the IM occurred regardless of the amino-acid residues found in the +2 or +3 positions. IM localization was dependent upon an intact N-terminal domain (amino acids +1 to +61), suggesting that this region plays a role in YscJ localization. In contrast, the YscJ C-terminal domain and TM domain were not required for IM localization. N-terminal sequence analysis demonstrated that a significant proportion of membrane-localized YscJ lacks -acylation, the final modification required for Lol-dependent transport of a lipoprotein to the OM. Interestingly, attachment of the N-terminus to the IM was required for YscJ function; however, the YscJ secretion signal and lipo-box could be functionally replaced by the first TM domain of the YscV protein, suggesting that the mechanism of attachment to the IM was not critical.

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2008-02-01
2019-11-22
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References

  1. Alexeyev, M. F. & Winkler, H. H. ( 1999; ). Membrane topology of the Rickettsia prowazekii ATP/ADP translocase revealed by novel dual pho-lac reporters. J Mol Biol 285, 1503–1513.[CrossRef]
    [Google Scholar]
  2. Ali, S. A. & Steinkasserer, A. ( 1995; ). PCR-ligation-PCR mutagenesis: a protocol for creating gene fusions and mutations. Biotechniques 18, 746–750.
    [Google Scholar]
  3. Allaoui, A., Sansonetti, P. J. & Parsot, C. ( 1992; ). MxiJ, a lipoprotein involved in secretion of Shigella Ipa invasins, is homologous to YscJ, a secretion factor of the Yersinia Yop proteins. J Bacteriol 174, 7661–7669.
    [Google Scholar]
  4. Allaoui, A., Schulte, R. & Cornelis, G. R. ( 1995; ). Mutational analysis of the Yersinia enterocolitica virC operon: characterization of yscE, F, G, I, J, K required for Yop secretion and yscH encoding YopR. Mol Microbiol 18, 343–355.[CrossRef]
    [Google Scholar]
  5. Blocker, A., Gounon, P., Larquet, E., Niebuhr, K., Cabiaux, V., Parsot, C. & Sansonetti, P. ( 1999; ). The tripartite type III secreton of Shigella flexneri inserts IpaB and IpaC into host membranes. J Cell Biol 147, 683–693.[CrossRef]
    [Google Scholar]
  6. 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.[CrossRef]
    [Google Scholar]
  7. Burghout, P., Beckers, F., de Wit, E., van Boxtel, R., Cornelis, G. R., Tommassen, J. & Koster, M. ( 2004; ). Role of the pilot protein YscW in the biogenesis of the YscC secretin in Yersinia enterocolitica. J Bacteriol 186, 5366–5375.[CrossRef]
    [Google Scholar]
  8. Cambau, E., Bordon, F., Collatz, E. & Gutmann, L. ( 1993; ). Novel gyrA point mutation in a strain of Escherichia coli resistant to fluoroquinolones but not to nalidixic acid. Antimicrob Agents Chemother 37, 1247–1252.[CrossRef]
    [Google Scholar]
  9. Cornelis, G. R. ( 2002; ). The Yersinia Ysc-Yop virulence apparatus. Int J Med Microbiol 291, 455–462.
    [Google Scholar]
  10. Crago, A. M. & Koronakis, V. ( 1998; ). Salmonella InvG forms a ring-like multimer that requires the InvH lipoprotein for outer membrane localization. Mol Microbiol 30, 47–56.[CrossRef]
    [Google Scholar]
  11. Crepin, V. F., Prasannan, S., Shaw, R. K., Wilson, R. K., Creasey, E., Abe, C. M., Knutton, S., Frankel, G. & Matthews, S. ( 2005; ). Structural and functional studies of the enteropathogenic Escherichia coli type III needle complex protein EscJ. Mol Microbiol 55, 1658–1670.[CrossRef]
    [Google Scholar]
  12. Fukuda, A., Matsuyama, S., Hara, T., Nakayama, J., Nagasawa, H. & Tokuda, H. ( 2002; ). Aminoacylation of the N-terminal cysteine is essential for Lol-dependent release of lipoproteins from membranes but does not depend on lipoprotein sorting signals. J Biol Chem 277, 43512–43518.[CrossRef]
    [Google Scholar]
  13. Goguen, J. D., Yother, J. & Straley, S. C. ( 1984; ). Genetic analysis of the low calcium response in Yersinia pestis Mu d1(Ap lac) insertion mutants. J Bacteriol 160, 842–848.
    [Google Scholar]
  14. 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 Bacteriol 177, 4121–4130.
    [Google Scholar]
  15. Haddix, P. L. & Straley, S. C. ( 1992; ). Structure and regulation of the Yersinia pestis yscBCDEF operon. J Bacteriol 174, 4820–4828.
