1887

Abstract

Biofilm formation in the human intestinal pathogen is in part regulated by norspermidine, spermidine and spermine. senses these polyamines through a signalling pathway consisting of the periplasmic protein, NspS, and the integral membrane c-di-GMP phosphodiesterase MbaA. NspS and MbaA belong to a proposed class of novel signalling systems composed of periplasmic ligand-binding proteins and membrane-bound c-di-GMP phosphodiesterases containing both GGDEF and EAL domains. In this signal transduction pathway, NspS is hypothesized to interact with MbaA in the periplasm to regulate its phosphodiesterase activity. Polyamine binding to NspS likely alters this interaction, leading to the activation or inhibition of biofilm formation depending on the polyamine. The purpose of this study was to determine the amino acids important for NspS function. We performed random mutagenesis of the gene, identified mutant clones deficient in biofilm formation, determined their responsiveness to norspermidine and mapped the location of these residues onto NspS homology models. Single mutants clustered on two lobes of the NspS model, but the majority were found on a single lobe that appeared to be more mobile upon norspermidine binding. We also identified residues in the putative ligand-binding site that may be important for norspermidine binding and interactions with MbaA. Ultimately, our results provide new insights into this novel signalling pathway in and highlight differences between periplasmic binding proteins involved in transport versus signal transduction.

Funding
This study was supported by the:
  • National Institute of Allergy and Infectious Diseases (Award R15AI096358)
    • Principle Award Recipient: ECEKARATAN
  • National Institute of General Medical Sciences (Award R35GM133506)
    • Principle Award Recipient: MistyL Kuhn
Loading

Article metrics loading...

/content/journal/micro/10.1099/mic.0.001023
2021-01-27
2021-10-16
Loading full text...

Full text loading...

