1887

Abstract

Expression of mannitol utilization genes in is directed by P, the promoter of the operon, and P, the promoter of the MtlR activator. MtlR contains phosphoenolpyruvate-dependent phosphotransferase system (PTS) regulation domains, called PRDs. The activity of PRD-containing MtlR is mainly regulated by the phosphorylation/dephosphorylation of its PRDII and EIIB-like domains. Replacing histidine 342 and cysteine 419 residues, which are the targets of phosphorylation in these two domains, by aspartate and alanine provided MtlR-H342D C419A, which permanently activates P. In the -H342D C419A mutant, P was active, even when the operon was deleted from the genome. The -H342D C419A allele was expressed in an strain lacking enzyme I of the PTS. Electrophoretic mobility shift assays using purified MtlR-H342D C419A showed an interaction between the MtlR double-mutant and the Cy5-labelled P and P DNA fragments. These investigations indicate that the activated MtlR functions regardless of the presence of the mannitol-specific transporter (MtlA). This is in contrast to the proposed model in which the sequestration of MtlR by the MtlA transporter is necessary for the activity of MtlR. Additionally, DNase I footprinting, construction of P-P hybrid promoters, as well as increasing the distance between the MtlR operator and the −35 box of P revealed that the activated MtlR molecules and RNA polymerase holoenzyme likely form a class II type activation complex at P and P during transcription initiation.

Loading

Article metrics loading...

/content/journal/micro/10.1099/mic.0.071233-0
2014-01-01
2020-07-10
Loading full text...

Full text loading...

/deliver/fulltext/micro/160/1/91.html?itemId=/content/journal/micro/10.1099/mic.0.071233-0&mimeType=html&fmt=ahah

