1887

Abstract

The transcriptome of was analysed at different time points (30, 60 and 90 min) after a temperature downshift from 37 to 18 °C using DNA macroarrays. This approach allowed the identification of around 50 genes exhibiting an increased mRNA level and around 50 genes exhibiting a decreased mRNA level under cold-shock conditions. Many of the repressed genes encode enzymes involved in the biosynthesis of amino acids, nucleotides and coenzymes, indicating metabolic adaptation of the cells to the decreased growth rate at the lower temperature. The strongest cold-inducible gene encodes fatty acid desaturase, which forms unsaturated fatty acids from saturated phospholipid precursors, thereby increasing membrane fluidity. The cold-shock-induced increase of mRNA levels of the classical cold-shock genes , and could be verified. Furthermore, besides many genes encoding proteins of unknown function, some genes encoding ribosomal proteins were transcriptionally up-regulated, which points to an adaptive reprogramming of the ribosomes under cold-shock conditions. Interestingly, the amount of mRNA specified by the operon ------, which encodes enzymes involved in degradation of branched-chain amino acids, also increases after a temperature downshift. As cells utilize the isoleucine and valine degradation intermediates α-methylbutyryl-CoA and isobutyryl-CoA for synthesis of branched-chain fatty acids, this finding reflects the adaptation of membrane lipid composition, ensuring the maintenance of appropriate membrane fluidity at low temperatures. The results of the DNA array analyses were verified for several selected genes by RNA slot-blot analysis and compared with two-dimensional PAGE analyses.

Loading

Article metrics loading...

/content/journal/micro/10.1099/00221287-148-11-3441
2002-11-01
2024-10-03
Loading full text...

Full text loading...

/deliver/fulltext/micro/148/11/1483441a.html?itemId=/content/journal/micro/10.1099/00221287-148-11-3441&mimeType=html&fmt=ahah

