1887

Abstract

The stressosome is a 1.8 MDa complex that is the focal point for activating the bacterium's general response to physical stress. studies demonstrated that the stressosome's core element can be formed from one or more of a family of paralogous proteins (RsbRA, -RB, -RC and -RD) onto which the system's activator protein (RsbT) and its principal inhibitor (RsbS) are bound. The RsbR components of the stressosome are envisioned to be the initial receptors of stress signalling with the stressosome structure itself serving as a device to integrate multiple stress signals for a coordinated response. In the current work, we examine several of the characteristics of the RsbR family members, including their expression and ability to form stressosomes to regulate . Translational fusions of to each paralogue revealed that , - and - are expressed at similar levels, which remain relatively constant during growth, ethanol stress and entry into stationary phase. , in contrast, is expressed at a level that is only slightly above background during growth, but is induced to 30 % of the expression level following ethanol stress. Velocity sedimentation analyses of extracts from strains expressing single paralogues demonstrated that each incorporates RsbS into fast-sedimenting complexes. However, consistent with 's lower expression, the RsbRD-dependent RsbS complexes were present at only 20 % of the level of the complexes seen in a wild-type strain. The lower stressosome level in the RsbRD strain is still able to hold RsbT's activity in check, implying that the RsbR/S component of stressosomes is normally in excess for the control of RsbT. Consistent with such a notion, reporter gene and Western blot assays demonstrate that although RsbT is synthesized at the same rate as RsbRA and RsbS, RsbT's ultimate level in growing is only 10 % that of RsbRA. Apparently, RsbT's inherent structure and/or its passage between the stressosome and its activation target compromises its persistence.

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2010-04-01
2019-12-07
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References

  1. Akbar, S., Kang, C. M., Gaidenko, T. A. & Price, C. W. ( 1997; ). Modulator protein RsbR regulates environmental signaling in the general stress pathway of Bacillus subtilis. Mol Microbiol 24, 567–578.[CrossRef]
    [Google Scholar]
  2. Akbar, S., Gaidenko, T. A., Kang, C. M., O'Reilly, M., Devine, K. M. & Price, C. W. ( 2001; ). New family of regulators in the environmental signaling pathway which activates the general stress transcription factor σ B of Bacillus subtilis. J Bacteriol 183, 1329–1338.[CrossRef]
    [Google Scholar]
  3. Avila-Perez, M., Hellingwerf, K. J. & Kort, R. ( 2006; ). Blue light activates the σ B dependent stress response of Bacillus subtilis via YtvA. J Bacteriol 188, 6411–6414.[CrossRef]
    [Google Scholar]
  4. Benson, A. K. & Haldenwang, W. G. ( 1992; ). Characterization of a regulatory network that controls σ B expression in Bacillus subtilis. J Bacteriol 174, 749–757.
    [Google Scholar]
  5. Benson, A. K. & Haldenwang, W. G. ( 1993; ). Bacillus subtilis σ B is regulated by a binding protein (RsbW) that blocks its association with core RNA polymerase. Proc Natl Acad Sci U S A 90, 2330–2334.[CrossRef]
    [Google Scholar]
  6. Brody, M. S., Vijay, K. & Price, C. W. ( 2001; ). Catalytic function of an α/β hydrolase is responsible for energy stress activation of the σ B transcription factor in Bacillus subtilis. J Bacteriol 183, 6422–6428.[CrossRef]
    [Google Scholar]
  7. Chen, C.-C., Lewis, R. J., Harris, R., Yudkin, M. D. & Delumeau, O. ( 2003; ). A supermolecular complex in the environmental stress signalling pathway of Bacillus subtilis. Mol Microbiol 49, 1657–1669.[CrossRef]
    [Google Scholar]
  8. Chen, C.-C., Yudkin, M. D. & Delumeau, O. ( 2004; ). Phosphorylation and RsbX-dependent dephosphorylation of RsbR in the RsbR–RsbS complex of Bacillus subtilis. J Bacteriol 186, 6830–6836.[CrossRef]
    [Google Scholar]
  9. Delumeau, O., Lewis, R. J. & Yudkin, M. D. ( 2002; ). Protein–protein interactions that regulate the energy stress activation of σ B in Bacillus subtilis. J Bacteriol 184, 5583–5589.[CrossRef]
    [Google Scholar]
  10. Delumeau, O., Chen, C.-C., Murray, J. W., Yudkin, M. D. & Lewis, R. J. ( 2006; ). High molecular weight complexes of RsbR and paralogues in the environmental signaling pathway. J Bacteriol 188, 7885–7892.[CrossRef]
    [Google Scholar]
  11. Dufour, A. & Haldenwang, W. G. ( 1994; ). Interactions between a Bacillus subtilis anti-σ factor (RsbW) and its antagonist (RsbV). J Bacteriol 176, 1813–1820.