References

  1. Ali M, Nelson AR, Lopez AL, Sack DA. Updated global burden of cholera in endemic countries. PLoS Negl Trop Dis 2015; 9:e0003832 [View Article]
    [Google Scholar]
  2. Kierek K, Watnick PI. Environmental determinants of Vibrio cholerae biofilm development. Appl Environ Microbiol 2003; 69:5079–5088 [View Article]
    [Google Scholar]
  3. Colwell RR, Spira WM. The ecology of Vibrio cholerae . In Barua D, Greenough WB. (editors) Cholera. Current Topics in Infectious Disease Boston, MA: Springer; 1992 pp 107–127
    [Google Scholar]
  4. Kaper JB, Morris JG, Levine MM. Cholera. Clin Microbiol Rev 1995; 8:48–86 [View Article]
    [Google Scholar]
  5. Wernick NLB, Chinnapen DJ-F, Cho JA, Lencer WI. Cholera toxin: an intracellular journey into the cytosol by way of the endoplasmic reticulum. Toxins 2010; 2:310–325 [View Article]
    [Google Scholar]
  6. Herrington DA, Hall RH, Losonsky G, Mekalanos JJ, Taylor RK et al. Toxin, toxin-coregulated pili, and the toxR regulon are essential for Vibrio cholerae pathogenesis in humans. J Exp Med 1988; 168:1487–1492 [View Article]
    [Google Scholar]
  7. Faruque SM, Biswas K, Udden SMN, Ahmad QS, Sack DA et al. Transmissibility of cholera: in vivo-formed biofilms and their relationship to infectivity and persistence in the environment. Proc Natl Acad Sci U S A 2006; 103:6350–6355 [View Article][PubMed]
    [Google Scholar]
  8. Richardson K. Roles of motility and flagellar structure in pathogenicity of Vibrio cholerae: analysis of motility mutants in three animal models. Infect Immun 1991; 59:2727–2736 [View Article]
    [Google Scholar]
  9. Watnick PI, Kolter R. Steps in the development of a Vibrio cholerae El Tor biofilm. Mol Microbiol 1999; 34:586–595 [View Article]
    [Google Scholar]
  10. Zhu J, Mekalanos JJ. Quorum sensing-dependent biofilms enhance colonization in Vibrio cholerae . Dev Cell 2003; 5:647–656 [View Article]
    [Google Scholar]
  11. Costerton JW, Cheng KJ, Geesey GG, Ladd TI, Nickel JC et al. Bacterial biofilms in nature and disease. Annu Rev Microbiol 1987; 41:435–464 [View Article]
    [Google Scholar]
  12. Costerton JW, Lewandowski Z, Caldwell DE, Korber DR, Lappin-Scott HM. Microbial biofilms. Annu Rev Microbiol 1995; 49:711–745 [View Article]
    [Google Scholar]
  13. Tischler AD, Camilli A. Cyclic diguanylate (c-di-GMP) regulates Vibrio cholerae biofilm formation. Mol Microbiol 2004; 53:857–869 [View Article]
    [Google Scholar]
  14. Romling 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 [View Article]
    [Google Scholar]
  15. Römling U, Gomelsky M, Galperin MY. C-di-GMP: the dawning of a novel bacterial signalling system. Mol Microbiol 2005; 57:629–639 [View Article]
    [Google Scholar]
  16. Beyhan S, Odell LS, Yildiz FH. Identification and characterization of cyclic diguanylate signaling systems controlling rugosity in Vibrio cholerae . J Bacteriol 2008; 190:7392–7405 [View Article]
    [Google Scholar]
  17. Karatan E, Watnick P. Signals, regulatory networks, and materials that build and break bacterial biofilms. Microbiol Mol Biol Rev 2009; 73:310–347 [View Article][PubMed]
    [Google Scholar]
  18. Tabor CW, Tabor H. Polyamines. Annu Rev Biochem 1984; 53:749–790 [View Article]
    [Google Scholar]
  19. Tabor CW, Tabor H. Polyamines in microorganisms. Microbiol Rev 1985; 49:81–99 [View Article]
    [Google Scholar]
  20. Igarashi K, Kashiwagi K. Polyamines: mysterious modulators of cellular functions. Biochem Biophys Res Commun 2000; 271:559–564 [View Article]
    [Google Scholar]
  21. Michael AJ. Polyamine function in archaea and bacteria. J Biol Chem 2018; 293:18693–18701 [View Article]
    [Google Scholar]
  22. Karatan E, Duncan TR, Watnick PI. NspS, a predicted polyamine sensor, mediates activation of Vibrio cholerae biofilm formation by norspermidine. J Bacteriol 2005; 187:7434–7443 [View Article]
    [Google Scholar]
  23. McGinnis MW, Parker ZM, Walter NE, Rutkovsky AC, Cartaya-Marin C et al. Spermidine regulates Vibrio cholerae biofilm formation via transport and signaling pathways. FEMS Microbiol Lett 2009; 299:166–174 [View Article]
    [Google Scholar]
  24. Cockerell SR, Rutkovsky AC, Zayner JP, Cooper RE, Porter LR et al. Vibrio cholerae NspS, a homologue of ABC-type periplasmic solute binding proteins, facilitates transduction of polyamine signals independent of their transport. Microbiology 2014; 160:832–843 [View Article]
    [Google Scholar]
