1887

Abstract

The bacterial pathogen requires colonizination of the human small intestine to cause cholera. The anaerobic and slightly acidic conditions predominating there enhance toxicity of low copper concentrations and create a selective environment for bacteria with evolved detoxifying mechanisms. We reported previously that the VCA0260, VCA0261 and VC2216 gene products were synthesized only in grown in microaerobiosis or anaerobiosis. Here we show that ORFs VCA0261 and VCA0260 are actually combined into a single gene encoding a 18.7 kDa protein. Bioinformatic analyses linked this protein and the VC2216 gene product to copper tolerance. Following the approach of predict-mutate and test, we describe for the first time, to our knowledge, the copper tolerance systems operating in . Copper susceptibility analyses of mutants in VCA0261–0260, VC2216 or in the putative copper-tolerance-related VC2215 ( ATPase) and VC0974 (), under aerobic and anaerobic growth, revealed that CopA represents the main tolerance system under both conditions. The VC2216-encoded periplasmic protein contributes to resistance only under anaerobiosis in a CopA-functional background. The locus tag VCA0261–0260 encodes a copper-inducible, CueR-dependent, periplasmic protein, which mediates tolerance in aerobiosis, but under anaerobiosis its role is only evident in CopA knock-out mutants. None of the genes involved in copper homeostasis were required for virulence or colonization in the mouse model. We conclude that copper tolerance in , which lacks orthologues of the periplasmic copper tolerance proteins CueO, CusCFBA and CueP, involves CopA and CueR proteins along with the periplasmic Cot (VCA0261–0260) and CopG (VC2216) homologues.

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2012-08-01
2020-01-25
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References

  1. Abd H., Saeed A., Weintraub A., Nair G. B., Sandström G.. ( 2007;). Vibrio cholerae O1 strains are facultative intracellular bacteria, able to survive and multiply symbiotically inside the aquatic free-living amoeba Acanthamoeba castellanii. FEMS Microbiol Ecol60:33–39 [CrossRef][PubMed]
    [Google Scholar]
  2. Ausubel F. M., Brent R., Kingston R. E., Moore D. D., Seidman J. G., Smith J. A., Struhl K.. ( 1995;). Short Protocols in Molecular Biology New York: John Wiley & Sons, Inc;
    [Google Scholar]
  3. Bagai I., Rensing C., Blackburn N. J., McEvoy M. M.. ( 2008;). Direct metal transfer between periplasmic proteins identifies a bacterial copper chaperone. Biochemistry47:11408–11414 [CrossRef][PubMed]
    [Google Scholar]
  4. Barry A. N., Otoikhian A., Bhatt S., Shinde U., Tsivkovskii R., Blackburn N. J., Lutsenko S.. ( 2011;). The lumenal loop Met672-Pro707 of copper-transporting ATPase ATP7A binds metals and facilitates copper release from the intramembrane sites. J Biol Chem286:26585–26594 [CrossRef][PubMed]
    [Google Scholar]
  5. Charbonnier J. B., Belin P., Moutiez M., Stura E. A., Quéméneur E.. ( 1999;). On the role of the cis-proline residue in the active site of DsbA. Protein Sci8:96–105 [CrossRef][PubMed]
    [Google Scholar]
  6. Choi M., Davidson V. L.. ( 2011;). Cupredoxins–a study of how proteins may evolve to use metals for bioenergetic processes. Metallomics3:140–151 [CrossRef][PubMed]
    [Google Scholar]
  7. Donnenberg M. S., Kaper J. B.. ( 1991;). Construction of an eae deletion mutant of enteropathogenic Escherichia coli by using a positive-selection suicide vector. Infect Immun59:4310–4317[PubMed]
    [Google Scholar]
  8. Egler M., Grosse C., Grass G., Nies D. H.. ( 2005;). Role of the extracytoplasmic function protein family sigma factor RpoE in metal resistance of Escherichia coli. J Bacteriol187:2297–2307 [CrossRef][PubMed]
