1887

Microbiology Society logo

Volume 156, Issue 5

Analysis of HmsH and its role in plague biofilm formation Free

Abstract

The Hms phenotype is a manifestation of biofilm formation that causes adsorption of Congo red and haemin at 26 °C but not at 37 °C. This phenotype is required for blockage of the proventricular valve of the oriental rat flea and plays a role in transmission of bubonic plague from fleas to mammals. Genes responsible for this phenotype are located in three separate operons, , and HmsH and HmsF are outer membrane (OM) proteins, while the other four Hms proteins are located in the inner membrane. According to the Hidden Markov Method-based predictor, HmsH has a large N terminus in the periplasm, a -barrel structure with 16 -strands that traverse the OM, eight surface-exposed loops, and seven short turns connecting the -strands on the periplasmic side. Here, we demonstrate that HmsH is a heat-modifiable protein, a characteristic of other -barrel proteins, thereby supporting the bioinformatics analysis. Alanine scanning mutagenesis was used to identify conserved amino acids in the HmsH-like family that are critical for the function of HmsH in biofilm formation. Of 23 conserved amino acids mutated, four residues affected HmsH function and three likely caused protein instability. We used formaldehyde cross-linking to demonstrate that HmsH interacts with HmsF but not with HmsR, HmsS, HmsT or HmsP. Loss-of-function HmsH variants with single alanine substitutions retained their -structure and interaction with HmsF. Finally, using a polar  : : mini- mutant, we demonstrated that biofilm development is not important for the pathogenesis of bubonic or pneumonic plague in mice.

  • Received:
  • Accepted:
  • Revised:
  • Published Online:
Keyword(s): c-di-GMP, cyclic-di-GMP , CR, Congo red , CV, crystal violet , EPS, exopolysaccharide , GT, glycosyltransferase , IM, inner membrane , OM, outer membrane , poly-β-1,6-GlcNac, poly-β-1,6-N-acetyl-d-glucosamine and RT, room temperature
SGM
Loading

Article metrics loading...

/content/journal/micro/10.1099/mic.0.036640-0
2010-05-01
2024-05-03
Loading full text...

Full text loading...

/deliver/fulltext/micro/156/5/1424.html?itemId=/content/journal/micro/10.1099/mic.0.036640-0&mimeType=html&fmt=ahah

