Extracellular DNA controls expression of genes involved in nutrient utilization, metal homeostasis, acid pH tolerance and virulence Free

Abstract

grows in extracellular DNA (eDNA)-enriched biofilms and infection sites. eDNA is generally considered to be a structural biofilm polymer required for aggregation and biofilm maturation. In addition, eDNA can sequester divalent metal cations, acidify growth media and serve as a nutrient source.

We wanted to determine the genome-wide influence on the transcriptome of planktonic PAO1 grown in the presence of eDNA.

RNA-seq analysis was performed to determine the genome-wide effects on gene expression of PAO1 grown with eDNA. Transcriptional fusions were used to confirm eDNA regulation and to validate phenotypes associated with growth in eDNA.

The transcriptome of eDNA-regulated genes included 89 induced and 76 repressed genes (FDR<0.05). A large number of eDNA-induced genes appear to be involved in utilizing DNA as a nutrient. Several eDNA-induced genes are also induced by acidic pH 5.5, and eDNA/acidic pH promoted an acid tolerance response in . The terminal oxidase is induced by both eDNA and pH 5.5, and contributed to the acid tolerance phenotype. Quantitative metal analysis confirmed that DNA binds to diverse metals, which helps explain why many genes involved in a general uptake of metals were controlled by eDNA. Growth in the presence of eDNA also promoted intracellular bacterial survival and influenced virulence in the acute infection model of fruit flies.

The diverse functions of the eDNA-regulated genes underscore the important role of this extracellular polymer in promoting antibiotic resistance, virulence, acid tolerance and nutrient utilization; phenotypes that contribute to long-term survival.

Funding
This study was supported by the:
  • Cystic Fibrosis Canada (Award na)
    • Principle Award Recipient: Shawn Lewenza
  • Cystic Fibrosis Canada (Award na)
    • Principle Award Recipient: Heidi Mulcahy-O'Grady
Loading

Article metrics loading...

/content/journal/jmm/10.1099/jmm.0.001184
2020-04-03
2024-03-28
Loading full text...

Full text loading...

/deliver/fulltext/jmm/69/6/895.html?itemId=/content/journal/jmm/10.1099/jmm.0.001184&mimeType=html&fmt=ahah

