Skip to main content
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1996 Aug;178(16):4861–4869. doi: 10.1128/jb.178.16.4861-4869.1996

Effects of mecA and mecB (clpC) mutations on expression of sigD, which encodes an alternative sigma factor, and autolysin operons and on flagellin synthesis in Bacillus subtilis.

M H Rashid 1, A Tamakoshi 1, J Sekiguchi 1
PMCID: PMC178268  PMID: 8759849

Abstract

The expression of the major vegetative phase-specific autolysin genes (cwlB [lytC] and cwlG [lytD]) was greatly reduced by mecA and mecB null mutations. In contrast to the negative effects on late competence genes (such as comG) and levansucrase gene (sacB) expression, this positive effect of mec genes on autolysin gene expression was not mediated through the ComK protein but apparently through the level of the SigD protein. The pleiotropic effects of the mec mutations, i.e., the reduction of sigD expression and the overexpression of the ComK protein, seem not to be interwoven since the SigD- and ComK-dependent functions are clearly separable in the mec mutants. We also show that the synthesis of the flagellin protein, which is encoded by the SigD-dependent hag gene, was similarly affected by the mec mutations. Complementation analysis with a SigD-overproducing plasmid, pHYSigD, in mec mutants revealed the reversion of almost all of the SigD-dependent phenotypes except motility. This finding suggested that Mec proteins act on motility genes at two levels, one of which is apparently SigD independent. Finally, we discuss the transcriptional regulation of the sigD gene by multiple regulators, i.e., MecA, MecB, SinR (FlaD), and DegS-DegU, and its implications for cells in a global context.

