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Genes & Genomics

, Volume 41, Issue 1, pp 43–59 | Cite as

Spermine and oxacillin stress response on the cell wall synthesis and the global gene expression analysis in Methicillin-resistance Staphylococcus aureus

  • Shrikant Pawar
  • Xiangyu Yao
  • Chung-Dar LuEmail author
Research Article

Abstract

Methicillin-resistant Staphylococcus aureus (MRSA) is a rapidly emerging bacteria causing infection, which has developed resistance to most of the beta-lactam antibiotics because of newly acquired low-affinity penicillin-binding protein (PBP2a), which can continue to build the cell wall when beta-lactams block other PBPs. Exogenous spermine exerts a dose-dependent inhibition effect on the growth of Escherichia coli, Salmonella enterica serovar, and S. aureus. Selection of an MRSA Mu50 derivative which harbors mutation on PBP2 gene (named as MuM) showing spermine resistance and which confers a complete abolishment of spermine-beta-lactam synergy was identified. To further investigate the gene expression changes, a transcriptome profiling of MuM against Mu50 (wild-type) without any treatment, MuM and Mu50 in response to high dose spermine and Mu50 in response to spermine-beta-lactam synergy at 15, 30 and 60 min time points was performed. Functional annotation was further performed to delineate the metabolic pathways associated with the significant genes. A significant down-regulation in the iron regulatory system, potassium channel uptake and polyamine transport system with an up-regulation in general stress response sigB dependent operon in MuM strain at 15, 30 and 60 min time points with spermine treatment compared to Mu50 strain was observed. Analysis of spermine-dependent synergy with beta-lactams on cell wall synthesis revealed that it significantly reduces the degree of cross-linkage on cell wall with no change in trypsin digestion pattern of purified PBPs and without affecting PBPs expression or PBPs acylation by Bocillin. A strong relation between PBP2 protein and general stress sigB response, iron, potassium and polyamine transport systems was observed. SigB regulon should be activated on stress, which was not seen in some of our previous studies where it was down-regulated in wild-type Mu50 strain with spermine stress. Here, an intriguing finding is made where there seems to be a correction of this abnormal response of no SigB induction to a significant induction by PBP2 mutation. In MuM strain, a significant down-regulation of KdpABC operon genes at 15, 30 and 60 min time points on spermine stress is seen, which seems to be absent without spermine treatment. Since KCL has been found to protect the cell against spermine stress in wild-type strain by induction of KdpABC operon, it fails to do so in MuM strain underlying the importance of PBP2 protein in spermine stress. Analysis of spermine-dependent synergy with beta-lactams on cell wall synthesis revealed that it significantly reduces the degree of cross-linkage on cell wall with no change in trypsin digestion patterns of purified PBPs and without affecting PBPs expression or PBPs acylation by Bocillin. Furthermore, spermine does not help in enhancing the binding of beta-lactams to PBPs and binding of spermine to PBPs does not cause conformational changes to PBPs, as tested with trypsin digestion patterns. Future studies on the molecular mechanism of spermine interactions with these systems hold great potential for the development of new therapeutics for MRSA infections.

Keywords

Staphylococcus aureus Microarrays Spermine Oxacillin Penicillin binding protein (PBPs) 

Notes

Acknowledgements

This work was supported by National Science Foundation Grant (NSF0950217) to CD Lu from Georgia State University.

Author contributions

C-DL conceived and designed the study; SP carried out data and statistical analysis with assistance in the writing of the manuscript; XY carried out the isolation of spermine resistant mutants, complementation assay and microarray transcriptional profiling. All authors read and approved the final manuscript.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interests.