References

  1. Altenbuchner J., Viell P., Pelletier I.. ( 1992;). Positive selection vectors based on palindromic DNA sequences. Methods Enzymol216:457–466 [CrossRef][PubMed]
    [Google Scholar]
  2. Arnaud M., Vary P., Zagorec M., Klier A., Debarbouille M., Postma P., Rapoport G.. ( 1992;). Regulation of the sacPA operon of Bacillus subtilis: identification of phosphotransferase system components involved in SacT activity. J Bacteriol174:3161–3170[PubMed]
    [Google Scholar]
  3. Baba T., Ara T., Hasegawa M., Takai Y., Okumura Y., Baba M., Datsenko K. A., Tomita M., Wanner B. L., Mori H.. ( 2006;). Construction of Escherichia coli K-12 in-frame, single-gene knockout mutants: the Keio collection. Mol Syst Biol2:0008 [CrossRef][PubMed]
    [Google Scholar]
  4. Bouraoui H., Ventroux M., Noirot-Gros M. F., Deutscher J., Joyet P.. ( 2013;). Membrane sequestration by the EIIB domain of the mannitol permease MtlA activates the Bacillus subtilis mtl operon regulator MtlR. Mol Microbiol87:789–801 [CrossRef][PubMed]
    [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 Biochem72:248–254 [CrossRef][PubMed]
    [Google Scholar]
  6. Browning D. F., Busby S. J.. ( 2004;). The regulation of bacterial transcription initiation. Nat Rev Microbiol2:57–65 [CrossRef][PubMed]
    [Google Scholar]
  7. Crutz A. M., Steinmetz M., Aymerich S., Richter R., Le Coq D.. ( 1990;). Induction of levansucrase in Bacillus subtilis: an antitermination mechanism negatively controlled by the phosphotransferase system. J Bacteriol172:1043–1050[PubMed]
    [Google Scholar]
  8. Deutscher J., Galinier A., Martin-Verstraete I.. ( 2002;). Carbohydrate uptake and metabolism. Bacillus Subtilis and its Closest Relatives: from Genes to Cells129–150 Sonenshein A. L., Hoch J. A., Losick R.. Washington, DC: American Society for Microbiology;[CrossRef]
    [Google Scholar]
  9. Deutscher J., Francke C., Postma P. W.. ( 2006;). How phosphotransferase system-related protein phosphorylation regulates carbohydrate metabolism in bacteria. Microbiol Mol Biol Rev70:939–1031 [CrossRef][PubMed]
    [Google Scholar]
  10. Fujita Y.. ( 2009;). Carbon catabolite control of the metabolic network in Bacillus subtilis. . Biosci Biotechnol Biochem73:245–259 [CrossRef][PubMed]
    [Google Scholar]
  11. Greenberg D. B., Stülke J., Saier M. H. Jr. ( 2002;). Domain analysis of transcriptional regulators bearing PTS regulatory domains. Res Microbiol153:519–526 [CrossRef][PubMed]
    [Google Scholar]
  12. Guérout-Fleury A. M., Frandsen N., Stragier P.. ( 1996;). Plasmids for ectopic integration in Bacillus subtilis. . Gene180:57–61 [CrossRef][PubMed]
    [Google Scholar]
  13. Harwood C. R., Cutting S. M.. ( 1990;). Molecular Biological Methods for Bacillus New York: Wiley;
    [Google Scholar]
  14. Heravi K. M., Wenzel M., Altenbuchner J.. ( 2011;). Regulation of mtl operon promoter of Bacillus subtilis: requirements of its use in expression vectors. Microb Cell Fact10:83 [CrossRef][PubMed]
    [Google Scholar]
  15. Hoffmann J., Bóna-Lovász J., Beuttler H., Altenbuchner J.. ( 2012;). In vivo and in vitro studies on the carotenoid cleavage oxygenases from Sphingopyxis alaskensis RB2256 and Plesiocystis pacifica SIR-1 revealed their substrate specificities and non-retinal-forming cleavage activities. FEBS J279:3911–3924 [CrossRef][PubMed]
    [Google Scholar]
  16. Joyet P., Derkaoui M., Poncet S., Deutscher J.. ( 2010;). Control of Bacillus subtilis mtl operon expression by complex phosphorylation-dependent regulation of the transcriptional activator MtlR. Mol Microbiol76:1279–1294 [CrossRef][PubMed]
    [Google Scholar]
  17. Joyet P., Bouraoui H., Aké F. M., Derkaoui M., Zébré A. C., Cao T. N., Ventroux M., Nessler S., Noirot-Gros M. F.. & other authors ( 2013;). Transcription regulators controlled by interaction with enzyme IIB components of the phosphoenolpyruvate: sugar phosphotransferase system. Biochim Biophys Acta1834:1415–1424 [CrossRef][PubMed]
    [Google Scholar]
  18. Lee D. J., Minchin S. D., Busby S. J.. ( 2012;). Activating transcription in bacteria. Annu Rev Microbiol66:125–152 [CrossRef][PubMed]
    [Google Scholar]
  19. Lindner C., Galinier A., Hecker M., Deutscher J.. ( 1999;). Regulation of the activity of the Bacillus subtilis antiterminator LicT by multiple PEP-dependent, enzyme I- and HPr-catalysed phosphorylation. Mol Microbiol31:995–1006 [CrossRef][PubMed]
    [Google Scholar]
  20. Lindner C., Hecker M., Le Coq D., Deutscher J.. ( 2002;). Bacillus subtilis mutant LicT antiterminators exhibiting enzyme I- and HPr-independent antitermination affect catabolite repression of the bglPH operon. J Bacteriol184:4819–4828 [CrossRef][PubMed]
    [Google Scholar]
  21. Lopian L., Nussbaum-Shochat A., O’Day-Kerstein K., Wright A., Amster-Choder O.. ( 2003;). The BglF sensor recruits the BglG transcription regulator to the membrane and releases it on stimulation. Proc Natl Acad Sci U S A100:7099–7104 [CrossRef][PubMed]
    [Google Scholar]
  22. Luria S. E., Adams J. N., Ting R. C.. ( 1960;). Transduction of lactose-utilizing ability among strains of E. coli and S. dysenteriae and the properties of the transducing phage particles. Virology12:348–390 [CrossRef][PubMed]
