1887

Abstract

In , expression of the gene, encoding glutamate dehydrogenase, and the operon, involved in arginine catabolism, requires SigL (σ)-containing RNA polymerase as well as RocR, a positive regulator of the NtrC/NifA family. The RocR protein was purified and shown to bind specifically to the intergenic region located between and the operon. DNaseI footprinting experiments were used to define the RocR-binding site as an 8 bp inverted repeat, separated by one base pair, forming an imperfect palindrome which is present twice within the intergenic region, acting as both a downstream activating sequence (DAS) and an upstream activating sequence (UAS). Point mutations in either of these two sequences significantly lowered expression of both and . This bidirectional enhancer element retained partial activity even when moved 9 kb downstream of the promoter. Electron microscopy experiments indicated that an intrinsically curved region is located between the UAS/DAS region and the promoter of the operon. This curvature could facilitate interaction of RocR with σ-RNA polymerase at the promoter.

Loading

Article metrics loading...

/content/journal/micro/10.1099/mic.0.26013-0
2003-03-01
2019-11-12
Loading full text...

Full text loading...

/deliver/fulltext/micro/149/3/mic149739.html?itemId=/content/journal/micro/10.1099/mic.0.26013-0&mimeType=html&fmt=ahah

References

  1. Baumberg, S. & Klingel, U. ( 1993; ). Biosynthesis of arginine, proline and related compounds. In Bacillus subtilis and Other Gram-positive Bacteria: Biochemistry, Physiology and Molecular Genetics, pp. 229–306. Edited by A. L. Sonenshein, J. A. Hoch & R. Losick. Washington, DC: American Society for Microbiology.
  2. Belitsky, B. & Sonenshein, A. L. ( 1998; ). Role and regulation of Bacillus subtilis glutamate dehydrogenase genes. J Bacteriol 180, 6298–6305.
    [Google Scholar]
  3. Belitsky, B. & Sonenshein, A. L. ( 1999; ). An enhancer element located downstream of the major glutamate dehydrogenase gene of Bacillus subtilis. Proc Natl Acad Sci U S A 96, 10290–10295.[CrossRef]
    [Google Scholar]
  4. Bolshoy, A., McNamara, P., Harrington, R. E. & Trifonov, E. N. ( 1991; ). Curved DNA without A-A: experimental estimation of all 16 DNA wedge angles. Proc Natl Acad Sci U S A 88, 2312–2316.[CrossRef]
    [Google Scholar]
  5. Bradford, M. M. ( 1976; ). A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein–dye binding. Anal Biochem 72, 248–254.[CrossRef]
    [Google Scholar]
  6. Calogero, S., Gardan, R., Glaser, P., Schweitzer, J., Rapoport, G. & Débarbouillé, M. ( 1994; ). RocR, a novel regulatory protein controlling arginine utilization in Bacillus subtilis, belongs to the NtrC/NifA family of transcriptional activators. J Bacteriol 176, 1234–1241.
    [Google Scholar]
  7. Cunin, R., Glandsdorff, N., Pierard, A. & Stalon, V. ( 1986; ). Biosynthesis and metabolism of arginine in bacteria. Microbiol Rev 50, 314–352.
    [Google Scholar]
  8. De Santis, P., Palleschi, A., Savino, M. & Scipioni, A. ( 1988; ). A theoretical model of DNA curvature. Biophys Chem 32, 305–317.[CrossRef]
    [Google Scholar]
  9. Derré, I., Rapoport, G. & Msadek, T. ( 1999; ). CtsR, a novel regulator of stress and heat shock response, controls clp and molecular chaperone gene expression in Gram-positive bacteria. Mol Microbiol 31, 117–132.[CrossRef]
    [Google Scholar]
  10. Dubochet, J., Ducommun, M., Zollinger, M. & Kellenberger, E. ( 1971; ). A new preparation method for dark-field electron microscopy of macromolecules. J Ultrastruct Res 35, 147–167.[CrossRef]
    [Google Scholar]
