1887

Abstract

The switch from a motile, planktonic existence to an attached biofilm is a major bacterial lifestyle transition that is often mediated by complex regulatory pathways. In this report, we describe a CheY-like protein required for control of the motile-to-sessile switch in the plant pathogen Agrobacterium tumefaciens. This regulator, which we have designated ClaR, possesses two distinct CheY-like receiver (REC) domains and is involved in the negative regulation of biofilm formation, through production of the unipolar polysaccharide (UPP) adhesin and cellulose. The ClaR REC domains share predicted structural homology with characterized REC domains and contain the majority of active site residues known to be essential for protein phosphorylation. REC1 is missing the conserved aspartate (N72) residue and although present in REC 2 (D193), it is not required for ClaR-dependent regulation suggesting that phosphorylation, which modulates the activity of many CheY-like proteins, appears not to be essential for ClaR activity. We also show that ClaR-dependent negative regulation of attachment is diminished significantly in mutants for PruA and PruR, proteins known to be involved in a pterin-mediated attachment regulation pathway. In A. tumefaciens, pterins are required for control of the intracellular signal cyclic diguanylate monophosphate through the DcpA regulator, but our findings suggest that pterin-dependent ClaR control of attachment can function independently from DcpA, including dampening of c-di-GMP levels. This report of a novel CheY-type biofilm regulator in A. tumefaciens thus also adds significant details to the role of pterin-mediated signalling.

Loading

Article metrics loading...

/content/journal/micro/10.1099/mic.0.000558
2017-10-25
2019-10-20
Loading full text...

Full text loading...

/deliver/fulltext/micro/163/11/1680.html?itemId=/content/journal/micro/10.1099/mic.0.000558&mimeType=html&fmt=ahah