    [Google Scholar]
  9. Espariz M., Checa S. K., Audero M. E., Pontel L. B., Soncini F. C.. ( 2007;). Dissecting the Salmonella response to copper. Microbiology153:2989–2997 [CrossRef][PubMed]
    [Google Scholar]
  10. Faruque S. M., Nair G. B., Mekalanos J. J.. ( 2004;). Genetics of stress adaptation and virulence in toxigenic Vibrio cholerae. DNA Cell Biol23:723–741 [CrossRef][PubMed]
    [Google Scholar]
  11. González-Guerrero M., Raimunda D., Cheng X., Argüello J. M.. ( 2010;). Distinct functional roles of homologous Cu+ efflux ATPases in Pseudomonas aeruginosa. Mol Microbiol78:1246–1258 [CrossRef][PubMed]
    [Google Scholar]
  12. Grass G., Rensing C.. ( 2001;). Genes involved in copper homeostasis in Escherichia coli. J Bacteriol183:2145–2147 [CrossRef][PubMed]
    [Google Scholar]
  13. Gupta S. D., Lee B. T., Camakaris J., Wu H. C.. ( 1995;). Identification of cutC and cutF (nlpE) genes involved in copper tolerance in Escherichia coli. J Bacteriol177:4207–4215[PubMed]
    [Google Scholar]
  14. Heidelberg J. F., Eisen J. A., Nelson W. C., Clayton R. A., Gwinn M. L., Dodson R. J., Haft D. H., Hickey E. K., Peterson J. D.. & other authors ( 2000;). DNA sequence of both chromosomes of the cholera pathogen Vibrio cholerae. Nature406:477–483 [CrossRef][PubMed]
    [Google Scholar]
  15. Hiniker A., Collet J. F., Bardwell J. C.. ( 2005;). Copper stress causes an in vivo requirement for the Escherichia coli disulfide isomerase DsbC. J Biol Chem280:33785–33791 [CrossRef][PubMed]
    [Google Scholar]
  16. Hirano Y., Hossain M. M., Takeda K., Tokuda H., Miki K.. ( 2006;). Purification, crystallization and preliminary X-ray crystallographic analysis of the outer membrane lipoprotein NlpE from Escherichia coli. Acta Crystallogr Sect F Struct Biol Cryst Commun62:1227–1230 [CrossRef][PubMed]
    [Google Scholar]
  17. Hirst T. R., Holmgren J.. ( 1987;). Transient entry of enterotoxin subunits into the periplasm occurs during their secretion from Vibrio cholerae. J Bacteriol169:1037–1045[PubMed]
    [Google Scholar]
  18. Janssen G. R., Bibb M. J.. ( 1993;). Derivatives of pUC18 that have BglII sites flanking a modified multiple cloning site and that retain the ability to identify recombinant clones by visual screening of Escherichia coli colonies. Gene124:133–134 [CrossRef][PubMed]
    [Google Scholar]
  19. Kim E. H., Rensing C., McEvoy M. M.. ( 2010;). Chaperone-mediated copper handling in the periplasm. Nat Prod Rep27:711–719 [CrossRef][PubMed]
    [Google Scholar]
  20. Kinch L. N., Baker D., Grishin N. V.. ( 2003;). Deciphering a novel thioredoxin-like fold family. Proteins52:323–331 [CrossRef][PubMed]
    [Google Scholar]
  21. Li J., Ji C., Chen J., Yang Z., Wang Y., Fei X., Zheng M., Gu X., Wen G.. & other authors ( 2005;). Identification and characterization of a novel Cut family cDNA that encodes human copper transporter protein CutC. Biochem Biophys Res Commun337:179–183 [CrossRef][PubMed]
    [Google Scholar]
  22. Lim S. Y., Joe M. H., Song S. S., Lee M. H., Foster J. W., Park Y. K., Choi S. Y., Lee I. S.. ( 2002;). CuiD is a crucial gene for survival at high copper environment in Salmonella enterica serovar Typhimurium. Mol Cells14:177–184[PubMed]
    [Google Scholar]
  23. Macomber L., Imlay J. A.. ( 2009;). The iron-sulfur clusters of dehydratases are primary intracellular targets of copper toxicity. Proc Natl Acad Sci U S A106:8344–8349 [CrossRef][PubMed]
    [Google Scholar]
  24. Macomber L., Rensing C., Imlay J. A.. ( 2007;). Intracellular copper does not catalyze the formation of oxidative DNA damage in Escherichia coli. J Bacteriol189:1616–1626 [CrossRef][PubMed]
    [Google Scholar]
