1887

Abstract

, a plant-growth-promoting rhizobacterium, is exposed to changes in its abiotic environment, including fluctuations in temperature, salinity, osmolarity, oxygen concentration and nutrient concentration, in the rhizosphere and in the soil. Since extra-cytoplasmic function (ECF) sigma factors play an important role in stress adaptation, we analysed the role of ECF sigma factor (also known as RpoE or ) in abiotic stress tolerance in . An in-frame deletion mutant of Sp7 was carotenoidless and slow-growing, and was sensitive to salt, ethanol and methylene blue stress. Expression of in the deletion mutant complemented the defects in growth, carotenoid biosynthesis and sensitivity to different stresses. Based on data from reverse transcriptase-PCR, a two-hybrid assay and a pull-down assay, we present evidence that is cotranscribed with and the proteins synthesized from these two overlapping genes interact with each other. Identification of the transcription start site by 5′ rapid amplification of cDNA ends showed that the operon was transcribed by two promoters. The proximal promoter was less active than the distal promoter, whose consensus sequence was characteristic of RpoE-dependent promoters found in alphaproteobacteria. Whereas the proximal promoter was RpoE-independent and constitutively expressed, the distal promoter was RpoE-dependent and strongly induced in response to stationary phase and elevated levels of ethanol, salt, heat and methylene blue. This study shows the involvement of RpoE in controlling carotenoid synthesis as well as in tolerance to some abiotic stresses in , which might be critical in the adaptation, survival and proliferation of this rhizobacterium in the soil and rhizosphere under stressful conditions.

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2011-04-01
2019-10-14
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References