    [Google Scholar]
  12. Dufour, A., Voelker, U., Voelker, A. & Haldenwang, W. G. ( 1996; ). Relative levels and fractionation properties of Bacillus subtilis σ B and its regulators during balanced growth and stress. J Bacteriol 178, 3701–3709.
    [Google Scholar]
  13. Errington, J. ( 1986; ). A general method for fusion of the Escherichia coli lacZ gene to chromosomal genes in Bacillus subtilis. J Gen Microbiol 132, 2953–2966.
    [Google Scholar]
  14. Eymann, C., Becher, D., Bernhardt, J., Gronau, K., Klutzny, A. & Hecker, M. ( 2007; ). Dynamics of protein phosphorylation on Ser/Thr/Tyr in Bacillus subtilis. Proteomics 7, 3509–3526.[CrossRef]
    [Google Scholar]
  15. Gaidenko, T. A., Yang, X., Lee, Y. M. & Price, C. W. ( 1999; ). Threonine phosphorylation of modulator protein RsbR governs its ability to regulate a serine kinase in the stress signaling pathway of Bacillus subtilis. J Mol Biol 288, 29–39.[CrossRef]
    [Google Scholar]
  16. Gaidenko, T. A., Kim, T.-J., Weigel, A. L., Brody, M. S. & Price, C. W. ( 2006; ). The blue-light receptor YtvA acts as an environmental stress signaling pathway of Bacillus subtilis. J Bacteriol 188, 6387–6395.[CrossRef]
    [Google Scholar]
  17. Guérout-Fleury, A.-M., Shazand, K., Frandsen, N. & Stragier, P. ( 1995; ). Antibiotic-resistance cassettes for Bacillus subtilis. Gene 167, 335–336.[CrossRef]
    [Google Scholar]
  18. Hardwick, S. W., Pane-Farre, J., Delumeau, O., Marles-Wright, J., Murray, J. W., Hecker, M. & Lewis, R. ( 2007; ). Structural and functional characterization of partner switching regulating the environmental stress response in Bacillus subtilis. J Biol Chem 282, 11562–11572.[CrossRef]
    [Google Scholar]
  19. Hecker, M., Schumann, W. & Voelker, U. ( 1996; ). Heat-shock and general stress response in Bacillus subtilis. Mol Microbiol 19, 417–428.[CrossRef]
    [Google Scholar]
  20. Hecker, M., Pane-Farre, J. & Volker, U. ( 2007; ). SigB-dependent general stress response in Bacillus subtilis and related Gram-positive bacteria. Annu Rev Microbiol 61, 215–236.[CrossRef]
    [Google Scholar]
  21. Jonas, R. M., Peters, H. K., III & Haldenwang, W. G. ( 1990; ). Phenotypes of Bacillus subtilis mutants altered in the precursor-specific region of σ E. J Bacteriol 172, 4178–4186.
    [Google Scholar]
  22. Kang, C. M., Brody, M. S., Akbar, S., Yang, X. & Price, C. W. ( 1996; ). Homologous pairs of regulatory proteins control activity of Bacillus subtilis transcription factor σ B in response to environmental stress. J Bacteriol 178, 3846–3853.
    [Google Scholar]
  23. Kenney, T. J. & Moran, C. P., Jr ( 1987; ). Organization and regulation of an operon that encodes a sporulation-essential sigma factor of Bacillus subtilis. J Bacteriol 169, 3329–3339.