References

  1. Hall-Stoodley L, Costerton JW, Stoodley P. Bacterial biofilms: from the natural environment to infectious diseases. Nat Rev Microbiol 2004; 2: 95– 108 [CrossRef] [PubMed]
    [Google Scholar]
  2. Flemming HC, Wingender J. The biofilm matrix. Nat Rev Microbiol 2010; 8: 623– 633 [CrossRef] [PubMed]
    [Google Scholar]
  3. Mckew BA, Taylor JD, Mcgenity TJ, Underwood GJ. Resistance and resilience of benthic biofilm communities from a temperate saltmarsh to desiccation and rewetting. ISME J 2011; 5: 30– 41 [CrossRef] [PubMed]
    [Google Scholar]
  4. Anderl JN, Franklin MJ, Stewart PS. Role of antibiotic penetration limitation in Klebsiella pneumoniae biofilm resistance to ampicillin and ciprofloxacin. Antimicrob Agents Chemother 2000; 44: 1818– 1824 [CrossRef] [PubMed]
    [Google Scholar]
  5. Mah TF, O'Toole GA. Mechanisms of biofilm resistance to antimicrobial agents. Trends Microbiol 2001; 9: 34– 39 [CrossRef] [PubMed]
    [Google Scholar]
  6. Houry A, Gohar M, Deschamps J, Tischenko E, Aymerich S et al. Bacterial swimmers that infiltrate and take over the biofilm matrix. Proc Natl Acad Sci USA 2012; 109: 13088– 13093 [CrossRef] [PubMed]
    [Google Scholar]
  7. Marks LR, Reddinger RM, Hakansson AP. High levels of genetic recombination during nasopharyngeal carriage and biofilm formation in Streptococcus pneumoniae. MBio 2012; 3: e00200-12 [CrossRef] [PubMed]
    [Google Scholar]
  8. Marks LR, Mashburn-Warren L, Federle MJ, Hakansson AP. Streptococcus pyogenes biofilm growth in vitro and in vivo and its role in colonization, virulence, and genetic exchange. J Infect Dis 2014; 210: 25– 34 [CrossRef] [PubMed]
    [Google Scholar]
  9. del Pozo JL, Patel R. The challenge of treating biofilm-associated bacterial infections. Clin Pharmacol Ther 2007; 82: 204– 209 [CrossRef] [PubMed]
    [Google Scholar]
  10. van Larebeke N, Engler G, Holsters M, van den Elsacker S, Zaenen I et al. Large plasmid in Agrobacterium tumefaciens essential for crown gall-inducing ability. Nature 1974; 252: 169– 170 [CrossRef] [PubMed]
    [Google Scholar]
  11. Watson B, Currier TC, Gordon MP, Chilton MD, Nester EW. Plasmid required for virulence of Agrobacterium tumefaciens. J Bacteriol 1975; 123: 255– 264 [PubMed]
    [Google Scholar]
  12. Escobar MA, Dandekar AM. Agrobacterium tumefaciens as an agent of disease. Trends Plant Sci 2003; 8: 380– 386 [CrossRef] [PubMed]
    [Google Scholar]
  13. Ramey BE, Matthysse AG, Fuqua C. The FNR-type transcriptional regulator SinR controls maturation of Agrobacterium tumefaciens biofilms. Mol Microbiol 2004; 52: 1495– 1511 [CrossRef] [PubMed]
    [Google Scholar]
  14. Abarca-Grau AM, Penyalver R, López MM, Marco-Noales E. Pathogenic and non-pathogenic Agrobacterium tumefaciens, A. rhizogenes and A. vitis strains form biofilms on abiotic as well as on root surfaces. Plant Pathol 2011; 60: 416– 425 [CrossRef]
    [Google Scholar]
  15. Heindl JE, Wang Y, Heckel BC, Mohari B, Feirer N et al. Mechanisms and regulation of surface interactions and biofilm formation in Agrobacterium. Front Plant Sci 2014; 5: 176 [CrossRef] [PubMed]
    [Google Scholar]
  16. Tomlinson AD, Fuqua C. Mechanisms and regulation of polar surface attachment in Agrobacterium tumefaciens. Curr Opin Microbiol 2009; 12: 708– 714 [CrossRef] [PubMed]
    [Google Scholar]
  17. Li G, Brown PJ, Tang JX, Xu J, Quardokus EM et al. Surface contact stimulates the just-in-time deployment of bacterial adhesins. Mol Microbiol 2012; 83: 41– 51 [CrossRef] [PubMed]
    [Google Scholar]
  18. Xu J, Kim J, Danhorn T, Merritt PM, Fuqua C. Phosphorus limitation increases attachment in Agrobacterium tumefaciens and reveals a conditional functional redundancy in adhesin biosynthesis. Res Microbiol 2012; 163: 674– 684 [CrossRef] [PubMed]
    [Google Scholar]
  19. Matthysse AG, Holmes KV, Gurlitz RH. Elaboration of cellulose fibrils by Agrobacterium tumefaciens during attachment to carrot cells. J Bacteriol 1981; 145: 583– 595 [PubMed]
    [Google Scholar]