  25. Marchler-Bauer A., Lu S., Anderson J. B., Chitsaz F., Derbyshire M. K., DeWeese-Scott C., Fong J. H., Geer L. Y., Geer R. C.. & other authors ( 2011;). CDD: a Conserved Domain Database for the functional annotation of proteins. Nucleic Acids Res39:Database issueD225–D229 [CrossRef][PubMed]
    [Google Scholar]
  26. Marrero K., Sánchez A., Rodríguez-Ulloa A., González L. J., Castellanos-Serra L., Paz-Lago D., Campos J., Rodríguez B. L., Suzarte E.. & other authors ( 2009;). Anaerobic growth promotes synthesis of colonization factors encoded at the vibrio pathogenicity island in Vibrio cholerae El Tor. Res Microbiol160:48–56 [CrossRef][PubMed]
    [Google Scholar]
  27. Monchy S., Benotmane M. A., Wattiez R., van Aelst S., Auquier V., Borremans B., Mergeay M., Taghavi S., van der Lelie D., Vallaeys T.. ( 2006;). Transcriptomic and proteomic analyses of the pMOL30-encoded copper resistance in Cupriavidus metallidurans strain CH34. Microbiology152:1765–1776 [CrossRef][PubMed]
    [Google Scholar]
  28. Mueller R. S., McDougald D., Cusumano D., Sodhi N., Kjelleberg S., Azam F., Bartlett D. H.. ( 2007;). Vibrio cholerae strains possess multiple strategies for abiotic and biotic surface colonization. J Bacteriol189:5348–5360 [CrossRef][PubMed]
    [Google Scholar]
  29. Osman D., Cavet J. S.. ( 2008;). Copper homeostasis in bacteria. Adv Appl Microbiol65:217–247 [CrossRef][PubMed]
    [Google Scholar]
  30. Osman D., Waldron K. J., Denton H., Taylor C. M., Grant A. J., Mastroeni P., Robinson N. J., Cavet J. S.. ( 2010;). Copper homeostasis in Salmonella is atypical and copper-CueP is a major periplasmic metal complex. J Biol Chem285:25259–25268 [CrossRef][PubMed]
    [Google Scholar]
  31. Outten F. W., Outten C. E., Hale J., O’Halloran T. V.. ( 2000;). Transcriptional activation of an Escherichia coli copper efflux regulon by the chromosomal MerR homologue, cueR. J Biol Chem275:31024–31029 [CrossRef][PubMed]
    [Google Scholar]
  32. Outten F. W., Huffman D. L., Hale J. A., O’Halloran T. V.. ( 2001;). The independent cue and cus systems confer copper tolerance during aerobic and anaerobic growth in Escherichia coli. J Biol Chem276:30670–30677 [CrossRef][PubMed]
    [Google Scholar]
  33. Pérez Audero M. E., Podoroska B. M., Ibáñez M. M., Cauerhff A., Checa S. K., Soncini F. C.. ( 2010;). Target transcription binding sites differentiate two groups of MerR-monovalent metal ion sensors. Mol Microbiol78:853–865 [CrossRef][PubMed]
    [Google Scholar]
  34. Pontel L. B., Soncini F. C.. ( 2009;). Alternative periplasmic copper-resistance mechanisms in Gram negative bacteria. Mol Microbiol73:212–225 [CrossRef][PubMed]
    [Google Scholar]
  35. Raimunda D., González-Guerrero M., Leeber B. W. III, Argüello J. M.. ( 2011;). The transport mechanism of bacterial Cu+-ATPases: distinct efflux rates adapted to different function. Biometals24:467–475 [CrossRef][PubMed]
    [Google Scholar]
  36. Rensing C., Grass G.. ( 2003;). Escherichia coli mechanisms of copper homeostasis in a changing environment. FEMS Microbiol Rev27:197–213 [CrossRef][PubMed]
    [Google Scholar]
  37. Rensing C., Fan B., Sharma R., Mitra B., Rosen B. P.. ( 2000;). CopA: An Escherichia coli Cu(I)-translocating P-type ATPase. Proc Natl Acad Sci U S A97:652–656 [CrossRef][PubMed]
    [Google Scholar]
  38. Robinson N. J., Winge D. R.. ( 2010;). Copper metallochaperones. Annu Rev Biochem79:537–562 [CrossRef][PubMed]
    [Google Scholar]
  39. Roy A., Kucukural A., Zhang Y.. ( 2010;). I-TASSER: a unified platform for automated protein structure and function prediction. Nat Protoc5:725–738 [CrossRef][PubMed]
    [Google Scholar]
  40. Rubino J. T., Franz K. J.. ( 2012;). Coordination chemistry of copper proteins: how nature handles a toxic cargo for essential function. J Inorg Biochem107:129–143 [CrossRef][PubMed]