References

  1. Antignac A, Sieradzki K, Tomasz A (2007) Perturbation of cell wall synthesis suppresses autolysis in Staphylococcus aureus: evidence for coregulation of cell wall synthetic and hydrolytic enzymes. J Bacteriol 189:7573–7580CrossRefGoogle Scholar
  2. Ashraf M, Ong S-K, Mujawar S, Pawar S, More P, Paul S, Lahiri C (2018) A side-effect free method for identifying cancer drug targets. Sci Rep.  https://doi.org/10.1038/s41598-018-25042-2 Google Scholar
  3. Berger-Bächi B, Tschierske M (1998) Role of fem factors in methicillin resistance. Drug Resist Update 1:325–335CrossRefGoogle Scholar
  4. Bischoff M, Dunman P, Kormanec J, Macapagal D (2004) Microarray-based analysis of the Staphylococcus aureus sigmab regulon. J Bacteriol 186:4085–4099CrossRefGoogle Scholar
  5. Chakraborty R, Braun V, Hantke K, Cornelis P (2013) Iron uptake in bacteria with emphasis on E. coli and Pseudomonas, Springer briefs in bio metals. Springer, DordrechtCrossRefGoogle Scholar
  6. Chan PF, Foster SJ, Ingham E, Clements MO (1998) The Staphylococcus aureus alternative sigma factor sigmab controls the environmental stress response but not starvation survival or pathogenicity in a mouse abscess model. J Bacteriol 180:6082–6089Google Scholar
  7. Cimolai N (2008) MRSA and the environment: implications for comprehensive control measures. Eur J Clin Microbiol Infect Dis 27:481–493CrossRefGoogle Scholar
  8. Dalman M, Deeter A, Duan ZH (2012) Fold change and p-value cutoffs significantly alter microarray interpretations. BMC Bioinform 13:11CrossRefGoogle Scholar
  9. de Jonge BL, Tomasz A (1993) Abnormal peptidoglycan produced in a methicillin-resistant strain of Staphylococcus aureus grown in the presence of methicillin: functional role for penicillin-binding protein 2a in cell wall synthesis. Antimicrob Agents Chemother 37:342–346CrossRefGoogle Scholar
  10. De Jonge BL, Chang YS, Xu N, Gage D (1996) Effect of exogenous glycine on peptidoglycan composition and resistance in a methicillin-resistant Staphylococcus aureus strain. Antimicrob Agents Chemother 40, 1498–1503CrossRefGoogle Scholar
  11. Dong-Hyeon K, Lu CD (2007) Polyamine effects on antibiotic susceptibility in bacteria. Antimicrob Agents Chemother 51:2070–2077CrossRefGoogle Scholar
  12. Enright MC, Robinson DA, Randle G, Feil EJ (2002) The evolutionary history of methicillin-resistant Staphylococcus aureus. Proc Natl Acad Sci 99:7687–7692CrossRefGoogle Scholar
  13. Epstein W (2003) The roles and regulation of potassium in bacteria. Prog Nucleic Acid Res Mol Biol 75:293–320CrossRefGoogle Scholar
  14. Foster T (1996) Medical microbiology. University of Texas Medical Branch at Galveston, GalvestonGoogle Scholar
  15. Ghuysen JM (1991) Serine beta-lactamases and penicillin-binding proteins. Annu Rev Microbiol 45:37–67CrossRefGoogle Scholar
  16. Gregory R, Warnes G (2009) gplots: various R programming tools for plotting data. R package versionGoogle Scholar
  17. Grossowicz R, Razin NS (1955) Factors influencing the antibacterial action of spermine and spermidine on Staphylococcus aureus. J Gen Microbiol 13:436–441CrossRefGoogle Scholar
  18. Haag H, Fiedler HP, Meiwes J, Drechsel H (1994) Isolation and biological characterization of staphyloferrin b, a compound with siderophore activity from Staphylococci. FEMS Microbiol Lett 115:125–130CrossRefGoogle Scholar
  19. Hartman BJ, Tomasz A (1984) Low-affinity penicillin-binding protein associated with beta-lactam resistance in Staphylococcus aureus. J Bacteriol 158:513–516Google Scholar