Full Text

The Full Text of this article is available as a PDF (596.1 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Anagnostopoulos C., Spizizen J. REQUIREMENTS FOR TRANSFORMATION IN BACILLUS SUBTILIS. J Bacteriol. 1961 May;81(5):741–746. doi: 10.1128/jb.81.5.741-746.1961. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Dubnau D. Genetic competence in Bacillus subtilis. Microbiol Rev. 1991 Sep;55(3):395–424. doi: 10.1128/mr.55.3.395-424.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Dubnau D., Hahn J., Roggiani M., Piazza F., Weinrauch Y. Two-component regulators and genetic competence in Bacillus subtilis. Res Microbiol. 1994 Jun-Aug;145(5-6):403–411. doi: 10.1016/0923-2508(94)90088-4. [DOI] [PubMed] [Google Scholar]
  4. Dubnau D., Roggiani M. Growth medium-independent genetic competence mutants of Bacillus subtilis. J Bacteriol. 1990 Jul;172(7):4048–4055. doi: 10.1128/jb.172.7.4048-4055.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Dubnau D. The regulation of genetic competence in Bacillus subtilis. Mol Microbiol. 1991 Jan;5(1):11–18. doi: 10.1111/j.1365-2958.1991.tb01820.x. [DOI] [PubMed] [Google Scholar]
  6. Foster S. J. Analysis of the autolysins of Bacillus subtilis 168 during vegetative growth and differentiation by using renaturing polyacrylamide gel electrophoresis. J Bacteriol. 1992 Jan;174(2):464–470. doi: 10.1128/jb.174.2.464-470.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Hahn J., Bylund J., Haines M., Higgins M., Dubnau D. Inactivation of mecA prevents recovery from the competent state and interferes with cell division and the partitioning of nucleoids in Bacillus subtilis. Mol Microbiol. 1995 Nov;18(4):755–767. doi: 10.1111/j.1365-2958.1995.mmi_18040755.x. [DOI] [PubMed] [Google Scholar]
  8. Hahn J., Kong L., Dubnau D. The regulation of competence transcription factor synthesis constitutes a critical control point in the regulation of competence in Bacillus subtilis. J Bacteriol. 1994 Sep;176(18):5753–5761. doi: 10.1128/jb.176.18.5753-5761.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Hahn J., Roggiani M., Dubnau D. The major role of Spo0A in genetic competence is to downregulate abrB, an essential competence gene. J Bacteriol. 1995 Jun;177(12):3601–3605. doi: 10.1128/jb.177.12.3601-3605.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Haldenwang W. G. The sigma factors of Bacillus subtilis. Microbiol Rev. 1995 Mar;59(1):1–30. doi: 10.1128/mr.59.1.1-30.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Helmann J. D., Márquez L. M., Chamberlin M. J. Cloning, sequencing, and disruption of the Bacillus subtilis sigma 28 gene. J Bacteriol. 1988 Apr;170(4):1568–1574. doi: 10.1128/jb.170.4.1568-1574.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Herbold D. R., Glaser L. Bacillus subtilis N-acetylmuramic acid L-alanine amidase. J Biol Chem. 1975 Mar 10;250(5):1676–1682. [PubMed] [Google Scholar]
  13. Kong L., Dubnau D. Regulation of competence-specific gene expression by Mec-mediated protein-protein interaction in Bacillus subtilis. Proc Natl Acad Sci U S A. 1994 Jun 21;91(13):5793–5797. doi: 10.1073/pnas.91.13.5793. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Kong L., Siranosian K. J., Grossman A. D., Dubnau D. Sequence and properties of mecA, a negative regulator of genetic competence in Bacillus subtilis. Mol Microbiol. 1993 Jul;9(2):365–373. doi: 10.1111/j.1365-2958.1993.tb01697.x. [DOI] [PubMed] [Google Scholar]
  15. Krüger E., Völker U., Hecker M. Stress induction of clpC in Bacillus subtilis and its involvement in stress tolerance. J Bacteriol. 1994 Jun;176(11):3360–3367. doi: 10.1128/jb.176.11.3360-3367.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Kunst F., Msadek T., Bignon J., Rapoport G. The DegS/DegU and ComP/ComA two-component systems are part of a network controlling degradative enzyme synthesis and competence in Bacillus subtilis. Res Microbiol. 1994 Jun-Aug;145(5-6):393–402. doi: 10.1016/0923-2508(94)90087-6. [DOI] [PubMed] [Google Scholar]
  17. Kuroda A., Rashid M. H., Sekiguchi J. Molecular cloning and sequencing of the upstream region of the major Bacillus subtilis autolysin gene: a modifier protein exhibiting sequence homology to the major autolysin and the spoIID product. J Gen Microbiol. 1992 Jun;138(6):1067–1076. doi: 10.1099/00221287-138-6-1067. [DOI] [PubMed] [Google Scholar]
  18. Kuroda A., Sekiguchi J. High-level transcription of the major Bacillus subtilis autolysin operon depends on expression of the sigma D gene and is affected by a sin (flaD) mutation. J Bacteriol. 1993 Feb;175(3):795–801. doi: 10.1128/jb.175.3.795-801.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Kuroda A., Sekiguchi J. Molecular cloning and sequencing of a major Bacillus subtilis autolysin gene. J Bacteriol. 1991 Nov;173(22):7304–7312. doi: 10.1128/jb.173.22.7304-7312.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  21. Lazarevic V., Margot P., Soldo B., Karamata D. Sequencing and analysis of the Bacillus subtilis lytRABC divergon: a regulatory unit encompassing the structural genes of the N-acetylmuramoyl-L-alanine amidase and its modifier. J Gen Microbiol. 1992 Sep;138(9):1949–1961. doi: 10.1099/00221287-138-9-1949. [DOI] [PubMed] [Google Scholar]