  25. Sobe RC, Bond WG, Wotanis CK, Zayner JP, Burriss MA et al. Spermine inhibits Vibrio cholerae biofilm formation through the NspS–MbaA polyamine signaling system. J Biol Chem 2017; 292:17025–17036 [View Article]
    [Google Scholar]
  26. Zappia V, Porta R, Cartenì-Farina M, De Rosa M, Gambacorta A. Polyamine distribution in eukaryotes: occurrence of sym-nor-spermidine and sym-nor-spermine in arthropods. FEBS Lett 1978; 94:161–165 [View Article]
    [Google Scholar]
  27. Yamamoto S, Shinoda S, Kawaguchi M, Wakamatsu K, Makita M. Polyamine distribution in Vibrionaceae : norspermidine as a general constituent of Vibrio species. Can J Microbiol 1983; 29:724–728 [View Article]
    [Google Scholar]
  28. Hamana K, Matsuzaki S. Widespread occurrence of norspermidine and norspermine in eukaryotic algae. J Biochem 1982; 91:1321–1328 [View Article]
    [Google Scholar]
  29. Stillway LW, Walle T. Identification of the unusual polyamines 3,3′-diaminodipropylamine and n,n′-bis(3-aminopropyl)-1,3-propanediamine in the white shrimp Penaeus setiferus. Biochem Biophys Res Commun 1977; 77:1103–1107 [View Article]
    [Google Scholar]
  30. Michael AJ. Polyamines in eukaryotes, bacteria, and archaea. J Biol Chem 2016; 291:14896–14903 [View Article]
    [Google Scholar]
  31. Milovic V. Polyamines in the gut lumen: bioavailability and biodistribution. Eur J Gastroen Hepatol 2001; 13:1021–1025 [View Article]
    [Google Scholar]
  32. Murphy GM. Polyamines in the human gut. Eur J Gastroen Hepat 2001; 13:1011–1014 [View Article]
    [Google Scholar]
  33. Matsumoto M, Kibe R, Ooga T, Aiba Y, Kurihara S et al. Impact of intestinal microbiota on intestinal luminal metabolome. Sci Rep 2012; 2:233 [View Article]
    [Google Scholar]
  34. Bomchil N, Watnick P, Kolter R. Identification and characterization of a Vibrio cholerae gene, mbaA, involved in maintenance of biofilm architecture. J Bacteriol 2003; 185:1384–1390 [View Article]
    [Google Scholar]
  35. Davidson AL, Dassa E, Orelle C, Chen J. Structure, function, and evolution of bacterial ATP-binding cassette systems. Microbiol Mol Biol Rev 2008; 72:317–364 [View Article]
    [Google Scholar]
  36. Sugiyama S, Matsuo Y, Vassylyev DG, Matsushima M, Morikawa K et al. The 1.8-Å X-ray structure of the Escherichia coli PotD protein complexed with spermidine and the mechanism of polyamine binding. Protein Sci 1996; 5:1984–1990 [View Article]
    [Google Scholar]
  37. Waldor MK, Mekalanos JJ. Emergence of a new cholera pandemic: molecular analysis of virulence determinants in Vibrio cholerae 0139 and development of a live vaccine prototype. J Infect Dis 1994; 170:278–283 [View Article]
    [Google Scholar]
  38. O'Toole GA. Microtiter dish biofilm formation assay. J Vis Exp 2011; 47: [View Article]
    [Google Scholar]
  39. Waterhouse A, Bertoni M, Bienert S, Studer G, Tauriello G et al. SWISS-MODEL: homology modelling of protein structures and complexes. Nucleic Acids Res 2018; 46:W296–W303 [View Article]
    [Google Scholar]
  40. Wu D, Lim SC, Dong Y, Wu J, Tao F et al. Structural basis of substrate binding specificity revealed by the crystal structures of polyamine receptors SpuD and SpuE from Pseudomonas aeruginosa. J Mol Biol 2012; 416:697–712 [View Article]
    [Google Scholar]
  41. Almagro Armenteros JJ, Tsirigos KD, Sønderby CK, Petersen TN, Winther O et al. SignalP 5.0 improves signal peptide predictions using deep neural networks. Nat Biotechnol 2019; 37:420–423 [View Article]
    [Google Scholar]
  42. Pei J, Kim B-H, Grishin NV. PROMALS3D: a tool for multiple protein sequence and structure alignments. Nucleic Acids Res 2008; 36:2295–2300 [View Article]
    [Google Scholar]
  43. Morris GM, Huey R, Lindstrom W, Sanner MF, Belew RK et al. AutoDock4 and AutoDockTools4: automated docking with selective receptor flexibility. J Comput Chem 2009; 30:2785–2791 [View Article]
    [Google Scholar]
  44. Dolinsky TJ, Nielsen JE, McCammon JA, Baker NA. PDB2PQR: an automated pipeline for the setup of Poisson-Boltzmann electrostatics calculations. Nucleic Acids Res 2004; 32:W665–W667 [View Article]
    [Google Scholar]
  45. Scrutton NS, Raine ARC. Cation-pi bonding and amino-aromatic interactions in the biomolecular recognition of substituted ammonium ligands. Biochem J 1996; 319:1–8 [View Article]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/mic.0.001023
Loading
/content/journal/micro/10.1099/mic.0.001023
Loading

Data & Media loading...

Supplements

Supplementary material 1

PDF

Most cited this month 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