  20. Matthysse AG, Marry M, Krall L, Kaye M, Ramey BE et al. The effect of cellulose overproduction on binding and biofilm formation on roots by Agrobacterium tumefaciens. Mol Plant Microbe Interact 2005; 18: 1002– 1010 [CrossRef] [PubMed]
    [Google Scholar]
  21. Tomlinson AD, Ramey-Hartung B, Day TW, Merritt PM, Fuqua C. Agrobacterium tumefaciens ExoR represses succinoglycan biosynthesis and is required for biofilm formation and motility. Microbiology 2010; 156: 2670– 2681 [CrossRef] [PubMed]
    [Google Scholar]
  22. Heckel BC, Tomlinson AD, Morton ER, Choi JH, Fuqua C. Agrobacterium tumefaciens exoR controls acid response genes and impacts exopolysaccharide synthesis, horizontal gene transfer, and virulence gene expression. J Bacteriol 2014; 196: 3221– 3233 [CrossRef] [PubMed]
    [Google Scholar]
  23. Heindl JE, Hibbing ME, Xu J, Natarajan R, Buechlein AM et al. Discrete responses to limitation for Iron and Manganese in Agrobacterium tumefaciens: influence on attachment and biofilm formation. J Bacteriol 2015; 198: 816– 829 [CrossRef] [PubMed]
    [Google Scholar]
  24. Merritt PM, Danhorn T, Fuqua C. Motility and chemotaxis in Agrobacterium tumefaciens surface attachment and biofilm formation. J Bacteriol 2007; 189: 8005– 8014 [CrossRef] [PubMed]
    [Google Scholar]
  25. Xu J, Kim J, Koestler BJ, Choi JH, Waters CM et al. Genetic analysis of Agrobacterium tumefaciens unipolar polysaccharide production reveals complex integrated control of the motile-to-sessile switch. Mol Microbiol 2013; 89: 929– 948 [CrossRef] [PubMed]
    [Google Scholar]
  26. Schirmer T, Jenal U. Structural and mechanistic determinants of c-di-GMP signalling. Nat Rev Microbiol 2009; 7: 724– 735 [CrossRef] [PubMed]
    [Google Scholar]
  27. Römling U, Galperin MY, Gomelsky M. Cyclic di-GMP: the first 25 years of a universal bacterial second messenger. Microbiol Mol Biol Rev 2013; 77: 1– 52 [CrossRef] [PubMed]
    [Google Scholar]
  28. Sondermann H, Shikuma NJ, Yildiz FH. You've come a long way: c-di-GMP signaling. Curr Opin Microbiol 2012; 15: 140– 146 [CrossRef] [PubMed]
    [Google Scholar]
  29. Amikam D, Benziman M. Cyclic diguanylic acid and cellulose synthesis in Agrobacterium tumefaciens. J Bacteriol 1989; 171: 6649– 6655 [CrossRef] [PubMed]
    [Google Scholar]
  30. Feirer N, Xu J, Allen KD, Koestler BJ, Bruger EL et al. A pterin-dependent signaling pathway regulates a dual-function diguanylate cyclase-phosphodiesterase controlling surface attachment in Agrobacterium tumefaciens. MBio 2015; 6: e00156-15 [CrossRef] [PubMed]
    [Google Scholar]
  31. Howie AJ, Brewer DB. Optical properties of amyloid stained by Congo red: history and mechanisms. Micron 2009; 40: 285– 301 [CrossRef] [PubMed]
    [Google Scholar]
  32. Morton ER, Fuqua C. Genetic manipulation of Agrobacterium. Curr Protoc Microbiol 2012; Chapter 3: Unit 3D 2
    [Google Scholar]
  33. Tempé J, Petit A, Holsters M, Montagu M, Schell J. Thermosensitive step associated with transfer of the Ti plasmid during conjugation: possible relation to transformation in crown gall. Proc Natl Acad Sci USA 1977; 74: 2848– 2849 [CrossRef] [PubMed]
    [Google Scholar]
  34. Khan SR, Gaines J, Roop RM, Farrand SK. Broad-host-range expression vectors with tightly regulated promoters and their use to examine the influence of TraR and TraM expression on Ti plasmid quorum sensing. Appl Environ Microbiol 2008; 74: 5053– 5062 [CrossRef] [PubMed]
    [Google Scholar]
  35. Morton ER, Fuqua C. Phenotypic analyses of Agrobacterium. Curr Protoc Microbiol 2012; Chapter 3: Unit 3D [CrossRef] [PubMed]
    [Google Scholar]
  36. Massie JP, Reynolds EL, Koestler BJ, Cong JP, Agostoni M et al. Quantification of high-specificity cyclic diguanylate signaling. Proc Natl Acad Sci USA 2012; 109: 12746– 12751 [CrossRef] [PubMed]
    [Google Scholar]
  37. Stock AM, Robinson VL, Goudreau PN. Two-component signal transduction. Annu Rev Biochem 2000; 69: 183– 215 [CrossRef] [PubMed]
    [Google Scholar]