  11. Farez-Vidal, M. E., Wilson, T. J., Davidson, B. E., Howlett, G. J., Austin, S. & Dixon, R. A. ( 1996; ). Effector-induced self association and conformation changes in the enhancer-binding protein NtrC. Mol Microbiol 22, 779–788.[CrossRef]
    [Google Scholar]
  12. Fisher, S. ( 1993; ). Utilization of amino acids and other nitrogen containing compounds. In Bacillus subtilis and Other Gram-positive Bacteria: Biochemistry, Physiology and Molecular Genetics, pp. 221–228. Edited by A. L. Sonenshein, J. A. Hoch & R. Losick. Washington, DC: American Society for Microbiology.
  13. Fisher, S. & Débarbouillé, M. ( 2002; ). Nitrogen source utilization and its regulation. In Bacillus subtilis and its Closest Relatives. From Genes to Cells, pp. 181–191. Edited by A. L. Sonenshein, J. A. Hoch & R. Losick. Washington, DC: American Society for Microbiology.
  14. Fouet, A. & Sonenshein, A. L. ( 1990; ). A target for carbon source-dependent negative regulation of the citB promoter of Bacillus subtilis. J Bacteriol 172, 835–844.
    [Google Scholar]
  15. Gardan, R., Rapoport, G. & Débarbouillé, M. ( 1995; ). Expression of the rocDEF operon involved in arginine catabolism in Bacillus subtilis. J Mol Biol 249, 843–856.[CrossRef]
    [Google Scholar]
  16. Gardan, R., Rapoport, G. & Débarbouillé, M. ( 1997; ). Role of the transcriptional activator RocR in the arginine-degradation pathway of Bacillus subtilis. Mol Microbiol 24, 825–837.[CrossRef]
    [Google Scholar]
  17. Gibson, T. J. ( 1984; ). Studies on the Epstein–Barr Virus Genome. Cambridge: University of Cambridge.
  18. Harwood, C. R. & Baumberg, S. ( 1977; ). Arginine hydroxamate resistant mutants of Bacillus subtilis with altered control of arginine metabolism. J Gen Microbiol 100, 177–188.[CrossRef]
    [Google Scholar]
  19. Kabsch, W., Sander, S. & Trifonov, E. N. ( 1982; ). The ten helical twist angles of B-DNA. Nucleic Acids Res 10, 1097–1104.[CrossRef]
    [Google Scholar]
  20. Koo, H.-S. & Crothers, D. M. ( 1988; ). Calibration of DNA curvature and a unified description of sequence-directed bending. Proc Natl Acad Sci U S A 85, 1763–1767.[CrossRef]
    [Google Scholar]
  21. Kunkel, T. A., Roberts, J. D. & Zakour, R. A. ( 1987; ). Rapid and efficient site-specific mutagenesis without phenotypic selection. Methods Enzymol 154, 367–382.
    [Google Scholar]
  22. Kustu, S., Santero, E., Keener, J., Popham, D. & Weiss, D. ( 1989; ). Expression of sigma 54 (ntrA)-dependent genes is probably united by a common mechanism. Microbiol Rev 53, 367–376.
    [Google Scholar]
  23. Kustu, S., North, A. K. & Weiss, D. ( 1991; ). Prokaryotic transcriptional enhancers and enhancer-binding proteins. Trends Biochem Sci 16, 397–402.[CrossRef]
    [Google Scholar]
  24. Laemmli, U. K. ( 1970; ). Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227, 680–685.[CrossRef]
    [Google Scholar]
  25. Larquet, E., Furrer, P., Stasiak, A., Dubochet, J. & Revet, B. ( 1995; ). DNA ResCue: 3D reconstruction and simulation curvature of DNA from the analysis of electron micrographs. (Ninth Conversation in Biomolecular Stereodynamics, June 20–24, 1995). J Biomol Struct Dyn 12, A134.
    [Google Scholar]
  26. Le Cam, E. & Delain, E. ( 1995; ). Nucleic acids–ligand interactions. In Visualization of Nucleic Acids, pp. 333–358. Edited by G. Morel. Boca Raton, FL: CRC Press.
  27. Lereclus, D. & Arantès, O. ( 1992; ). spbA locus ensures the segregational stability of pHT1030, a novel type of Gram-positive replicon. Mol Microbiol 6, 35–46.[CrossRef]
    [Google Scholar]