    [Google Scholar]
  23. Martin-Verstraete I., Débarbouillé M., Klier A., Rapoport G.. ( 1992;). Mutagenesis of the Bacillus subtilis “-12, -24” promoter of the levanase operon and evidence for the existence of an upstream activating sequence. J Mol Biol226:85–99 [CrossRef][PubMed]
    [Google Scholar]
  24. Michel J. F., Millet J.. ( 1970;). Physiological studies on early-blocked sporulation mutants of Bacillus subtilis. . J Appl Bacteriol33:220–227 [CrossRef][PubMed]
    [Google Scholar]
  25. Miller J. H.. ( 1972;). Experiments in Molecular Genetics Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
    [Google Scholar]
  26. Motejadded H., Altenbuchner J.. ( 2007;). Integration of a lipase gene into the Bacillus subtilis chromosome: Recombinant strains without antibiotic resistance marker. Iranian J Biotechnol5:105–109
    [Google Scholar]
  27. Reizer J., Bachem S., Reizer A., Arnaud M., Saier M. H. Jr, Stülke J.. ( 1999;). Novel phosphotransferase system genes revealed by genome analysis - the complete complement of PTS proteins encoded within the genome of Bacillus subtilis . Microbiology145:3419–3429[PubMed][CrossRef]
    [Google Scholar]
  28. Rothe F. M., Wrede C., Lehnik-Habrink M., Görke B., Stülke J.. ( 2013;). Dynamic localization of a transcription factor in Bacillus subtilis: the LicT antiterminator relocalizes in response to inducer availability. J Bacteriol195:2146–2154 [CrossRef][PubMed]
    [Google Scholar]
  29. Sambrook J., Fritsch E. F., Maniatis T.. ( 1989;). Molecular Cloning: a Laboratory Manual, 2nd edn. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
    [Google Scholar]
  30. Sanger F., Nicklen S., Coulson A. R.. ( 1977;). DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A74:5463–5467 [CrossRef][PubMed]
    [Google Scholar]
  31. Schnetz K., Stülke J., Gertz S., Krüger S., Krieg M., Hecker M., Rak B.. ( 1996;). LicT, a Bacillus subtilis transcriptional antiterminator protein of the BglG family. J Bacteriol178:1971–1979[PubMed]
    [Google Scholar]
  32. Sonenshein A. L.. ( 2007;). Control of key metabolic intersections in Bacillus subtilis. . Nat Rev Microbiol5:917–927 [CrossRef][PubMed]
    [Google Scholar]
  33. Stülke J., Martin-Verstraete I., Zagorec M., Rose M., Klier A., Rapoport G.. ( 1997;). Induction of the Bacillus subtilis ptsGHI operon by glucose is controlled by a novel antiterminator, GlcT. Mol Microbiol25:65–78 [CrossRef][PubMed]
    [Google Scholar]
  34. Stülke J., Arnaud M., Rapoport G., Martin-Verstraete I.. ( 1998;). PRD–a protein domain involved in PTS-dependent induction and carbon catabolite repression of catabolic operons in bacteria. Mol Microbiol28:865–874 [CrossRef][PubMed]
    [Google Scholar]
  35. Sun T., Altenbuchner J.. ( 2010;). Characterization of a mannose utilization system in Bacillus subtilis. . J Bacteriol192:2128–2139 [CrossRef][PubMed]
    [Google Scholar]
  36. Titok M. A., Chapuis J., Selezneva Y. V., Lagodich A. V., Prokulevich V. A., Ehrlich S. D., Jannière L.. ( 2003;). Bacillus subtilis soil isolates: plasmid replicon analysis and construction of a new theta-replicating vector. Plasmid49:53–62 [CrossRef][PubMed]
    [Google Scholar]
  37. Tobisch S., Stülke J., Hecker M.. ( 1999;). Regulation of the lic operon of Bacillus subtilis and characterization of potential phosphorylation sites of the LicR regulator protein by site-directed mutagenesis. J Bacteriol181:4995–5003[PubMed]
    [Google Scholar]
  38. Tortosa P., Declerck N., Dutartre H., Lindner C., Deutscher J., Le Coq D.. ( 2001;). Sites of positive and negative regulation in the Bacillus subtilis antiterminators LicT and SacY. Mol Microbiol41:1381–1393 [CrossRef][PubMed]
    [Google Scholar]
  39. Watanabe S., Hamano M., Kakeshita H., Bunai K., Tojo S., Yamaguchi H., Fujita Y., Wong S. L., Yamane K.. ( 2003;). Mannitol-1-phosphate dehydrogenase (MtlD) is required for mannitol and glucitol assimilation in Bacillus subtilis: possible cooperation of mtl and gut operons. J Bacteriol185:4816–4824 [CrossRef][PubMed]
    [Google Scholar]
  40. Wenzel M., Altenbuchner J.. ( 2013;). The Bacillus subtilis mannose regulator, ManR, a DNA-binding protein regulated by HPr and its cognate PTS transporter ManP. Mol Microbiol88:562–576 [CrossRef][PubMed]
    [Google Scholar]
  41. Yamamoto H., Serizawa M., Thompson J., Sekiguchi J.. ( 2001;). Regulation of the glv operon in Bacillus subtilis: YfiA (GlvR) is a positive regulator of the operon that is repressed through CcpA and cre. . J Bacteriol183:5110–5121 [CrossRef][PubMed]
    [Google Scholar]
  42. Yanisch-Perron C., Vieira J., Messing J.. ( 1985;). Improved M13 phage cloning vectors and host strains: nucleotide sequences of the M13mp18 and pUC19 vectors. Gene33:103–119 [CrossRef][PubMed]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/mic.0.071233-0
Loading
/content/journal/micro/10.1099/mic.0.071233-0
Loading

Data & Media loading...

Most cited this month Most Cited RSS feed

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