  38. Wadhams GH, Armitage JP. Making sense of it all: bacterial chemotaxis. Nat Rev Mol Cell Biol 2004; 5: 1024– 1037 [CrossRef] [PubMed]
    [Google Scholar]
  39. Li J, Swanson RV, Simon MI, Weis RM. The response regulators CheB and CheY exhibit competitive binding to the kinase CheA. Biochemistry 1995; 34: 14626– 14636 [CrossRef] [PubMed]
    [Google Scholar]
  40. Welch M, Oosawa K, Aizawa S, Eisenbach M. Phosphorylation-dependent binding of a signal molecule to the flagellar switch of bacteria. Proc Natl Acad Sci USA 1993; 90: 8787– 8791 [CrossRef] [PubMed]
    [Google Scholar]
  41. Volz K, Matsumura P. Crystal structure of Escherichia coli CheY refined at 1.7-A resolution. J Biol Chem 1991; 266: 15511– 15519 [PubMed]
    [Google Scholar]
  42. Stock AM, Mottonen JM, Stock JB, Schutt CE. Three-dimensional structure of CheY, the response regulator of bacterial chemotaxis. Nature 1989; 337: 745– 749 [CrossRef] [PubMed]
    [Google Scholar]
  43. Volz K. Structural conservation in the CheY superfamily. Biochemistry 1993; 32: 11741– 11753 [CrossRef] [PubMed]
    [Google Scholar]
  44. Sanders DA, Gillece-Castro BL, Stock AM, Burlingame AL, Koshland DE. Identification of the site of phosphorylation of the chemotaxis response regulator protein, CheY. J Biol Chem 1989; 264: 21770– 21778 [PubMed]
    [Google Scholar]
  45. Lukat GS, Stock AM, Stock JB. Divalent metal ion binding to the CheY protein and its significance to phosphotransfer in bacterial chemotaxis. Biochemistry 1990; 29: 5436– 5442 [CrossRef] [PubMed]
    [Google Scholar]
  46. Lee SY, Cho HS, Pelton JG, Yan D, Henderson RK et al. Crystal structure of an activated response regulator bound to its target. Nat Struct Biol 2001; 8: 789– 794 [CrossRef] [PubMed]
    [Google Scholar]
  47. Lee SY, Cho HS, Pelton JG, Yan D, Berry EA et al. Crystal structure of activated CheY. Comparison with other activated receiver domains. J Biol Chem 2001; 276: 16425– 16431 [CrossRef] [PubMed]
    [Google Scholar]
  48. Kelley LA, Mezulis S, Yates CM, Wass MN, Sternberg MJ. The Phyre2 web portal for protein modeling, prediction and analysis. Nat Protoc 2015; 10: 845– 858 [CrossRef] [PubMed]
    [Google Scholar]
  49. Wright EL, Deakin WJ, Shaw CH. A chemotaxis cluster from Agrobacterium tumefaciens. Gene 1998; 220: 83– 89 [CrossRef] [PubMed]
    [Google Scholar]
  50. Harighi B. Role of CheY1 and CheY2 in the chemotaxis of A. tumefaciens toward acetosyringone. Curr Microbiol 2008; 56: 547– 552 [CrossRef] [PubMed]
    [Google Scholar]
  51. Sourjik V, Schmitt R. Different roles of CheY1 and CheY2 in the chemotaxis of Rhizobium meliloti. Mol Microbiol 1996; 22: 427– 436 [CrossRef] [PubMed]
    [Google Scholar]
  52. Sourjik V, Schmitt R. Phosphotransfer between CheA, CheY1, and CheY2 in the chemotaxis signal transduction chain of Rhizobium meliloti. Biochemistry 1998; 37: 2327– 2335 [CrossRef] [PubMed]
    [Google Scholar]
  53. Aldridge P, Jenal U. Cell cycle-dependent degradation of a flagellar motor component requires a novel-type response regulator. Mol Microbiol 1999; 32: 379– 391 [CrossRef] [PubMed]
    [Google Scholar]
  54. Paul R, Weiser S, Amiot NC, Chan C, Schirmer T et al. Cell cycle-dependent dynamic localization of a bacterial response regulator with a novel di-guanylate cyclase output domain. Genes Dev 2004; 18: 715– 727 [CrossRef] [PubMed]
    [Google Scholar]
  55. Chan C, Paul R, Samoray D, Amiot NC, Giese B et al. Structural basis of activity and allosteric control of diguanylate cyclase. Proc Natl Acad Sci USA 2004; 101: 17084– 17089 [CrossRef] [PubMed]
    [Google Scholar]
  56. Curtis PD, Brun YV. Getting in the loop: regulation of development in Caulobacter crescentus. Microbiol Mol Biol Rev 2010; 74: 13– 41 [CrossRef] [PubMed]
    [Google Scholar]
  57. Barnhart DM, Su S, Baccaro BE, Banta LM, Farrand SK. CelR, an ortholog of the diguanylate cyclase PleD of Caulobacter, regulates cellulose synthesis in Agrobacterium tumefaciens. Appl Environ Microbiol 2013; 79: 7188– 7202 [CrossRef] [PubMed]