References

  1. Aguilar P. S, Cronan J. E. Jr, De Mendoza D. 1998; A Bacillus subtilis gene induced by cold shock encodes a membrane phospholipid desaturase. J Bacteriol 180:2194–2200
    [Google Scholar]
  2. Aguilar P. S, Lopez P., De Mendoza D. 1999; Transcriptional control of the low-temperature-inducible des gene, encoding the delta5 desaturase of Bacillus subtilis . J Bacteriol 181:7028–7033
    [Google Scholar]
  3. Aguilar P. S, Hernandez-Arriaga A. M, Cybulski L. E, Erazo A. C., De Mendoza D. 2001; Molecular basis of thermosensing: a two-component signal transduction thermometer in Bacillus subtilis . EMBO J 20:1681–1691
    [Google Scholar]
  4. Anagnostopoulos C., Spizizen J. 1961; Requirements for transformation in Bacillus subtilis . J Bacteriol 81:741–746
    [Google Scholar]
  5. Antelmann H, Engelmann S, Schmid R., Hecker M. 1996; General and oxidative stress responses in Bacillus subtilis : cloning, expression and mutation of the alkyl hydroperoxide reductase operon. J Bacteriol 178:6571–6578
    [Google Scholar]
  6. Begley T. P, Downs D. M, Ealick S. E. 8 other authors 1999; Thiamin biosynthesis in prokaryotes. Arch Microbiol 71:293–300
    [Google Scholar]
  7. Bernhardt J., Werner H. 1999 The 2D protein index of Bacillus subtilis – Sub2D http://microbio2.biologie.uni-greifswald.de:8880/ (Ernst-Moritz-Arndt-Universität Greifswald, Greifswald, Germany:
    [Google Scholar]
  8. Bernhardt J, Völker U, Völker A, Antelmann H, Schmid R, Mach H., Hecker M. 1997; Specific and general stress proteins in Bacillus subtilis – a two dimensional protein electrophoresis study. Microbiology 143:999–1017
    [Google Scholar]
  9. Bernhardt J, Büttner K, Scharf C., Hecker M. 1999; Dual channel imaging of two-dimensional electropherograms in Bacillus subtilis . Electrophoresis 20:2225–2240
    [Google Scholar]
  10. Bower S, Perkins J. B, Yocum R. R, Howitt C. L, Rahaim P., Pero J. 1996; Cloning, sequencing, and characterization of the Bacillus subtilis biotin biosynthetic operon. J Bacteriol 178:4122–4130
    [Google Scholar]
  11. Brandi A, Pietroni P, Gualerzi C. O., Pon C. L. 1996; Post-transcriptional regulation of CspA expression in Escherichia coli . Mol Microbiol 19:231–240
    [Google Scholar]
  12. Chomczynski P., Sacchi N. 1987; Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal Biochem 162:156–159
    [Google Scholar]
  13. Debarbouille M, Gardan R, Arnaud M., Rapoport G. 1999; Role of bkdR , a transcriptional activator of the SigL-dependent isoleucine and valine degradation pathway in Bacillus subtilis . J Bacteriol 181:2059–2066
    [Google Scholar]
  14. Ebbole D. J., Zalkin H. 1987; Cloning and characterization of a 12-gene cluster from Bacillus subtilis encoding nine enzymes for de novo purine nucleotide synthesis. J Biol Chem 262:8274–8287
    [Google Scholar]
  15. Fang L, Jiang W, Bae W., Inouye M. 1997; Promoter-independent cold shock induction of cspA and its derepression at 37 °C by mRNA stabilisation. Mol Microbiol 23:355–364
    [Google Scholar]
  16. Foster S. J. 1993; Molecular analysis of three major wall-associated proteins of Bacillus subtilis 168: evidence for processing of the product of a gene encoding a 258 kDa precursor two-domain ligand-binding protein. Mol Microbiol 8:299–310
    [Google Scholar]
  17. Goldenberg D, Azar I., Oppenheim A. B. 1996; Differential mRNA stability of the cspA gene in the cold-shock response of Escherichia coli . Mol Microbiol 19:241–248
    [Google Scholar]
  18. Grandoni J. A, Zahler S. A., Calvo J. M. 1992; Transcriptional regulation of the ilv - leu operon of Bacillus subtilis. J Bacteriol 174:3212–3219
    [Google Scholar]
  19. Graumann P, Schröder K, Schmid R., Marahiel M. A. 1996; Cold shock stress-induced proteins in Bacillus subtilis . J Bacteriol 178:4611–4619
    [Google Scholar]
  20. Graumann P, Wendrich T. M, Weber M. H, Schröder K., Marahiel M. A. 1997; A family of cold shock proteins in Bacillus subtilis is essential for cellular growth and for efficient protein synthesis at optimal and low temperatures. Mol Microbiol 25:741–756
    [Google Scholar]
  21. Graumann P. L., Marahiel M. A. 1999; Cold shock response in Bacillus subtilis . J Mol Microbiol Biotechnol 1:203–209
    [Google Scholar]
  22. Grundy F. J., Henkin T. M. 1998; The S box regulon: a new global transcription termination control system for methionine and cysteine biosynthesis genes in gram-positive bacteria. Mol Microbiol 30:737–749
    [Google Scholar]
  23. Kaan T, Jürgen B., Schweder T. 1999; Regulation of the expression of the cold shock proteins CspB and CspC in Bacillus subtilis . Mol Gen Genet 262:351–354
    [Google Scholar]
  24. Kandror O., Goldberg A. L. 1997; Trigger factor is induced upon cold shock and enhances viability of Escherichia coli at low temperatures. Proc Natl Acad Sci USA 94:4978–4981
    [Google Scholar]
  25. Kappes R. M, Kempf B, Kneip S, Boch J, Gade J, Meier-Wagner J., Bremer E. 1999; Two evolutionarily closely related ABC transporters mediate the uptake of choline for synthesis of the osmoprotectant glycine betaine in Bacillus subtilis . Mol Microbiol 32:203–216