  1. Alba, B. M. & Gross, C. A. ( 2004; ). Regulation of the Escherichia coli sigma-dependent envelope stress response. Mol Microbiol 52, 613–619.[CrossRef]
    [Google Scholar]
  2. Alvarez-Martinez, C. E., Lourenço, R. F., Baldini, R. L., Laub, M. T. & Gomes, S. L. ( 2007; ). The ECF sigma factor σ T is involved in osmotic and oxidative stress responses in Caulobacter crescentus. Mol Microbiol 66, 1240–1255.[CrossRef]
    [Google Scholar]
  3. Anthony, J. R., Newman, J. D. & Donohue, T. J. ( 2004; ). Interactions between the Rhodobacter sphaeroides ECF sigma factor, sigma(E), and its anti-sigma factor, ChrR. J Mol Biol 341, 345–360.[CrossRef]
    [Google Scholar]
  4. Anthony, J. R., Warczak, K. L. & Donohue, T. J. ( 2005; ). A transcriptional response to singlet oxygen, a toxic byproduct of photosynthesis. Proc Natl Acad Sci U S A 102, 6502–6507.[CrossRef]
    [Google Scholar]
  5. Baldani, J. I., Krieg, N. R., Baldani, V. L. D., Hartmann, A. & Dobereiner, J. ( 1979; ). Genus II. Azospirillum Tarrand, Krieg and Dobereiner, 79AL. In Bergey's Manual of Systematic Bacteriology, vol. 2. Edited by Garrity, G. M., Brenner, D. J., Krieg, N. R. & Staley, J. T.. New York, USA. : Springer.
    [Google Scholar]
  6. Becker, L. A., Cetin, M. S., Hutkins, R. W. & Benson, A. K. ( 1998; ). Identification of the gene encoding the alternative sigma factor σ B from Listeria monocytogenes and its role in osmotolerance. J Bacteriol 180, 4547–4554.
    [Google Scholar]
  7. Benachour, A., Muller, C., Dabrowski-Coton, M., Le Breton, Y., Giard, J.-C., Rincé, A., Auffray, Y. & Hartke, A. ( 2005; ). The Enterococcus faecalis SigV protein is an extracytoplasmic function sigma factor contributing to survival following heat, acid, and ethanol treatments. J Bacteriol 187, 1022–1035.[CrossRef]
    [Google Scholar]
  8. Browning, D. F., Whitworth, D. E. & Hodgson, D. A. ( 2003; ). Light-induced carotenogenesis in Myxococcus xanthus: functional characterization of the ECF sigma factor CarQ and antisigma factor CarR. Mol Microbiol 48, 237–251.[CrossRef]
    [Google Scholar]
  9. Burger, M., Woods, R. G., McCarthy, C. & Beacham, I. R. ( 2000; ). Temperature regulation of protease in Pseudomonas fluorescens LS107d2 by an ECF sigma factor and a transmembrane activator. Microbiology 146, 3149–3155.
    [Google Scholar]
  10. Campbell, E. A., Greenwell, R., Anthony, J. R., Wang, S., Lim, L., Das, K., Sofia, H. J., Donohue, T. J. & Darst, S. A. ( 2007; ). A conserved structural module regulates transcriptional responses to diverse stress signals in bacteria. Mol Cell 27, 793–805.[CrossRef]
    [Google Scholar]
  11. da Silva Neto, J. F., Koide, T., Gomes, S. L. & Marques, M. V. ( 2007; ). The single extracytoplasmic-function sigma factor of Xylella fastidiosa is involved in the heat shock response and presents an unusual regulatory mechanism. J Bacteriol 189, 551–560.[CrossRef]
    [Google Scholar]
  12. De Las Peñas, A., Connolly, L. & Gross, C. A. ( 1997; ). The σ E-mediated response to extracytoplasmic stress in Escherichia coli is transduced by RseA and RseB, two negative regulators of σ E. Mol Microbiol 24, 373–385.[CrossRef]
    [Google Scholar]
  13. Dufour, Y. S., Landick, R. & Donohue, T. J. ( 2008; ). Organization and evolution of the biological response to singlet oxygen stress. J Mol Biol 383, 713–730.[CrossRef]
    [Google Scholar]
  14. Heimann, J. D. ( 2002; ). The extracytoplasmic function (ECF) sigma factors. Adv Microb Physiol 46, 47–110.
    [Google Scholar]
  15. Kadouri, D., Jurkevitch, E., Okon, Y. & Castro-Sowinski, S. ( 2005; ). Ecological and agricultural significance of bacterial polyhydroxyalkanoates. Crit Rev Microbiol 31, 55–67.[CrossRef]
    [Google Scholar]
  16. Karls, R. K., Wolf, J. R. & Donohue, T. J. ( 1999; ). Activation of the cycA P2 promoter for the Rhodobacter sphaeroides cytochrome c 2 gene by the photosynthesis response regulator. Mol Microbiol 34, 822–835.[CrossRef]
    [Google Scholar]
  17. Kumar, K., Tharad, M., Ganapathy, S., Ram, G., Narayan, A., Khan, J. A., Pratap, R., Ghosh, A., Samuchiwal, S. K. & other authors ( 2009; ). Phenylalanine-rich peptides potently bind ESAT6, a virulence determinant of Mycobacterium tuberculosis, and concurrently affect the pathogen's growth. PLoS ONE 4, e7615.[CrossRef]
    [Google Scholar]
  18. Lonetto, M., Gribskov, M. & Gross, C. A. ( 1992; ). The σ 70 family: sequence conservation and evolutionary relationships. J Bacteriol 174, 3843–3849.
    [Google Scholar]
  19. Miller, J. H. ( 1972; ). Experiments in Molecular Genetics. Cold Spring Harbor, NY. : Cold Spring Harbor Laboratory.