    [Google Scholar]
  24. Kim, T.-J., Gaidenko, T. A. & Price, C. W. ( 2004a; ). In vivo phosphorylation of partner switching regulators correlates with stress transmission in the environmental signaling pathway of Bacillus subtilis. J Bacteriol 186, 6124–6132.[CrossRef]
    [Google Scholar]
  25. Kim, T.-J., Gaidenko, T. A. & Price, C. W. ( 2004b; ). A multi-component protein complex mediates environmental stress signaling in Bacillus subtilis. J Mol Biol 341, 135–150.[CrossRef]
    [Google Scholar]
  26. Kuo, S., Zhang, S., Woodbury, R. L. & Haldenwang, W. G. ( 2004; ). Associations between Bacillus subtilis σ B regulators in cell extracts. Microbiology 150, 4125–4136.[CrossRef]
    [Google Scholar]
  27. Marles-Wright, J., Grant, T., Delumeau, O., van Duinin, G., Firbank, S. J., Lewis, P. J., Murray, J. W., Newman, J. A., Quin, M. B. & other authors ( 2008; ). Molecular architecture of the “stressosome,” a signal integration and transduction hub. Science 322, 92–96.[CrossRef]
    [Google Scholar]
  28. Petersohn, A., Bernhardt, J., Gerth, U., Hoper, D., Koburger, T., Voelker, U. & Hecker, M. ( 1999; ). Identification of σ B-dependent genes in Bacillus subtilis using a promoter consensus-directed search and oligonucleotide hybridization. J Bacteriol 181, 5718–5724.
    [Google Scholar]
  29. Price, C. W., Fawcett, P., Ceremonie, H., Su, N., Murphy, C. K. & Youngman, P. ( 2001; ). Genome-wide analysis of the general stress response in Bacillus subtilis. Mol Microbiol 41, 757–774.
    [Google Scholar]
  30. Reeves, A. & Haldenwang, W. G. ( 2007; ). Isolation and characterization of dominant mutations in the Bacillus subtilis stressosome components RsbR and RsbS. J Bacteriol 189, 1531–1541.[CrossRef]
    [Google Scholar]
  31. Reeves, A., Gerth, U., Volker, U. & Haldenwang, W. G. ( 2007; ). ClpP modulates the activity of Bacillus subtilis stress response transcription factor σ B. . J Bacteriol 189, 6168–6175.[CrossRef]
    [Google Scholar]
  32. Sambrook, J., Fritsch, E. F. & Maniatis, T. ( 1989; ). Molecular Cloning: a Laboratory Manual, 2nd edn. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory.
  33. Suzuki, N., Takaya, N., Hashino, T. & Nakamura, A. ( 2007; ). Enhancement of a σ B-dependent stress response in Bacillus subtilis by light via the YtvA photoreceptor. J Gen Appl Microbiol 53, 81–88.[CrossRef]
    [Google Scholar]
  34. Vijay, K., Brody, M. S., Fredlund, E. & Price, C. W. ( 2000; ). A PP2C phosphatase containing a PAS domain is required to convey signals of energy stress to the σ B transcription factor of Bacillus subtilis. Mol Microbiol 35, 180–188.[CrossRef]
    [Google Scholar]
  35. Voelker, U., Voelker, A., Maul, B., Hecker, M., Dufour, A. & Haldenwang, W. G. ( 1995; ). Separate mechanisms activate σ B of Bacillus subtilis in response to environmental and metabolic stresses. J Bacteriol 177, 3771–3780.
    [Google Scholar]
  36. Voelker, U., Voelker, A. & Haldenwang, W. G. ( 1996; ). Reactivation of the Bacillus subtilis anti-σ B antagonist, RsbV, by stress or starvation-induced phosphatase activities. J Bacteriol 178, 5456–5463.
    [Google Scholar]
  37. Voelker, U., Luo, T., Smirnova, N. & Haldenwang, W. G. ( 1997; ). Stress activation of Bacillus subtilis σ B can occur in the absence of the σ B negative regulator RsbX. J Bacteriol 179, 1980–1984.
    [Google Scholar]
  38. Wise, A. A. & Price, C. W. ( 1995; ). Four additional genes in the sigB operon of Bacillus subtilis that control activity of the general stress factor σ B in response to environmental signals. J Bacteriol 177, 123–133.
    [Google Scholar]
  39. Yang, X., Kang, C. M., Brody, M. S. & Price, C. W. ( 1996; ). Opposing pairs of serine protein kinases and phosphatases transmit signals of environmental stress to activate a bacterial transcription factor. Genes Dev 10, 2265–2275.[CrossRef]
    [Google Scholar]
  40. Yasbin, R. E., Wilson, G. A. & Young, F. E. ( 1973; ). Transformation and transfection of lysogenic strains of Bacillus subtilis 168. J Bacteriol 113, 540–548.
    [Google Scholar]
  41. Zhang, S. & Haldenwang, W. G. ( 2005; ). Contributions of ATP, GTP, and redox state to nutritional stress activation of the Bacillus subtilis σ B transcription factor. J Bacteriol 187, 7554–7560.[CrossRef]
    [Google Scholar]
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