    [Google Scholar]
  26. Klein W, Weber M. H., Marahiel M. A. 1999; Cold shock response of Bacillus subtilis : isoleucine-dependent switch in the fatty acid branching pattern for membrane adaptation to low temperatures. J Bacteriol 181:5341–5349
    [Google Scholar]
  27. Lopez M. M, Yutani K., Makhatadze G. I. 1999; Interactions of the major cold shock protein of Bacillus subtilis CspB with single-stranded DNA templates of different base composition. J Biol Chem 274:33601–33608
    [Google Scholar]
  28. Lopez M. M, Yutani K., Makhatadze G. I. 2001; Interactions of the cold shock protein CspB from Bacillus subtilis with single-stranded DNA. Importance of the T base content and position within the template. J Biol Chem 276:15511–15518
    [Google Scholar]
  29. Lottering E. A., Streips U. N. 1995; Induction of cold shock proteins in Bacillus subtilis . Curr Microbiol 30:193–199
    [Google Scholar]
  30. Mantsala P., Zalkin H. 1992; Cloning and sequence of Bacillus subtilis purA and guaA , involved in the conversion of IMP to AMP and GMP. J Bacteriol 174:1883–1890
    [Google Scholar]
  31. Murphy B. A, Grundy F. J., Henkin T. M. 2002; Prediction of gene function in methylthioadenosine recycling from regulatory signals. J Bacteriol 184:2314–2318
    [Google Scholar]
  32. Petersohn A, Brigulla M, Haas S, Hoheisel J. D, Volker U., Hecker M. 2001; Global analysis of the general stress response of Bacillus subtilis . J Bacteriol 183:5617–5631
    [Google Scholar]
  33. Que Q., Helmann J. D. 2000; Manganese homeostasis in Bacillus subtilis is regulated by MntR, a bifunctional regulator related to the diphtheria toxin repressor family of proteins. Mol Microbiol 35:1454–1468
    [Google Scholar]
  34. Saxild H. H., Nygaard P. 1988; Gene-enzyme relationships of the purine biosynthetic pathway in Bacillus subtilis . Mol Gen Genet 211:160–167
    [Google Scholar]
  35. Schindler T, Graumann P. L, Perl D, Ma S, Schmid F. X., Marahiel M. A. 1999; The family of cold shock proteins of Bacillus subtilis . Stability and dynamics in vitro and in vivo. J Biol Chem 274:3407–3413
    [Google Scholar]
  36. Schmid R, Bernhardt J, Antelmann H, Völker A, Mach H, Völker U., Hecker M. 1997; Identification of vegetative proteins for two dimensional protein index of Bacillus subtilis . Microbiology 143:991–998
    [Google Scholar]
  37. Schröder K, Graumann P, Schnuchel A, Holak T. A., Marahiel M. A. 1995; Mutational analysis of the putative nucleic acid-binding surface of the cold-shock domain, CspB, revealed an essential role of aromatic and basic residues in binding of single-stranded DNA containing the Y-box motif. Mol Microbiol 16:699–708
    [Google Scholar]
  38. Scotti C, Valbuzzi A, Perego M, Galizzi A., Albertini A. M. 1996; The Bacillus subtilis genes for ribonucleotide reductase are similar to the genes for the second class I NrdE/NrdF enzymes of Enterobacteriaceae. Microbiology 142:2995–3004
    [Google Scholar]
  39. Spizizen J. 1958; Transformation of biochemically deficient strains of Bacillus subtilis by deoxyribonucleate. Proc Natl Acad Sci USA 44:1072–1084
    [Google Scholar]
  40. Stülke J, Hanschke R., Hecker M. 1993; Temporal activation of the β-glucanase synthesis in Bacillus subtilis is mediated by the GTP pool. J Gen Microbiol 139:2041–2045
    [Google Scholar]
  41. Tobisch S, Zühlke D, Bernhardt J, Stülke J., Hecker M. 1999; Role of CcpA in regulation of the central pathways of carbon catabolism in Bacillus subtilis . J Bacteriol 181:6996–7004
    [Google Scholar]
  42. Vagner V, Dervyn E., Ehrlich S. D. 1998; A vector for systematic gene inactivation in Bacillus subtilis . Microbiology 144:3097–3104
    [Google Scholar]
  43. Völker U, Engelmann S, Maul B, Riethdorf S, Völker A, Schmid R, Mach H., Hecker M. 1994; Analysis of the induction of general stress proteins of Bacillus subtilis . Microbiology 140:741–752
    [Google Scholar]
  44. Weber M. H, Klein W, Müller L, Niess U. M., Marahiel M. A. 2001a; Role of the Bacillus subtilis fatty acid desaturase in membrane adaptation during cold shock. Mol Microbiol 39:1321–1329
    [Google Scholar]
  45. Weber M. H, Volkov A. V, Fricke I, Marahiel M. A., Graumann P. L. 2001b; Localization of cold shock proteins to cytosolic spaces surrounding nucleoids in Bacillus subtilis depends on active transcription. J Bacteriol 183:6435–6443
    [Google Scholar]
  46. Wetzstein M, Völker U, Dedio J, Löbau S, Zuber U, Schiesswohl M, Herget C, Hecker M., Schumann W. 1992; Cloning, sequencing and molecular analysis of the dnaK locus from Bacillus subtilis . J Bacteriol 174:3300–3310
    [Google Scholar]
  47. Wiegert T, Homuth G, Versteeg S., Schumann W. 2001; Alkaline shock induces the Bacillus subtilis sigmaW regulon. Mol Microbiol 41:59–71
    [Google Scholar]
/content/journal/micro/10.1099/00221287-148-11-3441
Loading
/content/journal/micro/10.1099/00221287-148-11-3441
Loading

Data & Media loading...

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error