References

  1. Ausubel F. M. 1987 Current protocols in Molecular Biology New York: Greene Publishing & Wiley-Interscience;
  2. Bacot A. W. 1915; LXXXI. Further notes on the mechanism of the transmission of plague by fleas. J Hyg (Lond 14 :Plague Suppl. 4744–776
     [Google Scholar]
  3. Bacot A. W., Martin C. J. 1914; LXVII. Observations on the mechanism of the transmission of plague by fleas. J Hyg (Lond 13 :Plague Suppl. 3423–439
     [Google Scholar]
  4. Bagos P. G., Liakopoulos T. D., Spyropoulos I. C., Hamodrakas S. J. 2004a; A Hidden Markov Model method, capable of predicting and discriminating beta-barrel outer membrane proteins. BMC Bioinformatics 5:29
     [Google Scholar]
  5. Bagos P. G., Liakopoulos T. D., Spyropoulos I. C., Hamodrakas S. J. 2004b; PRED-TMBB: a web server for predicting the topology of beta-barrel outer membrane proteins. Nucleic Acids Res 32:W400–W404
     [Google Scholar]
  6. Bazanova L. P., Zhovtyi I., Maevskii M. P., Klimov V. Y., Popkov A. F. 1991; The seasonal dynamics of blocking in the flea Citellophorus tesquorum altaicus from the Tuva natural plague focus. Med Parazitol (Mosk) Jan-Feb 24–26
     [Google Scholar]
  7. Beesley E. D., Brubaker R. R., Janssen W. A., Surgalla M. J. 1967; Pesticins. III. Expression of coagulase and mechanism of fibrinolysis. J Bacteriol 94:19–26
     [Google Scholar]
  8. Beher M. G., Schnaitman C. A., Pugsley A. P. 1980; Major heat-modifiable outer membrane protein in Gram-negative bacteria: comparison with the OmpA protein of Escherichia coli. J Bacteriol 143:906–913
     [Google Scholar]
  9. Bell K. S., Sebaihia M., Pritchard L., Holden M. T., Hyman L. J., Holeva M. C., Thomson N. R., Bentley S. D., Churcher L. J. other authors 2004; Genome sequence of the enterobacterial phytopathogen Erwinia carotovora subsp. atroseptica and characterization of virulence factors. Proc Natl Acad Sci U S A 101:11105–11110
     [Google Scholar]
  10. Birnboim H. C., Doly J. 1979; A rapid alkaline extraction procedure for screening recombinant plasmid DNA. Nucleic Acids Res 7:1513–1523
     [Google Scholar]
  11. Blattner F. R., Plunkett G. III, Bloch C. A., Perna N. T., Burland V., Riley M., Collado-Vides J., Glasner J. D., Rode C. K. other authors 1997; The complete genome sequence of Escherichia coli K-12. Science 277:1453–1474
     [Google Scholar]
  12. Bobrov A. G., Kirillina O., Perry R. D. 2005; The phosphodiesterase activity of the HmsP EAL domain is required for negative regulation of biofilm formation in Yersinia pestis. FEMS Microbiol Lett 247:123–130
     [Google Scholar]
  13. Bobrov A. G., Kirillina O., Forman S., Mack D., Perry R. D. 2008; Insights into Yersinia pestis biofilm development: topology and co-interaction of Hms inner membrane proteins involved in exopolysaccharide production. Environ Microbiol 10:1419–1432
     [Google Scholar]
  14. Burroughs A. L. 1947; Sylvatic plague studies. The vector efficiency of nine species of fleas compared with Xenopsylla cheopis. J Hyg (Lond 45:371–391
     [Google Scholar]
  15. Darby C., Hsu J. W., Ghori N., Falkow S. 2002; Caenorhabditis elegans: plague bacteria biofilm blocks food intake. Nature 417:243–244
     [Google Scholar]
  16. Eisen R. J., Bearden S. W., Wilder A. P., Montenieri J. A., Antolin M. F., Gage K. L. 2006; Early-phase transmission of Yersinia pestis by unblocked fleas as a mechanism explaining rapidly spreading plague epizootics. Proc Natl Acad Sci U S A 103:15380–15385
     [Google Scholar]
  17. Eisen R. J., Lowell J. L., Montenieri J. A., Bearden S. W., Gage K. L. 2007a; Temporal dynamics of early-phase transmission of Yersinia pestis by unblocked fleas: secondary infectious feeds prolong efficient transmission by Oropsylla montana (Siphonaptera: Ceratophyllidae). J Med Entomol 44:672–677
     [Google Scholar]
  18. Eisen R. J., Wilder A. P., Bearden S. W., Montenieri J. A., Gage K. L. 2007b; Early-phase transmission of Yersinia pestis by unblocked Xenopsylla cheopis (Siphonaptera: Pulicidae) is as efficient as transmission by blocked fleas. J Med Entomol 44:678–682
     [Google Scholar]
  19. Fetherston J. D., Schuetze P., Perry R. D. 1992; Loss of the pigmentation phenotype in Yersinia pestis is due to the spontaneous deletion of 102 kb of chromosomal DNA which is flanked by a repetitive element. Mol Microbiol 6:2693–2704
