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Volume 156, Issue 4

Mutations in that confer streptomycin resistance show pleiotropic effects on virulence and the production of a carbapenem antibiotic in Free

Abstract

Spontaneous streptomycin-resistant derivatives of subsp. strain ATTn10 were isolated. Sequencing of the locus (encoding the ribosomal protein S12) showed that each mutant was missense, with a single base change, resulting in the substitution of the wild-type lysine by arginine, threonine or asparagine at codon 43. Phenotypic analyses showed that the mutants could be segregated into two groups: K43R mutants showed reduced production of the -lactam secondary metabolite 1-carbapen-2-em-3 carboxylic acid (Car), but little effect on exoenzyme production or virulence in potato tuber tests. By contrast, the K43N and K43T mutations were pleiotropic, resulting in reduced exoenzyme production and virulence, as well as diminished Car production. The effect on Car production was due to reduced transcription of the quorum-sensing-dependent biosynthetic genes. The effects of K43N and K43T mutations on Car production were partially alleviated by provision of an excess of the quorum-sensing signalling molecule -(3-oxohexanoyl)--homoserine lactone. Finally, a proteomic analysis of the K43T mutant indicated that the abundance of a subset of intracellular proteins was affected by this mutation.

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Keyword(s): 3-oxo-C6-HSL, N-(3-oxohexanoyl)-l-homoserine lactone , Car, 1-carbapen-2-em-3 carboxylic acid , DiGE, difference gel electrophoresis , Eca, Erwinia carotovora subsp. atroseptica , Ecc, Erwinia carotovora subsp. carotovora and QS, quorum sensing
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2010-04-01
2024-04-25
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References