  20. Hecker M (2001) General stress response of Bacillus subtilis and other bacteria. Adv Microb Physiol 44:35–91CrossRefGoogle Scholar
  21. Hecker M, Pane-Farre UV (2007) Sigb-dependent general stress response in Bacillus subtilis and related gram-positive bacteria. Annu Rev Microbiol 61:215–236CrossRefGoogle Scholar
  22. Heermann R (2010) The complexity of the’simple’ two-component system kdpd/kdpe in Escherichia coli. FEMS Microbiol Lett 304:97–106CrossRefGoogle Scholar
  23. Huang da W, Sherman BT (2009) Bioinformatics enrichment tools: paths toward the comprehensive functional analysis of large gene lists. Nucleic Acids Res 37:1–13CrossRefGoogle Scholar
  24. Hyeon-Gyu S, Zhe L (2005) Mechanism of the voltage sensitivity of irk1 inward-rectifier k + channel block by the polyamine spermine. J Gen Physiol 125:413–426CrossRefGoogle Scholar
  25. Kohanski MA, Dwyer DJ, Hayete B, Lawrence CA (2007) A common mechanism of cellular death induced by bactericidal antibiotics. Cell 130:797–810CrossRefGoogle Scholar
  26. Kohanski M, Dwyer D, Collins J (2010) How antibiotics kill bacteria: from targets to networks. Nat Rev Microbiol 8:423–435CrossRefGoogle Scholar
  27. Kuwahara-Arai K, Kondo N, Hori S, Tateda-Suzuki E (1996) Suppression of methicillin resistance in a meca-containing pre-methicillin-resistant Staphylococcus aureus strain is caused by the meci-mediated repression of pbp 2′ production. Antimicrob Agents Chemother 40:2680–2685CrossRefGoogle Scholar
  28. Lahiri C, Shrikant P, Sabarinathan R, Ashraf M, Chakravortty D (2012) Identifying indispensable proteins of the type III secretion systems of Salmonella enterica serovar Typhimurium strain LT2. BMC Bioinform 13(Suppl 12):A10CrossRefGoogle Scholar
  29. Lahiri C, Pawar S, Sabarinathan R, Ashraf MI, Chand Y, Chakravortty D (2014) Interactome analyses of Salmonella pathogenicity islands reveal SicA indispensable for virulence. J Theor Biol 363:188–197CrossRefGoogle Scholar
  30. Lomozik L, Jastrzab R (2003) Copper(ii) complexes with uridine, uridine 5′-monophosphate, spermidine, or spermine in aqueous solution. J Inorg Biochem 93:132–140CrossRefGoogle Scholar
  31. Mademidis A (1998) Transport activity of fhua, fhuc, fhud, and fhub derivatives in a system free of polar effects, and stoichiometry of components involved in ferrichrome uptake. Mol Gen Genet 258:156–165CrossRefGoogle Scholar
  32. Marquardt JL, Siegele DA, Kolter R, Walsh CT (1992) Cloning and sequencing of Escherichia coli murz and purification of its product, a udp-n-acetylglucosamine enolpyruvyl transferase. J Bacteriol 174:5748–5752CrossRefGoogle Scholar
  33. Marzabadi MR (1996) Spermine prevent iron accumulation and depress lipofuscin accumulation in cultured myocardial cells. Free Radic Biol Med 21:375–381CrossRefGoogle Scholar
  34. Meiwes J, Fiedler HP, Haag H, Zähner H (1990) Isolation and characterization of staphyloferrin a, a compound with siderophore activity from Staphylococcus hyicus dsm 20459. FEMS Microbiol Lett 55:201–205CrossRefGoogle Scholar
  35. Mittal K, Choi DH, Klimov S, Pawar S, Kaur R, Mitra AK, Gupta MV, Sams R, Cantuaria G, Rida PCG, Aneja R (2016a) A centrosome clustering protein, KIFC1, predicts aggressive disease course in serous ovarian adenocarcinomas. J Ovarian Res 9:17CrossRefGoogle Scholar