     [Google Scholar]
  20. Fetherston J. D., Lillard J. W. Jr, Perry R. D. 1995; Analysis of the pesticin receptor from Yersinia pestis: role in iron-deficient growth and possible regulation by its siderophore. J Bacteriol 177:1824–1833
     [Google Scholar]
  21. Fields K. A., Nilles M. L., Cowan C., Straley S. C. 1999; Virulence role of V antigen of Yersinia pestis at the bacterial surface. Infect Immun 67:5395–5408
     [Google Scholar]
  22. Forman S., Bobrov A. G., Kirillina O., Craig S. K., Abney J., Fetherston J. D., Perry R. D. 2006; Identification of critical amino acid residues in the plague biofilm Hms proteins. Microbiology 152:3399–3410
     [Google Scholar]
  23. Forman S., Wulff C. R., Myers-Morales T., Cowan C., Perry R. D., Straley S. C. 2008; yadBC of Yersinia pestis, a new virulence determinant for bubonic plague. Infect Immun 76:578–587
     [Google Scholar]
  24. Gage K. L., Kosoy M. Y. 2005; Natural history of plague: perspectives from more than a century of research. Annu Rev Entomol 50:505–528
     [Google Scholar]
  25. Goller C. C., Romeo T. 2008; Environmental influences on biofilm development. Curr Top Microbiol Immunol 322:37–66
     [Google Scholar]
  26. Gong S., Bearden S. W., Geoffroy V. A., Fetherston J. D., Perry R. D. 2001; Characterization of the Yersinia pestis Yfu ABC inorganic iron transport system. Infect Immun 69:2829–2837
     [Google Scholar]
  27. Gotz F. 2002; Staphylococcus and biofilms. Mol Microbiol 43:1367–1378
     [Google Scholar]
  28. Hartzell P. L., Millstein J., LaPaglia C. 1999; Biofilm formation in hyperthermophilic Archaea. Methods Enzymol 310:335–349
     [Google Scholar]
  29. Higuchi K., Smith J. L. 1961; Studies on the nutrition and physiology of Pasteurella pestis. VI. A differential plating medium for the estimation of the mutation rate to avirulence. J Bacteriol 81:605–608
     [Google Scholar]
  30. Hinnebusch B. J., Erickson D. L. 2008; Yersinia pestis biofilm in the flea vector and its role in the transmission of plague. Curr Top Microbiol Immunol 322:229–248
     [Google Scholar]
  31. Hinnebusch B. J., Perry R. D., Schwan T. G. 1996; Role of the Yersinia pestis hemin storage ( hms) locus in the transmission of plague by fleas. Science 273:367–370
     [Google Scholar]
  32. Itoh Y., Wang X., Hinnebusch B. J., Preston J. F. III, Romeo T. 2005; Depolymerization of β-1,6- N-acetyl-d-glucosamine disrupts the integrity of diverse bacterial biofilms. J Bacteriol 187:382–387
     [Google Scholar]
  33. Itoh Y., Rice J. D., Goller C., Pannuri A., Taylor J., Meisner J., Beveridge T. J., Preston J. F. III, Romeo T. 2008; Roles of pgaABCD genes in synthesis, modification, and export of the Escherichia coli biofilm adhesin poly-beta-1,6- N-acetyl-d-glucosamine. J Bacteriol 190:3670–3680
     [Google Scholar]
  34. Izano E. A., Sadovskaya I., Vinogradov E., Mulks M. H., Velliyagounder K., Ragunath C., Kher W. B., Ramasubbu N., Jabbouri S. other authors 2007; Poly- N-acetylglucosamine mediates biofilm formation and antibiotic resistance in Actinobacillus pleuropneumoniae. Microb Pathog 43:1–9
     [Google Scholar]
  35. Jackson S., Burrows T. W. 1956; The pigmentation of Pasteurella pestis on a defined medium containing haemin. Br J Exp Pathol 37:570–576
     [Google Scholar]
  36. Jarrett C. O., Deak E., Isherwood K. E., Oyston P. C., Fischer E. R., Whitney A. R., Kobayashi S. D., DeLeo F. R., Hinnebusch B. J. 2004; Transmission of Yersinia pestis from an infectious biofilm in the flea vector. J Infect Dis 190:783–792
     [Google Scholar]
  37. Jefferson K. K. 2004; What drives bacteria to produce a biofilm?. FEMS Microbiol Lett 236:163–173
     [Google Scholar]
  38. Jones H. A., Lillard J. W. Jr, Perry R. D. 1999; HmsT, a protein essential for expression of the haemin storage (Hms+) phenotype of Yersinia pestis. Microbiology 145:2117–2128
     [Google Scholar]
  39. Kaplan J. B., Velliyagounder K., Ragunath C., Rohde H., Mack D., Knobloch J. K., Ramasubbu N. 2004; Genes involved in the synthesis and degradation of matrix polysaccharide in Actinobacillus actinomycetemcomitans and Actinobacillus pleuropneumoniae biofilms. J Bacteriol 186:8213–8220
     [Google Scholar]
  40. Kartman L., Prince F. M., Quan S. F., Stark H. E. 1958; New knowledge on the ecology of sylvatic plague. Ann N Y Acad Sci 70:668–711