    [Google Scholar]
  16. Hara, T., Matsuyama, S. & Tokuda, H. ( 2003; ). Mechanism underlying the inner membrane retention of Escherichia coli lipoproteins caused by Lol avoidance signals. J Biol Chem 278, 40408–40414.[CrossRef]
    [Google Scholar]
  17. Hayashi, S. & Wu, H. C. ( 1990; ). Lipoproteins in bacteria. J Bioenerg Biomembr 22, 451–471.[CrossRef]
    [Google Scholar]
  18. Huang, H. C., Sherman, M. Y., Kandror, O. & Goldberg, A. L. ( 2001; ). The molecular chaperone DnaJ is required for the degradation of a soluble abnormal protein in Escherichia coli. J Biol Chem 276, 3920–3928.[CrossRef]
    [Google Scholar]
  19. Hueck, C. J. ( 1998; ). Type III protein secretion systems in bacterial pathogens of animals and plants. Microbiol Mol Biol Rev 62, 379–433.
    [Google Scholar]
  20. Jackson, M. W. & Plano, G. V. ( 1999; ). DsbA is required for stable expression of outer membrane protein YscC and for efficient Yop secretion in Yersinia pestis. J Bacteriol 181, 5126–5130.
    [Google Scholar]
  21. Juris, S. J., Shao, F. & Dixon, J. E. ( 2002; ). Yersinia effectors target mammalian signalling pathways. Cell Microbiol 4, 201–211.[CrossRef]
    [Google Scholar]
  22. Kimbrough, T. G. & Miller, S. I. ( 2000; ). Contribution of Salmonella typhimurium type III secretion components to needle complex formation. Proc Natl Acad Sci U S A 97, 11008–11013.[CrossRef]
    [Google Scholar]
  23. Koster, M., Bitter, W., de Cock, H., Allaoui, A., Cornelis, G. R. & Tommassen, J. ( 1997; ). The outer membrane component, YscC, of the Yop secretion machinery of Yersinia enterocolitica forms a ring-shaped multimeric complex. Mol Microbiol 26, 789–797.[CrossRef]
    [Google Scholar]
  24. Kubori, T., Matsushima, Y., Nakamura, D., Uralil, J., Lara-Tejero, M., Sukhan, A., Galan, J. E. & Aizawa, S. I. ( 1998; ). Supramolecular structure of the Salmonella typhimurium type III protein secretion system. Science 280, 602–605.[CrossRef]
    [Google Scholar]
  25. Macnab, R. M. ( 1999; ). The bacterial flagellum: reversible rotary propellor and type III export apparatus. J Bacteriol 181, 7149–7153.
    [Google Scholar]
  26. Manoil, C., Boyd, D. & Beckwith, J. ( 1988; ). Molecular genetic analysis of membrane protein topology. Trends Genet 4, 223–226.[CrossRef]
    [Google Scholar]
  27. Marlovits, T. C., Kubori, T., Sukhan, A., Thomas, D. R., Galan, J. E. & Unger, V. M. ( 2004; ). Structural insights into the assembly of the type III secretion needle complex. Science 306, 1040–1042.[CrossRef]
    [Google Scholar]
  28. Masuda, K., Matsuyama, S. & Tokuda, H. ( 2002; ). Elucidation of the function of lipoprotein-sorting signals that determine membrane localization. Proc Natl Acad Sci U S A 99, 7390–7395.[CrossRef]
    [Google Scholar]
  29. Matsuyama, S., Tajima, T. & Tokuda, H. ( 1995; ). A novel periplasmic carrier protein involved in the sorting and transport of Escherichia coli lipoproteins destined for the outer membrane. EMBO J 14, 3365–3372.
    [Google Scholar]
  30. Matsuyama, S., Yokota, N. & Tokuda, H. ( 1997; ). A novel outer membrane lipoprotein, LolB (HemM), involved in the LolA (p20)-dependent localization of lipoproteins to the outer membrane of Escherichia coli. EMBO J 16, 6947–6955.[CrossRef]
    [Google Scholar]
  31. Michiels, T., Vanooteghem, J. C., Lambert de Rouvroit, C., China, B., Gustin, A., Boudry, P. & Cornelis, G. R. ( 1991; ). Analysis of virC, an operon involved in the secretion of Yop proteins by Yersinia enterocolitica. J Bacteriol 173, 4994–5009.
    [Google Scholar]
  32. 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.
    [Google Scholar]
  33. Pallen, M. J., Beatson, S. A. & Bailey, C. M. ( 2005; ). Bioinformatics, genomics and evolution of non-flagellar type-III secretion systems: a Darwinian perspective. FEMS Microbiol Rev 29, 201–229.[CrossRef]
    [Google Scholar]
  34. Plano, G. V. & Straley, S. C. ( 1995; ). Mutations in yscC, yscD, and yscG prevent high-level expression and secretion of V antigen and Yops in Yersinia pestis. J Bacteriol 177, 3843–3854.