  22. Mandic-Mulec I., Doukhan L., Smith I. The Bacillus subtilis SinR protein is a repressor of the key sporulation gene spo0A. J Bacteriol. 1995 Aug;177(16):4619–4627. doi: 10.1128/jb.177.16.4619-4627.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Mandic-Mulec I., Gaur N., Bai U., Smith I. Sin, a stage-specific repressor of cellular differentiation. J Bacteriol. 1992 Jun;174(11):3561–3569. doi: 10.1128/jb.174.11.3561-3569.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Margot P., Mauël C., Karamata D. The gene of the N-acetylglucosaminidase, a Bacillus subtilis 168 cell wall hydrolase not involved in vegetative cell autolysis. Mol Microbiol. 1994 May;12(4):535–545. doi: 10.1111/j.1365-2958.1994.tb01040.x. [DOI] [PubMed] [Google Scholar]
  25. Mirel D. B., Chamberlin M. J. The Bacillus subtilis flagellin gene (hag) is transcribed by the sigma 28 form of RNA polymerase. J Bacteriol. 1989 Jun;171(6):3095–3101. doi: 10.1128/jb.171.6.3095-3101.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Mohan S., Dubnau D. Transcriptional regulation of comC: evidence for a competence-specific transcription factor in Bacillus subtilis. J Bacteriol. 1990 Jul;172(7):4064–4071. doi: 10.1128/jb.172.7.4064-4071.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Msadek T., Kunst F., Rapoport G. MecB of Bacillus subtilis, a member of the ClpC ATPase family, is a pleiotropic regulator controlling competence gene expression and growth at high temperature. Proc Natl Acad Sci U S A. 1994 Jun 21;91(13):5788–5792. doi: 10.1073/pnas.91.13.5788. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Márquez-Magaña L. M., Chamberlin M. J. Characterization of the sigD transcription unit of Bacillus subtilis. J Bacteriol. 1994 Apr;176(8):2427–2434. doi: 10.1128/jb.176.8.2427-2434.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Márquez L. M., Helmann J. D., Ferrari E., Parker H. M., Ordal G. W., Chamberlin M. J. Studies of sigma D-dependent functions in Bacillus subtilis. J Bacteriol. 1990 Jun;172(6):3435–3443. doi: 10.1128/jb.172.6.3435-3443.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Rashid M. H., Kuroda A., Sekiguchi J. Bacillus subtilis mutant deficient in the major autolytic amidase and glucosaminidase is impaired in motility. FEMS Microbiol Lett. 1993 Sep 1;112(2):135–140. doi: 10.1111/j.1574-6968.1993.tb06438.x. [DOI] [PubMed] [Google Scholar]
  31. Rashid M. H., Mori M., Sekiguchi J. Glucosaminidase of Bacillus subtilis: cloning, regulation, primary structure and biochemical characterization. Microbiology. 1995 Oct;141(Pt 10):2391–2404. doi: 10.1099/13500872-141-10-2391. [DOI] [PubMed] [Google Scholar]
  32. Rogers H. J., Taylor C., Rayter S., Ward J. B. Purification and properties of autolytic endo-beta-N-acetylglucosaminidase and the N-acetylmuramyl-L-alanine amidase from Bacillus subtilis strain 168. J Gen Microbiol. 1984 Sep;130(9):2395–2402. doi: 10.1099/00221287-130-9-2395. [DOI] [PubMed] [Google Scholar]
  33. Roggiani M., Hahn J., Dubnau D. Suppression of early competence mutations in Bacillus subtilis by mec mutations. J Bacteriol. 1990 Jul;172(7):4056–4063. doi: 10.1128/jb.172.7.4056-4063.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Sekiguchi J., Ezaki B., Kodama K., Akamatsu T. Molecular cloning of a gene affecting the autolysin level and flagellation in Bacillus subtilis. J Gen Microbiol. 1988 Jun;134(6):1611–1621. doi: 10.1099/00221287-134-6-1611. [DOI] [PubMed] [Google Scholar]
  35. Sekiguchi J., Ohsu H., Kuroda A., Moriyama H., Akamatsu T. Nucleotide sequences of the Bacillus subtilis flaD locus and a B. licheniformis homologue affecting the autolysin level and flagellation. J Gen Microbiol. 1990 Jul;136(7):1223–1230. doi: 10.1099/00221287-136-7-1223. [DOI] [PubMed] [Google Scholar]
  36. Tokunaga T., Rashid M. H., Kuroda A., Sekiguchi J. Effect of degS-degU mutations on the expression of sigD, encoding an alternative sigma factor, and autolysin operon of Bacillus subtilis. J Bacteriol. 1994 Aug;176(16):5177–5180. doi: 10.1128/jb.176.16.5177-5180.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. van Sinderen D., Luttinger A., Kong L., Dubnau D., Venema G., Hamoen L. comK encodes the competence transcription factor, the key regulatory protein for competence development in Bacillus subtilis. Mol Microbiol. 1995 Feb;15(3):455–462. doi: 10.1111/j.1365-2958.1995.tb02259.x. [DOI] [PubMed] [Google Scholar]
  38. van Sinderen D., Venema G. comK acts as an autoregulatory control switch in the signal transduction route to competence in Bacillus subtilis. J Bacteriol. 1994 Sep;176(18):5762–5770. doi: 10.1128/jb.176.18.5762-5770.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. van Sinderen D., ten Berge A., Hayema B. J., Hamoen L., Venema G. Molecular cloning and sequence of comK, a gene required for genetic competence in Bacillus subtilis. Mol Microbiol. 1994 Feb;11(4):695–703. doi: 10.1111/j.1365-2958.1994.tb00347.x. [DOI] [PubMed] [Google Scholar]

Articles from Journal of Bacteriology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES