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Applied Microbiology and Biotechnology

, Volume 104, Issue 3, pp 1259–1271 | Cite as

OxyR-controlled surface polysaccharide production and biofilm formation in Acinetobacter oleivorans DR1

  • Bora Shin
  • Chulwoo Park
  • Woojun ParkEmail author
Applied microbial and cell physiology
  • 124 Downloads

Abstract

The genomes of several Acinetobacter species possess three distinct polysaccharide-producing operons [two poly-N-acetyl glucosamine (PNAG) and one K-locus]. Using a microfluidic device, an increased amount of polysaccharides and enhanced biofilm formation were observed following continuous exposure to H2O2 and removal of the H2O2-sensing key regulator, OxyR, in Acinetobacter oleivorans DR1 cells. Gene expression analysis revealed that genes located in PNAG1, but not those in PNAG2, were induced and that genes in the K-locus were expressed in the presence of H2O2. Interestingly, the expression of the K-locus gene was enhanced in the PNAG1 mutant and vice versa. The absence of either OxyR or PNAG1 resulted in enhanced biofilm formation, higher surface hydrophobicity, and increased motility, implying that K-locus-driven polysaccharide production in both the oxyR and PNAG1 deletion mutants may be related to these phenotypes. Both the oxyR and K-locus deletion mutants were more sensitive to H2O2 compared with the wildtype and PNAG1 mutant strains. Purified OxyR binds to the promoter regions of both polysaccharide operons with a higher affinity toward the K-locus promoter. Although oxidized OxyR could bind to both promoter regions, the addition of dithiothreitol further enhanced the binding efficiency of OxyR, suggesting that OxyR might function as a repressor for controlling these polysaccharide operons.

Keywords

Acinetobacter OxyR Exopolysaccharides Poly-N-acetyl glucosamine Capsular polysaccharides Biofilm 

Notes

Acknowledgements

This work was supported by grants from the National Research Foundation of Korea (No. NRF-2019R1A2C1088452).

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical statement

This article does not contain any studies with human participants or animals performed by any of the authors.

Supplementary material

253_2019_10303_MOESM1_ESM.pdf (1.4 mb)
ESM 1 (PDF 1443 kb)

References

  1. Arakawa Y, Wacharotayankun R, Nagatsuka T, Ito H, Kato N, Ohta M (1995) Genomic organization of the Klebsiella pneumoniae cps region responsible for serotype K2 capsular polysaccharide synthesis in the virulent strain Chedid. J Bacteriol 177(7):1788–1796.  https://doi.org/10.1128/jb.177.7.1788-1796.1995 CrossRefPubMedPubMedCentralGoogle Scholar
  2. Aslund F, Zheng M, Beckwith J, Storz G (1999) Regulation of the OxyR transcription factor by hydrogen peroxide and the cellular thiol-disulfide status. Proc Natl Acad Sci U S A 96(11):6161–6165.  https://doi.org/10.1073/pnas.96.11.6161 CrossRefPubMedPubMedCentralGoogle Scholar
  3. Chen H, Xu G, Zhao Y, Tian B, Lu H, Yu X, Xu Z, Ying N, Hu S, Hua Y (2008) A novel OxyR sensor and regulator of hydrogen peroxide stress with one cysteine residue in Deinococcus radiodurans. PLoS One 3(2):e1602.  https://doi.org/10.1371/journal.pone.0001602 CrossRefPubMedPubMedCentralGoogle Scholar
  4. Cywes-Bentley C, Skurnik D, Zaidi T, Roux D, Deoliveira RB, Garrett WS, Lu X, O'Malley J, Kinzel K, Zaidi T, Rey A, Perrin C, Fichorova RN, Kayatani AK, Maira-Litran T, Gening ML, Tsvetkov YE, Nifantiev NE, Bakaletz LO, Pelton SI, Golenbock DT, Pier GB (2013) Antibody to a conserved antigenic target is protective against diverse prokaryotic and eukaryotic pathogens. Proc Natl Acad Sci U S A 110(24):2209–2218.  https://doi.org/10.1073/pnas.1303573110 CrossRefGoogle Scholar
  5. Derouiche A, Shi L, Kalantari A, Mijakovic I (2016) Substrate specificity of the Bacillus subtilis BY-kinase PtkA is controlled by alternative activators: TkmA and SalA. Front Microbiol 7:1525.  https://doi.org/10.3389/fmicb.2016.01525 CrossRefPubMedPubMedCentralGoogle Scholar
  6. Doublet P, Vincent C, Grangeasse C, Cozzone AJ, Duclos B (1999) On the binding of ATP to the autophosphorylating protein, Ptk, of the bacterium Acinetobacter johnsonii. FEBS Lett 445(1):137–143.  https://doi.org/10.1016/s0014-5793(99)00111-8 CrossRefPubMedGoogle Scholar
  7. Epstein AK, Pokroy B, Seminara A, Aizenberg J (2011) Bacterial biofilm shows persistent resistance to liquid wetting and gas penetration. Proc Natl Acad Sci U S A 108(3):995–1000.  https://doi.org/10.1073/pnas.1011033108 CrossRefPubMedGoogle Scholar
  8. Fang FC (2011) Antimicrobial actions of reactive oxygen species. MBio 2(5):e00141–e00111.  https://doi.org/10.1128/mBio.00141-11 CrossRefPubMedPubMedCentralGoogle Scholar
  9. Geisinger E, Isberg RR (2015) Antibiotic modulation of capsular exopolysaccharide and virulence in Acinetobacter baumannii. PLoS Pathog 11(2):e1004691.  https://doi.org/10.1371/journal.ppat.1004691 CrossRefPubMedPubMedCentralGoogle Scholar
  10. Goller C, Wang X, Itoh Y, Romeo T (2006) The cation-responsive protein NhaR of Escherichia coli activates pgaABCD transcription, required for production of the biofilm adhesin poly-β-1,6-N-acetyl-D-glucosamine. J Bacteriol 188(23):8022–8032.  https://doi.org/10.1128/JB.01106-06 CrossRefPubMedPubMedCentralGoogle Scholar
  11. Haagensen JA, Hansen SK, Christensen BB, Pamp SJ, Molin S (2015) Development of spatial distribution patterns by biofilm cells. Appl Environ Microbiol 81(18):6120–6128.  https://doi.org/10.1128/AEM.01614-15 CrossRefPubMedPubMedCentralGoogle Scholar
  12. Hall-Stoodley L, Stoodley P (2009) Evolving concepts in biofilm infections. Cell Microbiol 11(7):1034–1043.  https://doi.org/10.1111/j.1462-5822.2009.01323.x CrossRefPubMedGoogle Scholar
  13. Harding CM, Hennon SW, Feldman MF (2018) Uncovering the mechanisms of Acinetobacter baumannii virulence. Nat Rev Microbiol 16(2):91–102.  https://doi.org/10.1038/nrmicro.2017.148 CrossRefPubMedGoogle Scholar
  14. He X, Lu F, Yuan F, Jiang D, Zhao P, Zhu J, Cheng H, Cao J, Lu G (2015) Biofilm formation caused by clinical Acinetobacter baumannii isolates is associated with overexpression of the AdeFGH efflux pump. Antimicrob Agents Chemother 59(8):4817–4825.  https://doi.org/10.1128/AAC.00877-15 CrossRefPubMedPubMedCentralGoogle Scholar
  15. Heo YJ, Chung IY, Cho WJ, Lee BY, Kim JH, Choi KH, Lee JW, Hassett DJ, Cho YH (2010) The major catalase gene (katA) of Pseudomonas aeruginosa PA14 is under both positive and negative control of the global transactivator OxyR in response to hydrogen peroxide. J Bacteriol 192(2):381–390.  https://doi.org/10.1128/JB.00980-09 CrossRefPubMedGoogle Scholar
  16. Hishinuma S, Yuki M, Fujimura M, Fukumori F (2006) OxyR regulated the expression of two major catalases, KatA and KatB, along with peroxiredoxin, AhpC in Pseudomonas putida. Environ Microbiol 8(12):2115–2124.  https://doi.org/10.1111/j.1462-2920.2006.01088.x CrossRefPubMedGoogle Scholar
  17. Hu D, Liu B, Dijkshoorn L, Wang L, Reeves PR (2013) Diversity in the major polysaccharide antigen of Acinetobacter baumannii assessed by DNA sequencing, and development of a molecular serotyping scheme. PLoS One 8(7):e70329.  https://doi.org/10.1371/journal.pone.0070329 CrossRefPubMedPubMedCentralGoogle Scholar
  18. Jang IA, Kim J, Park W (2016) Endogenous hydrogen peroxide increases biofilm formation by inducing exopolysaccharide production in Acinetobacter oleivorans DR1. Sci Rep 6:21121.  https://doi.org/10.1038/srep21121 CrossRefPubMedPubMedCentralGoogle Scholar
  19. Jordan IK, Natale DA, Koonin EV, Galperin MY (2001) Independent evolution of heavy metal-associated domains in copper chaperones and copper-transporting ATPases. J Mol Evol 53(6):622–633.  https://doi.org/10.1007/s002390010249 CrossRefPubMedGoogle Scholar
  20. Juttukonda LJ, Green ER, Lonergan ZR, Heffern MC, Chang CJ, Skaar EP (2019) Acinetobacter baumannii OxyR regulates the transcriptional response to hydrogen peroxide. Infect Immun 87(1):e00413–e00418.  https://doi.org/10.1128/IAI.00413-18 CrossRefPubMedGoogle Scholar
  21. Kenyon JJ, Hall RM (2013) Variation in the complex carbohydrate biosynthesis loci of Acinetobacter baumannii genomes. PLoS One 8(4):e62160.  https://doi.org/10.1371/journal.pone.0062160 CrossRefPubMedPubMedCentralGoogle Scholar
  22. Kim HJ, Eom HJ, Park C, Jung J, Shin B, Kim W, Chung N, Choi IG, Park W (2016) Calcium carbonate precipitation by Bacillus and Sporosarcina strains isolated from concrete and analysis of the bacterial community of concrete. J Microbiol Biotechnol 26(3):540–548.  https://doi.org/10.4014/jmb.1511.11008 CrossRefPubMedGoogle Scholar
  23. Kim J, Cho Y, Jang IA, Park W (2015) Molecular mechanism involved in the response to hydrogen peroxide stress in Acinetobacter oleivorans DR1. Appl Microbiol Biotechnol 99(24):10611–10626.  https://doi.org/10.1007/s00253-015-6914-5 CrossRefPubMedGoogle Scholar
  24. Kim J, Park W (2013) Indole inhibits bacterial quorum sensing signal transmission by interfering with quorum sensing regulator folding. Microbiology 159(Pt12):2616–2625.  https://doi.org/10.1099/mic.0.070615-0 CrossRefPubMedGoogle Scholar
  25. LeBlanc JJ, Brassinga AK, Ewann F, Davidson RJ, Hoffman PS (2008) An ortholog of OxyR in Legionella pneumophila is expressed postexponentially and negatively regulates the alkyl hydroperoxide reductase (ahpC2D) operon. J Bacteriol 190(10):3444–3455.  https://doi.org/10.1128/JB.00141-08 CrossRefPubMedPubMedCentralGoogle Scholar
  26. Little DJ, Pfoh R, Le Mauff F, Bamford NC, Notte C, Baker P, Guragain M, Robinson H, Pier GB, Nitz M, Deora R, Sheppard DC, Howell PL (2018) PgaB orthologues contain a glycoside hydrolase domain that cleaves deacetylated poly-β-(1,6)-N-acetylglucosamine and can disrupt bacterial biofilms. PLoS Pathog 14(4):e1006998.  https://doi.org/10.1371/journal.ppat.1006998 CrossRefPubMedPubMedCentralGoogle Scholar
  27. Milse J, Petri K, Ruckert C, Kalinowski J (2014) Transcriptional response of Corynebacterium glutamicum ATCC 13032 to hydrogen peroxide stress and characterization of the OxyR regulon. J Biotechnol 190:40–54.  https://doi.org/10.1016/j.jbiotec.2014.07.452 CrossRefPubMedGoogle Scholar
  28. Nagarajan S, Sherman DM, Shaw I, Sherman LA (2012) Functions of the duplicated hik31 operons in central metabolism and responses to light, dark, and carbon sources in Synechocystis sp. strain PCC 6803. J Bacteriol 194(2):448–459.  https://doi.org/10.1128/JB.06207-11 CrossRefGoogle Scholar
  29. Nakar D, Gutnick DL (2003) Involvement of a protein tyrosine kinase in production of the polymeric bioemulsifier emulsan from the oil-degrading strain Acinetobacter lwoffii RAG-1. J Bacteriol 185(3):1001–1009.  https://doi.org/10.1128/jb.185.3.1001-1009.2003 CrossRefPubMedPubMedCentralGoogle Scholar
  30. Omelchenko MV, Makarova KS, Wolf YI, Rogozin IB, Koonin EV (2003) Evolution of mosaic operons by horizontal gene transfer and gene displacement in situ. Genome Biol 4(9):55.  https://doi.org/10.1186/gb-2003-4-9-r55 CrossRefGoogle Scholar
  31. Pan YJ, Lin TL, Chen CT, Chen YY, Hsieh PF, Hsu CR, Wu MC, Wang JT (2015) Genetic analysis of capsular polysaccharide synthesis gene clusters in 79 capsular types of Klebsiella spp. Sci Rep 5:15573.  https://doi.org/10.1038/srep15573 CrossRefPubMedPubMedCentralGoogle Scholar