  28. Martin-Verstraete, I., Débarbouillé, M., Klier, A. & Rapoport, G. ( 1990; ). Levanase operon of Bacillus subtilis includes a fructose-specific phosphotransferase system regulating the expression of the operon. J Mol Biol 214, 657–671.[CrossRef]
    [Google Scholar]
  29. Martin-Verstraete, I., Débarbouillé, M., Klier, A. & Rapoport, G. ( 1992; ). Mutagenesis of the Bacillus subtilis “−12, −24” promotor of the levanase operon and evidence for the existence of an upstream activating sequence. J Mol Biol 226, 85–99.[CrossRef]
    [Google Scholar]
  30. Merrick, M. J. ( 1993; ). In a class of its own – the RNA polymerase sigma factor sigma 54 (sigma N). Mol Microbiol 10, 903–909.[CrossRef]
    [Google Scholar]
  31. Miller, C., Baumberg, S. & Stockley, P. G. ( 1997; ). Operator interactions by the Bacillus subtilis arginine repressor/activator AhrC: novel positioning and DNA-mediated assembly of a transcriptional activator at catabolic sites. Mol Microbiol 26, 37–48.[CrossRef]
    [Google Scholar]
  32. Miller, J. H. ( 1972; ). Experiments in Molecular Genetics. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory.
  33. Morett, E. & Segovia, L. ( 1993; ). The σ54 bacterial enhancer-binding protein family: mechanism of action and phylogenetic relationship of their functional domains. J Bacteriol 175, 6067–6074.
    [Google Scholar]
  34. Muzard, G., Theveny, B. & Revet, B. ( 1990; ). Electron microscopy mapping of pBR322 DNA curvature. Comparison with theoretical models. EMBO J 9, 1289–1298.
    [Google Scholar]
  35. Perez-Martin, J. & de Lorenzo, V. ( 1996; ). ATP binding to the sigma 54-dependent activator XylR triggers a protein multimerization cycle catalyzed by UAS DNA. Cell 86, 331–339.[CrossRef]
    [Google Scholar]
  36. Sambrook, J., Fritsch, E. F. & Maniatis, T. ( 1989; ). Molecular Cloning: a Laboratory Manual, 2nd edn. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory.
  37. Studier, F. W. & Moffatt, B. A. ( 1986; ). Use of bacteriophage T7 RNA polymerase to direct selective high-level expression of cloned genes. J Mol Biol 189, 113–130.[CrossRef]
    [Google Scholar]
  38. Thei Dame, R., Wyman, C. & Goosen, N. ( 2001; ). Structural basis for preferential binding of H-NS to curved DNA. Biochimie 83, 231–234.[CrossRef]
    [Google Scholar]
  39. Tricot, C., Stalon, V. & Legrain, C. ( 1991; ). Isolation and characterization of Pseudomonas putida mutants affected in arginine, ornithine and citrulline catabolism: function of the arginine oxidase and arginine succinyltransferase pathways. J Gen Microbiol 137, 2911–2918.[CrossRef]
    [Google Scholar]
  40. Ulanovsky, L., Bodner, M., Trifonov, E. N. & Choder, M. ( 1986; ). Curved DNA: design, synthesis and circularization. Proc Natl Acad Sci U S A 83, 862–866.[CrossRef]
    [Google Scholar]
  41. Wikstrom, P., O'Neil, E. & Shingler, V. ( 2001; ). The regulatory N-terminal region of the aromatic-responsive transcriptional activator DmpR constrains nucleotide-triggered multimerization. J Mol Biol 314, 971–984.[CrossRef]
    [Google Scholar]
  42. Wilson, T. J., Maroudas, G., Howlett, G. J. & Davidson, B. D. ( 1994; ). Ligand-induced self association of the Escherichia coli regulatory protein TyrR. J Mol Biol 238, 309–318.[CrossRef]
    [Google Scholar]
  43. Wyman, C., Rombel, I., North, A., Bustamante, C. & Kustu, S. ( 1997; ). Unusual oligomerization required for activity of NtrC, a bacterial enhancer-binding protein. Science 275, 1658–1661.[CrossRef]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/mic.0.26013-0
Loading
/content/journal/micro/10.1099/mic.0.26013-0
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