     [Google Scholar]
  41. Kirillina O., Fetherston J. D., Bobrov A. G., Abney J., Perry R. D. 2004; HmsP, a putative phosphodiesterase, and HmsT, a putative diguanylate cyclase, control Hms-dependent biofilm formation in Yersinia pestis. Mol Microbiol 54:75–88
     [Google Scholar]
  42. Kropec A., Maira-Litran T., Jefferson K. K., Grout M., Cramton S. E., Gotz F., Goldmann D. A., Pier G. B. 2005; Poly- N-acetylglucosamine production in Staphylococcus aureus is essential for virulence in murine models of systemic infection. Infect Immun 73:6868–6876
     [Google Scholar]
  43. Kutyrev V. V., Filippov A. A., Oparina O. S., Protsenko O. A. 1992; Analysis of Yersinia pestis chromosomal determinants Pgm+ and Psts associated with virulence. Microb Pathog 12:177–186
     [Google Scholar]
  44. Lee B. M., Park Y. J., Park D. S., Kang H. W., Kim J. G., Song E. S., Park I. C., Yoon U. H., Hahn J. H. other authors 2005; The genome sequence of Xanthomonas oryzae pathovar oryzae KACC10331, the bacterial blight pathogen of rice. Nucleic Acids Res 33:577–586
     [Google Scholar]
  45. Lillard J. W. Jr, Fetherston J. D., Pedersen L., Pendrak M. L., Perry R. D. 1997; Sequence and genetic analysis of the hemin storage ( hms) system of Yersinia pestis. Gene 193:13–21
     [Google Scholar]
  46. Lillard J. W. Jr, Bearden S. W., Fetherston J. D., Perry R. D. 1999; The haemin storage (Hms+) phenotype of Yersinia pestis is not essential for the pathogenesis of bubonic plague in mammals. Microbiology 145:197–209
     [Google Scholar]
  47. Mack D., Fischer W., Krokotsch A., Leopold K., Hartmann R., Egge H., Laufs R. 1996; The intercellular adhesin involved in biofilm accumulation of Staphylococcus epidermidis is a linear β-1,6-linked glucosaminoglycan: purification and structural analysis. J Bacteriol 178:175–183
     [Google Scholar]
  48. Mack D., Riedewald J., Rohde H., Magnus T., Feucht H. H., Elsner H. A., Laufs R., Rupp M. E. 1999; Essential functional role of the polysaccharide intercellular adhesin of Staphylococcus epidermidis in hemagglutination. Infect Immun 67:1004–1008
     [Google Scholar]
  49. Mack D., Davies A. P., Harris L. G., Rohde H., Horstkotte M. A., Knobloch J. K. 2007; Microbial interactions in Staphylococcus epidermidis biofilms. Anal Bioanal Chem 387:399–408
     [Google Scholar]
  50. Maniatis T., Fritsch E. F., Sambrook J. 1982 Molecular Cloning: a Laboratory Manual Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
  51. Nakamura K., Mizushima S. 1976; Effects of heating in dodecyl sulfate solution on the conformation and electrophoretic mobility of isolated major outer membrane proteins from Escherichia coli K-12. J Biochem 80:1411–1422
     [Google Scholar]
  52. Nandi B., Nandy R. K., Sarkar A., Ghose A. C. 2005; Structural features, properties and regulation of the outer-membrane protein W (OmpW) of Vibrio cholerae. Microbiology 151:2975–2986
     [Google Scholar]
  53. Nitzan Y., Pechatnikov I., Bar-El D., Wexler H. 1999; Isolation and characterization of heat-modifiable proteins from the outer membrane of Porphyromonas asaccharolytica and Acinetobacter baumannii. Anaerobe 5:43–50
     [Google Scholar]
  54. O'Gara J. P. 2007; ica and beyond: biofilm mechanisms and regulation in Staphylococcus epidermidis and Staphylococcus aureus. FEMS Microbiol Lett 270:179–188
     [Google Scholar]
  55. O'Toole G. A., Pratt L. A., Watnick P. I., Newman D. K., Weaver V. B., Kolter R. 1999; Genetic approaches to study of biofilms. Methods Enzymol 310:91–109
     [Google Scholar]
  56. Otto M. 2008; Staphylococcal biofilms. Curr Top Microbiol Immunol 322:207–228
     [Google Scholar]
  57. Paulsen I. T., Press C. M., Ravel J., Kobayashi D. Y., Myers G. S., Mavrodi D. V., DeBoy R. T., Seshadri R., Ren Q. other authors 2005; Complete genome sequence of the plant commensal Pseudomonas fluorescens Pf-5. Nat Biotechnol 23:873–878
     [Google Scholar]
  58. Pendrak M. L., Perry R. D. 1991; Characterization of a hemin-storage locus of Yersinia pestis. Biol Met 4:41–47
     [Google Scholar]
  59. Pendrak M. L., Perry R. D. 1993; Proteins essential for expression of the Hms+ phenotype of Yersinia pestis. Mol Microbiol 8:857–864
     [Google Scholar]