  32. Reid AN, Whitfield C (2005) Functional analysis of conserved gene products involved in assembly of Escherichia coli capsules and exopolysaccharides: evidence for molecular recognition between Wza and Wzc for colanic acid biosynthesis. J Bacteriol 187(15):5470–5481.  https://doi.org/10.1128/JB.187.15.5470-5481.2005 CrossRefPubMedPubMedCentralGoogle Scholar
  33. Shin B, Park C, Imlay JA, Park W (2018) 4-Hydroxybenzaldehyde sensitizes Acinetobacter baumannii to amphenicols. Appl Microbiol Biotechnol 102(5):2323–2335.  https://doi.org/10.1007/s00253-018-8791-1 CrossRefPubMedGoogle Scholar
  34. Shin B, Park W (2015) Synergistic effect of oleanolic acid on aminoglycoside antibiotics against Acinetobacter baumannii. PLoS One 10(9):e0137751.  https://doi.org/10.1371/journal.pone.0137751 CrossRefPubMedPubMedCentralGoogle Scholar
  35. Shu HY, Fung CP, Liu YM, Wu KM, Chen YT, Li LH, Liu TT, Kirby R, Tsai SF (2009) Genetic diversity of capsular polysaccharide biosynthesis in Klebsiella pneumoniae clinical isolates. Microbiology 155(Pt12):4170–4183.  https://doi.org/10.1099/mic.0.029017-0 CrossRefPubMedGoogle Scholar
  36. Steiner S, Lori C, Boehm A, Jenal U (2013) Allosteric activation of exopolysaccharide synthesis through cyclic di-GMP-stimulated protein-protein interaction. EMBO J 32(3):354–368.  https://doi.org/10.1038/emboj.2012.315 CrossRefPubMedGoogle Scholar
  37. Stewart PS, Franklin MJ (2008) Physiological heterogeneity in biofilms. Nat Rev Microbiol 6(3):199–210.  https://doi.org/10.1038/nrmicro1838 CrossRefPubMedGoogle Scholar
  38. Tartaglia LA, Storz G, Ames BN (1989) Identification and molecular analysis of oxyR-regulated promoters important for the bacterial adaptation to oxidative stress. J Mol Biol 210(4):709–719.  https://doi.org/10.1016/0022-2836(89)90104-6 CrossRefPubMedGoogle Scholar
  39. Teramoto H, Inui M, Yukawa H (2013) OxyR acts as a transcriptional repressor of hydrogen peroxide-inducible antioxidant genes in Corynebacterium glutamicum R. FEBS J 280(14):3298–3312.  https://doi.org/10.1111/febs.12312 CrossRefPubMedGoogle Scholar
  40. Tseng HJ, McEwan AG, Apicella MA, Jennings MP (2003) OxyR acts as a repressor of catalase expression in Neisseria gonorrhoeae. Infect Immun 71(1):550–556.  https://doi.org/10.1128/iai.71.1.550-556.2003 CrossRefPubMedPubMedCentralGoogle Scholar
  41. Tytgat HL, Lebeer S (2014) The sweet tooth of bacteria: common themes in bacterial glycoconjugates. Microbiol Mol Biol Rev 78(3):372–417.  https://doi.org/10.1128/MMBR.00007-14 CrossRefPubMedPubMedCentralGoogle Scholar
  42. Wan F, Kong L, Gao H (2018) Defining the binding determinants of Shewanella oneidensis OxyR: implications for the link between the contracted OxyR regulon and adaptation. J Biol Chem 293(11):4085–4096.  https://doi.org/10.1074/jbc.RA117.001530 CrossRefPubMedPubMedCentralGoogle Scholar
  43. Wang X, Dubey AK, Suzuki K, Baker CS, Babitzke P, Romeo T (2005) CsrA post-transcriptionally represses pgaABCD, responsible for synthesis of a biofilm polysaccharide adhesin of Escherichia coli. Mol Microbiol 56(6):1648–1663.  https://doi.org/10.1111/j.1365-2958.2005.04648.x CrossRefPubMedGoogle Scholar
  44. Wang Y, Andole Pannuri A, Ni D, Zhou H, Cao X, Lu X, Romeo T, Huang Y (2016) Structural basis for translocation of a biofilm-supporting exopolysaccharide across the bacterial outer membrane. J Biol Chem 291(19):10046–10057.  https://doi.org/10.1074/jbc.M115.711762 CrossRefPubMedPubMedCentralGoogle Scholar
  45. Whitfield C (2006) Biosynthesis and assembly of capsular polysaccharides in Escherichia coli. Annu Rev Biochem 75:39–68.  https://doi.org/10.1146/annurev.biochem.75.103004.142545 CrossRefPubMedGoogle Scholar
  46. Whitfield C, Paiment A (2003) Biosynthesis and assembly of group 1 capsular polysaccharides in Escherichia coli and related extracellular polysaccharides in other bacteria. Carbohydr Res 338(23):2491–2502.  https://doi.org/10.1016/j.carres.2003.08.010 CrossRefPubMedGoogle Scholar
  47. Whitney JC, Howell PL (2013) Synthase-dependent exopolysaccharide secretion in Gram-negative bacteria. Trends Microbiol 21(2):63–72.  https://doi.org/10.1016/j.tim.2012.10.001 CrossRefGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Laboratory of Molecular Environmental Microbiology, Department of Environmental Science and Ecological EngineeringKorea UniversitySeoulRepublic of Korea

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