  36. Mittal K, Choi DH, Klimov S, Pawar S, Kaur R, Mitra A, Gupta MV, Sams R, Cantuaria G, Rida PCG, Aneja R (2016b) Evaluation of centrosome clustering protein KIFC1 as a potential prognostic biomarker in serous ovarian adenocarcinomas. J Clin Oncol 34(15_suppl):e17083–e17083CrossRefGoogle Scholar
  37. Pawar S, Donthamsetty S, Pannu V, Rida P, Ogden A, Bowen N, Osan R, Cantuaria G, Aneja R (2014) KIFCI, a novel putative prognostic biomarker for ovarian adenocarcinomas: delineating protein interaction networks and signaling circuitries. J Ovarian Res 7:53CrossRefGoogle Scholar
  38. Pawar S, Ashraf M, Mehata K, Lahiri C (2017) Computational identification of indispensable virulence proteins of Salmonella Typhi CT18. Curr Top Salmonella Salmonellosis.  https://doi.org/10.5772/66489 Google Scholar
  39. Pawar S, Ashraf M, Mujawar S, Mishra R, Lahiri C (2018) In silico Identification of the indispensable quorum sensing proteins of multidrug resistant Proteus mirabilis. Front Cell Infect Microbiol 8:269CrossRefGoogle Scholar
  40. Pawar S, Davis C, Rinehart C (2011) Statistical analysis of microarray gene expression data from a mouse model of toxoplasmosis. BMC Bioinform 12(Suppl 7):A19CrossRefGoogle Scholar
  41. Petersohn A, Brigulla M, Haas S, Hoheisel JD (2001) Global analysis of the general stress response of Bacillus subtilis. J Bacteriol 183:5617–5631CrossRefGoogle Scholar
  42. Pinho MG, Lencastre H (1998) Transcriptional analysis of the Staphylococcus aureus penicillin binding protein 2 gene. J Bacteriol 180:6077–6081Google Scholar
  43. Pinho MG, de Lencastre H, Tomasz A (2001) An acquired and a native penicillin-binding protein cooperate in building the cell wall of drug-resistant Staphylococci. Proc Natl Acad Sci 98:10886–10891CrossRefGoogle Scholar
  44. Pootoolal J, Neu WJ (2002) Glycopeptide antibiotic resistance. Annu Rev Pharmacol Toxicol 42:381–408CrossRefGoogle Scholar
  45. Qamar A (2012) Dual roles of fmta in Staphylococcus aureus cell wall biosynthesis and autolysis. Antimicrob Agents Chemother 56:3797–3805CrossRefGoogle Scholar
  46. Rogers MB, Sexton J, DeCastro J, Calderwood S (2000) Identification of an operon required for ferrichrome iron utilization in Vibrio cholerae. J Bacteriol 182:2350–2353CrossRefGoogle Scholar
  47. Shah P (2008) A multifaceted role for polyamines in bacterial pathogens. Mol Microbiol 68:4–16CrossRefGoogle Scholar
  48. Utaida S, Dunman PM, Macapagal D, Murphy E (2003) Genome-wide transcriptional profiling of the response of Staphylococcus aureus to cell-wall-active antibiotics reveals a cell-wall-stress stimulon. Microbiology 149:2719–2732CrossRefGoogle Scholar
  49. Vollmer W, Joris B, Charlier P, Foster S (2008) Bacterial peptidoglycan (murein) hydrolases. FEMS Microbiol Rev 32:259–286CrossRefGoogle Scholar
  50. Yao X, Lu CD (2012) A pbp 2 mutant devoid of the transpeptidase domain abolishes spermine-beta-lactam synergy in Staphylococcus aureus mu50. Antimicrob Agents Chemother 56:83–91CrossRefGoogle Scholar
  51. Yao X, Lu CD (2014) Characterization of Staphylococcus aureus responses to spermine stress. Curr Microbiol 69:394–403CrossRefGoogle Scholar

Copyright information

© The Genetics Society of Korea and Springer Nature B.V. 2018

Authors and Affiliations

  1. 1.Department of Computer ScienceGeorgia State UniversityAtlantaUSA
  2. 2.Department of BiologyGeorgia State UniversityAtlantaUSA
  3. 3.National Institutes of HealthBethesdaUSA
  4. 4.Department of Clinical Laboratory and Nutritional SciencesUniversity of MassachusettsLowellUSA

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