References

  1. Lewenza S. Extracellular DNA-induced antimicrobial peptide resistance mechanisms in Pseudomonas aeruginosa . Front Microbiol 2013; 4:21 [View Article][PubMed]
    [Google Scholar]
  2. Okshevsky M, Meyer RL. The role of extracellular DNA in the establishment, maintenance and perpetuation of bacterial biofilms. Crit Rev Microbiol 2015; 41:1040–1841 [View Article][PubMed]
    [Google Scholar]
  3. Halverson TWR, Wilton M, Poon KKH, Petri B, Lewenza S. DNA is an antimicrobial component of neutrophil extracellular traps. PLoS Pathog 2015; 11:e1004593 [View Article][PubMed]
    [Google Scholar]
  4. Marcos V, Zhou Z, Yildirim AO, Bohla A, Hector A et al. CXCR2 mediates NADPH oxidase-independent neutrophil extracellular trap formation in cystic fibrosis airway inflammation. Nat Med 2010; 16:1018–1023 [View Article][PubMed]
    [Google Scholar]
  5. Manzenreiter R, Kienberger F, Marcos V, Schilcher K, Krautgartner WD et al. Ultrastructural characterization of cystic fibrosis sputum using atomic force and scanning electron microscopy. J Cyst Fibros 2012; 11:84–92 [View Article][PubMed]
    [Google Scholar]
  6. Shan Q, Dwyer M, Rahman S, Gadjeva M. Distinct susceptibilities of corneal Pseudomonas aeruginosa clinical isolates to neutrophil extracellular trap-mediated immunity. Infect Immun 2014; 82:4135–4143 [View Article][PubMed]
    [Google Scholar]
  7. Jakubovics NS, Shields RC, Rajarajan N, Burgess JG. Life after death: the critical role of extracellular DNA in microbial biofilms. Lett Appl Microbiol 2013; 57:467–475 [View Article][PubMed]
    [Google Scholar]
  8. Gloag ES, Turnbull L, Huang A, Vallotton P, Wang H et al. Self-Organization of bacterial biofilms is facilitated by extracellular DNA. Proc Natl Acad Sci U S A 2013; 110:11541–11546 [View Article][PubMed]
    [Google Scholar]
  9. Mulcahy H, Charron-Mazenod L, Lewenza S. Extracellular DNA chelates cations and induces antibiotic resistance in Pseudomonas aeruginosa biofilms. PLoS Pathog 2008; 4:e1000213 [View Article][PubMed]
    [Google Scholar]
  10. Johnson L, Mulcahy H, Kanevets U, Shi Y, Lewenza S. Surface-localized spermidine protects the Pseudomonas aeruginosa outer membrane from antibiotic treatment and oxidative stress. J Bacteriol 2012; 194:813–826 [View Article][PubMed]
    [Google Scholar]
  11. Wilton M, Charron-Mazenod L, Moore R, Lewenza S. Extracellular DNA Acidifies Biofilms and Induces Aminoglycoside Resistance in Pseudomonas aeruginosa . Antimicrob Agents Chemother 2016; 60:544–553 [View Article][PubMed]
    [Google Scholar]
  12. Palchevskiy V, Finkel SE. Escherichia coli competence gene homologs are essential for competitive fitness and the use of DNA as a nutrient. J Bacteriol 2006; 188:3902–3910 [View Article][PubMed]
    [Google Scholar]
  13. Pinchuk GE, Ammons C, Culley DE, Li S-MW, McLean JS et al. Utilization of DNA as a sole source of phosphorus, carbon, and energy by Shewanella spp.: ecological and physiological implications for dissimilatory metal reduction. Appl Environ Microbiol 2008; 74:1198–1208 [View Article][PubMed]
    [Google Scholar]
  14. Mulcahy H, Charron-Mazenod L, Lewenza S. Pseudomonas aeruginosa produces an extracellular deoxyribonuclease that is required for utilization of DNA as a nutrient source. Environ Microbiol 2010; 12:1621-9 [View Article][PubMed]
    [Google Scholar]
  15. Wilton M, Halverson TWR, Charron-Mazenod L, Parkins MD, Lewenza S. Secreted Phosphatase and Deoxyribonuclease Are Required by Pseudomonas aeruginosa To Defend against Neutrophil Extracellular Traps. Infect Immun 2018; 86: 22 08 2018 [View Article][PubMed]
    [Google Scholar]
  16. Lewenza S, Falsafi RK, Winsor G, Gooderham WJ, McPhee JB et al. Construction of a mini-Tn5-luxCDABE mutant library in Pseudomonas aeruginosa PAO1: a tool for identifying differentially regulated genes. Genome Res 2005; 15:583–589 [View Article][PubMed]
    [Google Scholar]
  17. Winsor GL, Van Rossum T, Lo R, Khaira B, Whiteside MD et al. Pseudomonas genome database: facilitating user-friendly, comprehensive comparisons of microbial genomes. Nucleic Acids Res 2009; 37:D483–D488 [View Article][PubMed]
    [Google Scholar]