  60. Perry R. D., Fetherston J. D. 1997; Yersinia pestis – etiologic agent of plague. Clin Microbiol Rev 10:35–66
     [Google Scholar]
  61. Perry R. D., Pendrak M. L., Schuetze P. 1990; Identification and cloning of a hemin storage locus involved in the pigmentation phenotype of Yersinia pestis. J Bacteriol 172:5929–5937
     [Google Scholar]
  62. Perry R. D., Bobrov A. G., Kirillina O., Jones H. A., Pedersen L. L., Abney J., Fetherston J. D. 2004; Temperature regulation of the hemin storage (Hms+) phenotype of Yersinia pestis is posttranscriptional. J Bacteriol 186:1638–1647
     [Google Scholar]
  63. Prossnitz E., Nikaido K., Ulbrich S. J., Ames G. F. 1988; Formaldehyde and photoactivatable cross-linking of the periplasmic binding protein to a membrane component of the histidine transport system of Salmonella typhimurium. J Biol Chem 263:17917–17920
     [Google Scholar]
  64. Reed L. J., Muench H. 1938; A simple method of estimating fifty percent endpoints. Am J Hyg 27:493–497
     [Google Scholar]
  65. Simm R., Fetherston J. D., Kader A., Romling U., Perry R. D. 2005; Phenotypic convergence mediated by GGDEF-domain-containing proteins. J Bacteriol 187:6816–6823
     [Google Scholar]
  66. Skare J. T., Ahmer B. M., Seachord C. L., Darveau R. P., Postle K. 1993; Energy transduction between membranes. TonB, a cytoplasmic membrane protein, can be chemically cross-linked in vivo to the outer membrane receptor FepA. J Biol Chem 268:16302–16308
     [Google Scholar]
  67. Stoodley P., Sauer K., Davies D. G., Costerton J. W. 2002; Biofilms as complex differentiated communities. Annu Rev Microbiol 56:187–209
     [Google Scholar]
  68. Straley S. C., Bowmer W. S. 1986; Virulence genes regulated at the transcriptional level by Ca2+ in Yersinia pestis include structural genes for outer membrane proteins. Infect Immun 51:445–454
     [Google Scholar]
  69. Surgalla M. J., Beesley E. D. 1969; Congo red-agar plating medium for detecting pigmentation in Pasteurella pestis. Appl Microbiol 18:834–837
     [Google Scholar]
  70. Sutherland I. 2001; Biofilm exopolysaccharides: a strong and sticky framework. Microbiology 147:3–9
     [Google Scholar]
  71. Sutherland B. W., Toews J., Kast J. 2008; Utility of formaldehyde cross-linking and mass spectrometry in the study of protein-protein interactions. J Mass Spectrom 43:699–715
     [Google Scholar]
  72. Towbin H., Staehelin T., Gordon J. 1979; Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A 76:4350–4354
     [Google Scholar]
  73. Van Houdt R., Michiels C. W. 2005; Role of bacterial cell surface structures in Escherichia coli biofilm formation. Res Microbiol 156:626–633
     [Google Scholar]
  74. Vasilescu J., Guo X., Kast J. 2004; Identification of protein-protein interactions using in vivo cross-linking and mass spectrometry. Proteomics 4:3845–3854
     [Google Scholar]
  75. Vuong C., Kocianova S., Voyich J. M., Yao Y., Fischer E. R., DeLeo F. R., Otto M. 2004; A crucial role for exopolysaccharide modification in bacterial biofilm formation, immune evasion, and virulence. J Biol Chem 279:54881–54886
     [Google Scholar]
  76. Wang X., Preston J. F. III, Romeo T. 2004; The pgaABCD locus of Escherichia coli promotes the synthesis of a polysaccharide adhesin required for biofilm formation. J Bacteriol 186:2724–2734
     [Google Scholar]
  77. Warren M. A., Kucharski L. M., Veenstra A., Shi L., Grulich P. F., Maguire M. E. 2004; The CorA Mg2+ transporter is a homotetramer. J Bacteriol 186:4605–4612
     [Google Scholar]
  78. Wilder A. P., Eisen R. J., Bearden S. W., Montenieri J. A., Gage K. L., Antolin M. F. 2008; Oropsylla hirsuta (Siphonaptera: Ceratophyllidae) can support plague epizootics in black-tailed prairie dogs ( Cynomys ludovicianus) by early-phase transmission of Yersinia pestis. Vector Borne Zoonotic Dis 8:359–367
     [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/mic.0.036640-0
Loading
Analysis of HmsH and its role in plague biofilm formation
Microbiology 156, 1424 (2010); https://doi.org/10.1099/mic.0.036640-0
/content/journal/micro/10.1099/mic.0.036640-0
/content/journal/micro/10.1099/mic.0.036640-0
Loading

Data & Media loading...

Supplements

Supplementary material 1

PDF

Supplementary material 2

PDF

Supplementary material 3

PDF

Most cited Most Cited RSS feed

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