    [Google Scholar]
  58. Kim J, Heindl JE, Fuqua C. Coordination of division and development influences complex multicellular behavior in Agrobacterium tumefaciens. PLoS One 2013; 8: e56682 [CrossRef] [PubMed]
    [Google Scholar]
  59. Cho HS, Lee SY, Yan D, Pan X, Parkinson JS et al. NMR structure of activated CheY. J Mol Biol 2000; 297: 543– 551 [CrossRef] [PubMed]
    [Google Scholar]
  60. Villali J, Pontiggia F, Clarkson MW, Hagan MF, Kern D. Evidence against the "Y-T coupling" mechanism of activation in the response regulator NtrC. J Mol Biol 2014; 426: 1554– 1567 [CrossRef] [PubMed]
    [Google Scholar]
  61. Desai SK, Kenney LJ. To approximately P or Not to approximately P? Non-canonical activation by two-component response regulators. Mol Microbiol 2017; 103: 203– 213 [Crossref]
    [Google Scholar]
  62. Dahl MK, Msadek T, Kunst F, Rapoport G. The phosphorylation state of the DegU response regulator acts as a molecular switch allowing either degradative enzyme synthesis or expression of genetic competence in Bacillus subtilis. J Biol Chem 1992; 267: 14509– 14514 [PubMed]
    [Google Scholar]
  63. Kobayashi K. Gradual activation of the response regulator DegU controls serial expression of genes for flagellum formation and biofilm formation in Bacillus subtilis. Mol Microbiol 2007; 66: 395– 409 [CrossRef] [PubMed]
    [Google Scholar]
  64. Lin W, Wang Y, Han X, Zhang Z, Wang C et al. Atypical OmpR/PhoB subfamily response regulator GlnR of actinomycetes functions as a homodimer, stabilized by the unphosphorylated conserved Asp-focused charge interactions. J Biol Chem 2014; 289: 15413– 15425 [CrossRef] [PubMed]
    [Google Scholar]
  65. Wuichet K, Zhulin IB. Origins and diversification of a complex signal transduction system in prokaryotes. Sci Signal 2010; 3: ra50 [CrossRef] [PubMed]
    [Google Scholar]
  66. He K, Bauer CE. Chemosensory signaling systems that control bacterial survival. Trends Microbiol 2014; 22: 389– 398 [CrossRef] [PubMed]
    [Google Scholar]
  67. Kirby JR, Zusman DR. Chemosensory regulation of developmental gene expression in Myxococcus xanthus. Proc Natl Acad Sci USA 2003; 100: 2008– 2013 [CrossRef] [PubMed]
    [Google Scholar]
  68. Black WP, Yang Z. Myxococcus xanthus chemotaxis homologs DifD and DifG negatively regulate fibril polysaccharide production. J Bacteriol 2004; 186: 1001– 1008 [CrossRef] [PubMed]
    [Google Scholar]
  69. Berleman JE, Bauer CE. A che-like signal transduction cascade involved in controlling flagella biosynthesis in Rhodospirillum centenum. Mol Microbiol 2005; 55: 1390– 1402 [CrossRef] [PubMed]
    [Google Scholar]
  70. Berleman JE, Bauer CE. Involvement of a Che-like signal transduction cascade in regulating cyst cell development in Rhodospirillum centenum. Mol Microbiol 2005; 56: 1457– 1466 [CrossRef] [PubMed]
    [Google Scholar]
  71. D'Argenio DA, Calfee MW, Rainey PB, Pesci EC. Autolysis and autoaggregation in Pseudomonas aeruginosa colony morphology mutants. J Bacteriol 2002; 184: 6481– 6489 [CrossRef] [PubMed]
    [Google Scholar]
  72. Hickman JW, Tifrea DF, Harwood CS. A chemosensory system that regulates biofilm formation through modulation of cyclic diguanylate levels. Proc Natl Acad Sci USA 2005; 102: 14422– 14427 [CrossRef] [PubMed]
    [Google Scholar]
  73. Güvener ZT, Harwood CS. Subcellular location characteristics of the Pseudomonas aeruginosa GGDEF protein, WspR, indicate that it produces cyclic-di-GMP in response to growth on surfaces. Mol Microbiol 2007; 66: 1459– 1473 [CrossRef] [PubMed]
    [Google Scholar]
  74. Huangyutitham V, Güvener ZT, Harwood CS. Subcellular clustering of the phosphorylated WspR response regulator protein stimulates its diguanylate cyclase activity. MBio 2013; 4: e00242-13 [CrossRef] [PubMed]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/mic.0.000558
Loading
/content/journal/micro/10.1099/mic.0.000558
Loading

Data & Media loading...

Supplements

Supplementary File 1

PDF
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