  18. Anders S, Huber W. Differential expression analysis for sequence count data. Genome Biol 2010; 11:R106 [View Article][PubMed]
    [Google Scholar]
  19. Lee CK, Roberts AL, Finn TM, Knapp S, Mekalanos JJ. A new assay for invasion of HeLa 229 cells by Bordetella pertussis: effects of inhibitors, phenotypic modulation, and genetic alterations. Infect Immun 1990; 58:2516–2522 [View Article][PubMed]
    [Google Scholar]
  20. Mead CG. A deoxyribonucleic acid-associated ribonucleic acid from Drosophila melanogaster. J Biol Chem 1964; 239:550–554[PubMed]
    [Google Scholar]
  21. Mulcahy H, Sibley CD, Surette MG, Lewenza S. Drosophila melanogaster as an animal model for the study of Pseudomonas aeruginosa biofilm infections in vivo. PLoS Pathog 2011; 7:e1002299 [View Article][PubMed]
    [Google Scholar]
  22. Arai H, Kawakami T, Osamura T, Hirai T, Sakai Y et al. Enzymatic characterization and in vivo function of five terminal oxidases in Pseudomonas aeruginosa . J Bacteriol 2014; 196:4206–4215 [View Article][PubMed]
    [Google Scholar]
  23. Hirai T, Osamura T, Ishii M, Arai H. Expression of multiple cbb 3 cytochrome c oxidase isoforms by combinations of multiple isosubunits in Pseudomonas aeruginosa . Proc Natl Acad Sci U S A 2016; 113:12815–12819 [View Article][PubMed]
    [Google Scholar]
  24. Jo JTH, Brinkman FSL, Hancock REW. Aminoglycoside efflux in Pseudomonas aeruginosa: involvement of novel outer membrane proteins. Antimicrob Agents Chemother 2003; 47:1101–1111 [View Article][PubMed]
    [Google Scholar]
  25. Perron K, Caille O, Rossier C, Van Delden C, Dumas J-L et al. CzcR-CzcS, a two-component system involved in heavy metal and carbapenem resistance in Pseudomonas aeruginosa . J Biol Chem 2004; 279:8761–8768 [View Article][PubMed]
    [Google Scholar]
  26. Caille O, Rossier C, Perron K. A copper-activated two-component system interacts with zinc and imipenem resistance in Pseudomonas aeruginosa . J Bacteriol 2007; 189:4561–4568 [View Article][PubMed]
    [Google Scholar]
  27. Hauser AR. The type III secretion system of Pseudomonas aeruginosa: infection by injection. Nat Rev Microbiol 2009; 7:654–665 [View Article][PubMed]
    [Google Scholar]
  28. Sana TG, Berni B, Bleves S. The T6SSs of Pseudomonas aeruginosa Strain PAO1 and Their Effectors: Beyond Bacterial-Cell Targeting. Front Cell Infect Microbiol 2016; 6:61 [View Article][PubMed]
    [Google Scholar]
  29. Wilton M, Wong MJQ, Tang L, Liang X, Moore R et al. Chelation of Membrane-Bound Cations by Extracellular DNA Activates the Type VI Secretion System in Pseudomonas aeruginosa . Infect Immun 2016; 84:2355–2361 [View Article][PubMed]
    [Google Scholar]
  30. Horsman SR, Moore RA, Lewenza S. Calcium chelation by alginate activates the type III secretion system in mucoid Pseudomonas aeruginosa biofilms. PLoS One 2012; 7:e46826 [View Article][PubMed]
    [Google Scholar]
  31. Foster JW, Hall HK. Adaptive acidification tolerance response of Salmonella typhimurium . J Bacteriol 1990; 172:771–778 [View Article][PubMed]
    [Google Scholar]
  32. Kanjee U, Houry WA. Mechanisms of acid resistance in Escherichia coli . Annu Rev Microbiol 2013; 67:65–81 [View Article][PubMed]
    [Google Scholar]
  33. Whitchurch CB, Tolker-Nielsen T, Ragas PC, Mattick JS. Extracellular DNA required for bacterial biofilm formation. Science 2002; 295:1487 [View Article][PubMed]
    [Google Scholar]
  34. Teitzel GM, Parsek MR. Heavy metal resistance of biofilm and planktonic Pseudomonas aeruginosa . Appl Environ Microbiol 2003; 69:2313–2320 [View Article][PubMed]
    [Google Scholar]
  35. Mulcahy H, Lewenza S. Magnesium limitation is an environmental trigger of the Pseudomonas aeruginosa biofilm lifestyle. PLoS One 2011; 6:e23307 [View Article][PubMed]
    [Google Scholar]
  36. Goodman AL, Kulasekara B, Rietsch A, Boyd D, Smith RS et al. A signaling network reciprocally regulates genes associated with acute infection and chronic persistence in Pseudomonas aeruginosa . Dev Cell 2004; 7:745–754 [View Article][PubMed]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/jmm/10.1099/jmm.0.001184
Loading
/content/journal/jmm/10.1099/jmm.0.001184
Loading

Data & Media loading...

Supplements

Supplementary material 1

EXCEL

Most cited Most Cited RSS feed