  1. Ames B. N., Martin R. G., Garry B. J. 1961; The first step of histidine biosynthesis. J Biol Chem 236:2019–2026
     [Google Scholar]
  2. Bainton N. J., Stead P., Chhabra S. R., Bycroft B. W., Salmond G. P. C., Stewart G. S. A. B., Williams P. 1992a; N-(3-Oxohexanoyl)-l-homoserine lactone regulates carbapenem antibiotic production in Erwinia carotovora. Biochem J 288:997–1004
     [Google Scholar]
  3. Bainton N. J., Bycroft B. W., Chhabra S. R., Stead P., Gledhill L., Hill P. J., Rees C. E. D., Winson M. K., Salmond G. P. C. other authors 1992b; A general role for the lux autoinducer in bacterial cell signalling: control of antibiotic synthesis in Erwinia. Gene 116:87–91
     [Google Scholar]
  4. Barnard A. M. L., Salmond G. P. C. 2005; Quorum sensing: the complexities of chemical communication between bacteria. Complexus 2:87–101
     [Google Scholar]
  5. Bell K. S., Sebaihia M., Pritchard L., Holden M. T., Hyman L. J., Holeva M. C., Thomson N. R., Bentley S. D., Churcher L. J. other authors 2004; Genome sequence of the enterobacterial phytopathogen Erwinia carotovora subsp. atroseptica and characterization of virulence factors. Proc Natl Acad Sci U S A 101:11105–11110
     [Google Scholar]
  6. Björkman J., Hughes D., Andersson D. I. 1998; Virulence of antibiotic-resistant Salmonella typhimurium. Proc Natl Acad Sci U S A 95:3949–3953
     [Google Scholar]
  7. Burr T., Barnard A. M. L., Corbett M. J., Pemberton C. L., Simpson N. J. L., Salmond G. P. C. 2006; Identification of the central quorum sensing regulator of virulence in the enteric phytopathogen, Erwinia carotovora: the VirR repressor. Mol Microbiol 59:113–125
     [Google Scholar]
  8. Carter A. P., Clemons W. M., Brodersen D. E., Morgan-Warren R. J., Wimberly B. T., Ramakrishnan V. 2000; Functional insights from the structure of the 30S ribosomal subunit and its interactions with antibiotics. Nature 407:340–348
     [Google Scholar]
  9. Coulthurst S. J., Lilley K. S., Salmond G. P. C. 2006; Genetic and proteomic analysis of the role of luxS in the enteric phytopathogen, Erwinia carotovora. Mol Plant Pathol 7:31–45
     [Google Scholar]
  10. Cox A. R., Thomson N. R., Bycroft B. W., Stewart G. S., Williams P., Salmond G. P. 1998; A pheromone-independent CarR protein controls carbapenem antibiotic synthesis in the opportunistic human pathogen Serratia marcescens. Microbiology 144:201–209
     [Google Scholar]
  11. Gregory S. T., Cate J. H., Dahlberg A. E. 2001; Streptomycin-resistant and streptomycin-dependent mutants of the extreme thermophile Thermus thermophilus. J Mol Biol 309:333–338
     [Google Scholar]
  12. Holden M. T. G., McGowan S. J., Bycroft B. W., Stewart G. S. A. B., Williams P., Salmond G. P. C. 1998; Cryptic carbapenem antibiotic production genes are widespread in Erwinia carotovora: facile trans activation by the carR transcriptional regulator. Microbiology 144:1495–1508
     [Google Scholar]
  13. Hosaka T., Xu J., Ochi K. 2006; Increased expression of ribosome recycling factor is responsible for the enhanced protein synthesis during the late growth phase in an antibiotic-overproducing Streptomyces coelicolor ribosomal rpsL mutant. Mol Microbiol 61:883–897
     [Google Scholar]
  14. Hu H., Ochi K. 2001; Novel approach for improving the productivity of antibiotic-producing strains by inducing combined resistant mutations. Appl Environ Microbiol 67:1885–1892
     [Google Scholar]
  15. Jones S. J., Yu B., Bainton N. J., Birdsall M., Bycroft B. W., Chhabra S. R., Cox A. R. J., Golby P., Reeves P. J. other authors 1993; The lux autoinducer regulates the production of exoenzyme virulence determinants in Erwinia carotovora and Pseudomonas aeruginosa. EMBO J 12:2477–2482
     [Google Scholar]
  16. Kornder J. D. 2002; Streptomycin revisited: molecular action in the microbial cell. Med Hypotheses 58:34–46
     [Google Scholar]
  17. Kurosawa K., Hosaka T., Tamehiro N., Inaoka T., Ochi K. 2006; Improvement of α-amlyase production by modulation of ribosomal component protein S12 in Bacillus subtilis 168. Appl Environ Microbiol 72:71–77
     [Google Scholar]
  18. Laemmli U. K. 1970; Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685
     [Google Scholar]
  19. McGowan S. J., Sebaihia M., Jones S., Yu B., Bainton N., Chan P. F., Bycroft B., Stewart G. S. A. B., Williams P., Salmond G. P. C. 1995; Carbapenem antibiotic production in Ewinia carotovora is regulated by CarR, a homologue of the LuxR transcriptional activator. Microbiology 141:541–550