    [Google Scholar]
  20. Mishra, M. N., Thirunavukkarasu, N., Sharma, I. M., Jagnnadham, M. V. & Tripathi, A. K. ( 2008; ). Mutation in a gene encoding anti-sigma factor in A. brasilense confers tolerance to elevated temperature, antibacterial peptide and PEG-200 via carotenoid synthesis. FEMS Microbiol Lett 287, 221–229.[CrossRef]
    [Google Scholar]
  21. Newman, J. D., Falkowski, M. J., Schilke, B. A., Anthony, L. C. & Donohue, T. J. ( 1999; ). The Rhodobacter sphaeroides ECF sigma factor, σ E, and the target promoters cycA P3 and rpoE P1. J Mol Biol 294, 307–320.[CrossRef]
    [Google Scholar]
  22. Norman, R. A., Poh, C. L., Pearl, L. H., O'Hara, B. P. & Drew, R. E. ( 2000; ). Steric hindrance regulation of the Pseudomonas aeruginosa amidase operon. J Biol Chem 275, 30660–30667.[CrossRef]
    [Google Scholar]
  23. Nur, I., Steinitz, Y. L., Okon, Y. & Henis, Y. ( 1981; ). Carotenoid composition and function in nitrogen-fixing bacteria of the genus Azospirillum. J Gen Microbiol 122, 27–32.
    [Google Scholar]
  24. Schnider-Keel, U., Lejbølle, K. B., Baehler, E., Haas, D. & Keel, C. ( 2001; ). The sigma factor AlgU (AlgT) controls exopolysaccharide production and tolerance towards desiccation and osmotic stress in the biocontrol agent Pseudomonas fluorescens CHA0. Appl Environ Microbiol 67, 5683–5693.[CrossRef]
    [Google Scholar]
  25. Schurr, M. J., Yu, H., Boucher, J. C., Hibler, N. S. & Deretic, V. ( 1995; ). Multiple promoters and induction by heat shock of the gene encoding the alternative sigma factor AlgU (σ E) which controls mucoidy in cystic fibrosis isolates of Pseudomonas aeruginosa. J Bacteriol 177, 5670–5679.
    [Google Scholar]
  26. Sevciková, B., Benada, O., Kofronova, O. & Kormanec, J. ( 2001; ). Stress-response sigma factor σ H is essential for morphological differentiation of Streptomyces coelicolor A3(2). Arch Microbiol 177, 98–106.[CrossRef]
    [Google Scholar]
  27. Simon, R., Priefer, U. & Puhler, A. ( 1983; ). A broad host range mobilization system for in vivo genetic engineering: transposon mutagenesis in Gram negative bacteria. Biotechnology 1, 784–791.[CrossRef]
    [Google Scholar]
  28. Steenhoudt, O. & Vanderleyden, J. ( 2000; ). Azospirillum, a free-living nitrogen-fixing bacterium closely associated with grasses: genetic, biochemical and ecological aspects. FEMS Microbiol Rev 24, 487–506.[CrossRef]
    [Google Scholar]
  29. Tarrand, J. J., Krieg, N. R. & Döbereiner, J. ( 1978; ). A taxonomic study of the Spirillum lipoferum group, with descriptions of a new genus, Azospirillum gen. nov. and two species, Azospirillum lipoferum (Beijerinck) comb. nov. and Azospirillum brasilense sp. nov. Can J Microbiol 24, 967–980.[CrossRef]
    [Google Scholar]
  30. Thakur, K. G., Jaiswal, R. K., Shukla, J. K., Praveena, T. & Gopal, B. ( 2010; ). Over-expression and purification strategies for recombinant multi-protein oligomers: a case study of Mycobacterium tuberculosis σ/anti-σ factor protein complexes. Protein Expr Purif 74, 223–230.[CrossRef]
    [Google Scholar]
  31. Thirunavukkarasu, N., Mishra, M. N., Spaepen, S., Vanderleyden, J., Gross, C. A. & Tripathi, A. K. ( 2008; ). An extra-cytoplasmic function sigma factor and anti-sigma factor control carotenoid biosynthesis in Azospirillum brasilense. Microbiology 154, 2096–2105.[CrossRef]
    [Google Scholar]
  32. van Veen, J. A., van Overbeek, L. S. & van Elsas, J. D. ( 1997; ). Fate and activity of microorganisms introduced into soil. Microbiol Mol Biol Rev 61, 121–135.
    [Google Scholar]
  33. Vanstockem, M., Michiels, K., Vanderleyden, J. & Van Gool, A. P. ( 1987; ). Transposon mutagenesis of Azospirillum brasilense and Azospirillum lipoferum: physical analysis of Tn5 and Tn5-mob insertion mutants. Appl Environ Microbiol 53, 410–415.
    [Google Scholar]
  34. Zahran, H. H. ( 1999; ). Rhizobium-legume symbiosis and nitrogen fixation under severe conditions and in an arid climate. Microbiol Mol Biol Rev 63, 968–989.
    [Google Scholar]
  35. Ziegelhoffer, E. C. & Donohue, T. J. ( 2009; ). Bacterial responses to photo-oxidative stress. Nat Rev Microbiol 7, 856–863.
    [Google Scholar]
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vol. , part 4, pp. 988 - 999

Agarose gel showing amplicons obtained by RT-PCR with -specific primers, -specific primers, and with forward and reverse primers using total RNA extracted from late-exponential phase Sp7. Primers used in this study. [Single PDF](107 KB)



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