    [Google Scholar]
  41. Sandström G., Saeed A., Abd H.. ( 2010;). Acanthamoeba polyphaga is a possible host for Vibrio cholerae in aquatic environments. Exp Parasitol126:65–68 [CrossRef][PubMed]
    [Google Scholar]
  42. Senanayake S. D., Brian D. A.. ( 1995;). Precise large deletions by the PCR-based overlap extension method. Mol Biotechnol4:13–15 [CrossRef][PubMed]
    [Google Scholar]
  43. Simon R., Priefer U., Pühler A.. ( 1983;). A broad host range mobilization system for in vivo genetic engineering: transposon mutagenesis in Gram-negative bacteria. Biotechnology (N Y)1:784–791 [CrossRef]
    [Google Scholar]
  44. Singh S. K., Grass G., Rensing C., Montfort W. R.. ( 2004;). Cuprous oxidase activity of CueO from Escherichia coli. J Bacteriol186:7815–7817 [CrossRef][PubMed]
    [Google Scholar]
  45. Stoebner J. A., Butterton J. R., Calderwood S. B., Payne S. M.. ( 1992;). Identification of the vibriobactin receptor of Vibrio cholerae. J Bacteriol174:3270–3274[PubMed]
    [Google Scholar]
  46. Stoyanov J. V., Hobman J. L., Brown N. L.. ( 2001;). CueR (YbbI) of Escherichia coli is a MerR family regulator controlling expression of the copper exporter CopA. Mol Microbiol39:502–512 [CrossRef][PubMed]
    [Google Scholar]
  47. Stoyanov J. V., Magnani D., Solioz M.. ( 2003;). Measurement of cytoplasmic copper, silver, and gold with a lux biosensor shows copper and silver, but not gold, efflux by the CopA ATPase of Escherichia coli. FEBS Lett546:391–394 [CrossRef][PubMed]
    [Google Scholar]
  48. Su D., Berndt C., Fomenko D. E., Holmgren A., Gladyshev V. N.. ( 2007;). A conserved cis-proline precludes metal binding by the active site thiolates in members of the thioredoxin family of proteins. Biochemistry46:6903–6910 [CrossRef][PubMed]
    [Google Scholar]
  49. Taylor L. A., Rose R. E.. ( 1988;). A correction in the nucleotide sequence of the Tn903 kanamycin resistance determinant in pUC4K. Nucleic Acids Res16:358 [CrossRef][PubMed]
    [Google Scholar]
  50. Teitzel G. M., Geddie A., De Long S. K., Kirisits M. J., Whiteley M., Parsek M. R.. ( 2006;). Survival and growth in the presence of elevated copper: transcriptional profiling of copper-stressed Pseudomonas aeruginosa. J Bacteriol188:7242–7256 [CrossRef][PubMed]
    [Google Scholar]
  51. Tom-Petersen A., Hosbond C., Nybroe O.. ( 2001;). Identification of copper-induced genes in Pseudomonas fluorescens and use of a reporter strain to monitor bioavailable copper in soil. FEMS Microbiol Ecol38:59–67 [CrossRef]
    [Google Scholar]
  52. Valle E., Ledón T., Cedré B., Campos J., Valmaseda T., Rodríguez B., García L., Marrero K., Benítez J.. & other authors ( 2000;). Construction and characterization of a nonproliferative El Tor cholera vaccine candidate derived from strain 638. Infect Immun68:6411–6418 [CrossRef][PubMed]
    [Google Scholar]
  53. von Krüger W. M. A., Santos Lery L. M., Soares M. R., Saloum de Neves-Manta F., Batista e Silva C. M., da Costa Neves-Ferreira A. G., Perales J., Bisch P. M.. ( 2006;). The phosphate-starvation response in Vibrio cholerae O1 and phoB mutant under proteomic analysis: disclosing functions involved in adaptation, survival and virulence. Proteomics6:1495–1511 [CrossRef][PubMed]
    [Google Scholar]
  54. Weissman Z., Berdicevsky I., Cavari B. Z., Kornitzer D.. ( 2000;). The high copper tolerance of Candida albicans is mediated by a P-type ATPase. Proc Natl Acad Sci U S A97:3520–3525 [CrossRef][PubMed]
    [Google Scholar]
  55. Zhang X., Bremer H.. ( 1995;). Control of the Escherichia coli rrnB P1 promoter strength by ppGpp. J Biol Chem270:11181–11189 [CrossRef][PubMed]
    [Google Scholar]
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