     [Google Scholar]
  20. McGowan S. J., Sebaihia M., Porter L. E., Stewart G. S. A. B., Williams P., Bycroft B. W., Salmond G. P. C. 1996; Analysis of bacterial carbapenem antibiotic production genes reveals a novel β-lactam biosynthesis pathway. Mol Microbiol 22:415–426
     [Google Scholar]
  21. McGowan S. J., Sebaihia M., O'Leary S., Hardie K. R., Williams P., Stewart G. S. A. B., Bycroft B. W., Salmond G. P. C. 1997; Analysis of the carbapenem gene cluster of Erwinia carotovora: definition of the antibiotic biosynthetic genes and evidence for a novel β-lactam resistance mechanism. Mol Microbiol 26:545–556
     [Google Scholar]
  22. McGowan S. J., Barnard A. M. L., Bosgelmez G., Sebaihia M., Simpson N. J. L., Thomson N. R., Todd D. E., Welch M., Whitehead N. A., Salmond G. P. C. 2005; Carbapenem antibiotic biosynthesis in Erwinia carotovora is regulated by physiological and genetic factors modulating the quorum sensing-dependent control pathway. Mol Microbiol 55:526–545
     [Google Scholar]
  23. Nair J., Rouse D. A., Bai G.-H., Morris S. L. 1993; The rpsL gene and streptomcyin resistance in single and multiple drug-resistant strains of Mycobacterium tuberculosis. Mol Microbiol 10:521–527
     [Google Scholar]
  24. Okamoto-Hosoya Y., Hosaka T., Ochi K. 2003a; An aberrant protein synthesis activity is linked with antibiotic overproduction in rpsL mutants of Streptomyces coelicolor A3(2. Microbiology 149:3299–3309
     [Google Scholar]
  25. Okamoto-Hosoya Y., Okamoto S., Ochi K. 2003b; Development of antibiotic-overproducing strains by site-directed mutagenesis of the rpsL gene in Streptomyces lividans. Appl Environ Microbiol 69:4256–4259
     [Google Scholar]
  26. Rivet M. M. 1998 I nvestigation into the regulation of exoenzyme production in Erwinia carotovora subspecies carotovora PhD thesis University of; Warwick, UK:
     [Google Scholar]
  27. Sambrook J., Fritsch E. F., Maniatis T. 1989 Molecular Cloning: a Laboratory Manual, 2nd edn. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
  28. Shima J., Hesketh A., Okamoto S., Kawamoto S., Ochi K. 1996; Induction of actinorhodin production by rpsL (encoding ribosomal protein S12) mutations that confer streptomycin resistance in Streptomyces lividans and Streptomyces coelicolor A3(2. J Bacteriol 178:7276–7284
     [Google Scholar]
  29. Sreevatsan S., Pan X., Stockbauer K. E., Williams D. L., Kreiswirth B. N., Musser J. M. 1996; Characterization of rpsL and rrs mutants in streptomycin-resistant Mycobacterium tuberculosis isolates from diverse geographic localities. Antimicrob Agents Chemother 40:1024–1026
     [Google Scholar]
  30. Thomson N. R., Cox A., Bycroft B. W., Stewart G. S. A. B., Williams P., Salmond G. P. C. 1997; The Rap and Hor proteins of Erwinia, Serratia and Yersinia: a novel subgroup in a growing superfamily of proteins regulating diverse physiological processes in bacterial pathogens. Mol Microbiol 26:531–544
     [Google Scholar]
  31. Toth I. K., Birch P. R. J. 2005; Rotting softly and stealthily. Curr Opin Plant Biol 8:424–429
     [Google Scholar]
  32. Toth I., Perombelon M., Salmond G. 1993; Bacteriophage φKP mediated generalized transduction in Erwinia carotovora subspecies carotovora. J Gen Microbiol 139:2705–2709
     [Google Scholar]
  33. Voll M. J., Appella E., Martin R. G. 1967; Purification and composition studies of phosphoribosyladenosine triphosphate: pyrophosphate phosphoribosyltransferase, the first enzyme of histidine biosynthesis. J Biol Chem 242:1760–1767
     [Google Scholar]
  34. Waters C. M., Bassler B. L. 2005; Quorum sensing: cell-to-cell communication in bacteria. Annu Rev Cell Dev Biol 21:319–346
     [Google Scholar]
  35. Whitehead N. A., Barnard A. M. L., Slater H., Simpson N. J. L., Salmond G. P. C. 2001; Quorum-sensing in Gram-negative bacteria. FEMS Microbiol Rev 25:365–404
     [Google Scholar]
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Mutations in rpsL that confer streptomycin resistance show pleiotropic effects on virulence and the production of a carbapenem antibiotic in Erwinia carotovora
Microbiology 156, 1030 (2010); https://doi.org/10.1099/mic.0.034595-0
/content/journal/micro/10.1099/mic.0.034595-0
/content/journal/micro/10.1099/mic.0.034595-0
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