    [Google Scholar]
  35. Plano, G. V., Barve, S. S. & Straley, S. C. ( 1991; ). LcrD, a membrane-bound regulator of the Yersinia pestis low-calcium response. J Bacteriol 173, 7293–7303.
    [Google Scholar]
  36. Robichon, C., Bonhivers, M. & Pugsley, A. P. ( 2003; ). An intramolecular disulphide bond reduces the efficacy of a lipoprotein plasma membrane sorting signal. Mol Microbiol 49, 1145–1154.[CrossRef]
    [Google Scholar]
  37. Robichon, C., Vidal-Ingigliardi, D. & Pugsley, A. P. ( 2005; ). Depletion of apolipoprotein N-acyltransferase causes mislocalization of outer membrane lipoproteins in Escherichia coli. J Biol Chem 280, 974–983.[CrossRef]
    [Google Scholar]
  38. Rosqvist, R., Magnusson, K. E. & Wolf-Watz, H. ( 1994; ). Target cell contact triggers expression and polarized transfer of Yersinia YopE cytotoxin into mammalian cells. EMBO J 13, 964–972.
    [Google Scholar]
  39. 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 A 98, 11638–11643.[CrossRef]
    [Google Scholar]
  40. Skrzypek, E., Haddix, P. L., Plano, G. V. & Straley, S. C. ( 1993; ). New suicide vector for gene replacement in yersiniae and other gram-negative bacteria. Plasmid 29, 160–163.[CrossRef]
    [Google Scholar]
  41. Stoker, N. G., Fairweather, N. F. & Spratt, B. G. ( 1982; ). Versatile low-copy-number plasmid vectors for cloning in Escherichia coli. Gene 18, 335–341.[CrossRef]
    [Google Scholar]
  42. Terada, M., Kuroda, T., Matsuyama, S. I. & Tokuda, H. ( 2001; ). Lipoprotein sorting signals evaluated as the LolA-dependent release of lipoproteins from the cytoplasmic membrane of Escherichia coli. J Biol Chem 276, 47690–47694.[CrossRef]
    [Google Scholar]
  43. Tokuda, H. & Matsuyama, S. ( 2004; ). Sorting of lipoproteins to the outer membrane in E. coli. Biochim Biophys Acta 1693, 5–13.[CrossRef]
    [Google Scholar]
  44. Tokunaga, M., Tokunaga, H. & Wu, H. C. ( 1982; ). Post-translational modification and processing of Escherichia coli prolipoprotein in vitro. Proc Natl Acad Sci U S A 79, 2255–2259.[CrossRef]
    [Google Scholar]
  45. Troisfontaines, P. & Cornelis, G. R. ( 2005; ). Type III secretion: more systems than you think. Physiology (Bethesda) 20, 326–339.[CrossRef]
    [Google Scholar]
  46. Ueno, T., Oosawa, K. & Aizawa, S. ( 1994; ). Domain structures of the MS ring component protein (FliF) of the flagellar basal body of Salmonella typhimurium. J Mol Biol 236, 546–555.[CrossRef]
    [Google Scholar]
  47. Une, T. & Brubaker, R. R. ( 1984; ). In vivo comparison of avirulent Vwa and Pgm or Pstr phenotypes of yersiniae. Infect Immun 43, 895–900.
    [Google Scholar]
  48. Viboud, G. I. & Bliska, J. B. ( 2005; ). Yersinia outer proteins: role in modulation of host cell signalling responses and pathogenesis. Annu Rev Microbiol 59, 69–89.[CrossRef]
    [Google Scholar]
  49. Yakushi, T., Masuda, K., Narita, S., Matsuyama, S. & Tokuda, H. ( 2000; ). A new ABC transporter mediating the detachment of lipid-modified proteins from membranes. Nat Cell Biol 2, 212–218.[CrossRef]
    [Google Scholar]
  50. Yamaguchi, K., Yu, F. & Inouye, M. ( 1988; ). A single amino acid determinant of the membrane localization of lipoproteins in E. coli. Cell 53, 423–432.[CrossRef]
    [Google Scholar]
  51. Yip, C. K. & Strynadka, N. C. ( 2006; ). New structural insights into the bacterial type III secretion system. Trends Biochem Sci 31, 223–230.[CrossRef]
    [Google Scholar]
  52. Yip, C. K., Kimbrough, T. G., Felise, H. B., Vuckovic, M., Thomas, N. A., Pfuetzner, R., Frey, E. A., Finlay, B. B., Miller, S. I. & Strynadka, N. C. ( 2005; ). Structural characterization of the molecular platform for type III secretion system assembly. Nature 435, 702–707.[CrossRef]
    [Google Scholar]
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