Skip to main content
PMC home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1997 Dec;179(23):7351–7359. doi: 10.1128/jb.179.23.7351-7359.1997

A null mutation in the Bacillus subtilis aconitase gene causes a block in Spo0A-phosphate-dependent gene expression.

J E Craig 1, M J Ford 1, D C Blaydon 1, A L Sonenshein 1
PMCID: PMC179685  PMID: 9393699

Abstract

The citB gene of Bacillus subtilis encodes aconitase, the enzyme of the Krebs citric acid cycle, which is responsible for the interconversion of citrate and isocitrate. A B. subtilis strain with an insertion mutation in the citB gene was devoid of aconitase activity and aconitase protein, required glutamate for growth in minimal medium, and was unable to sporulate efficiently in nutrient broth sporulation medium. Mutant cells failed to form the asymmetric septum characteristic of sporulating cells and were defective in transcription of the earliest-expressed spo genes, that is, the genes dependent on the Spo0A phosphorelay. However, this early block in sporulation was partially overcome when cells of the citB mutant were induced to sporulate by resuspension in a poor medium. Accumulation of citrate in the mutant cells or in their culture fluid may be responsible for the early block, possibly because citrate can chelate divalent cations needed for the activity of the phosphorelay.

Full Text

The Full Text of this article is available as a PDF (921.1 KB).

Selected References

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

  1. Belitsky B. R., Janssen P. J., Sonenshein A. L. Sites required for GltC-dependent regulation of Bacillus subtilis glutamate synthase expression. J Bacteriol. 1995 Oct;177(19):5686–5695. doi: 10.1128/jb.177.19.5686-5695.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Burbulys D., Trach K. A., Hoch J. A. Initiation of sporulation in B. subtilis is controlled by a multicomponent phosphorelay. Cell. 1991 Feb 8;64(3):545–552. doi: 10.1016/0092-8674(91)90238-t. [DOI] [PubMed] [Google Scholar]
  3. CHARNEY J., FISHER W. P., HEGARTY C. P. Managanese as an essential element for sporulation in the genus Bacillus. J Bacteriol. 1951 Aug;62(2):145–148. doi: 10.1128/jb.62.2.145-148.1951. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Craig J. E., Zhang Y., Gallagher M. P. Cloning of the nupC gene of Escherichia coli encoding a nucleoside transport system, and identification of an adjacent insertion element, IS 186. Mol Microbiol. 1994 Mar;11(6):1159–1168. doi: 10.1111/j.1365-2958.1994.tb00392.x. [DOI] [PubMed] [Google Scholar]
  5. Cutting S., Oke V., Driks A., Losick R., Lu S., Kroos L. A forespore checkpoint for mother cell gene expression during development in B. subtilis. Cell. 1990 Jul 27;62(2):239–250. doi: 10.1016/0092-8674(90)90362-i. [DOI] [PubMed] [Google Scholar]
  6. Dingman D. W., Rosenkrantz M. S., Sonenshein A. L. Relationship between aconitase gene expression and sporulation in Bacillus subtilis. J Bacteriol. 1987 Jul;169(7):3068–3075. doi: 10.1128/jb.169.7.3068-3075.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Dingman D. W., Sonenshein A. L. Purification of aconitase from Bacillus subtilis and correlation of its N-terminal amino acid sequence with the sequence of the citB gene. J Bacteriol. 1987 Jul;169(7):3062–3067. doi: 10.1128/jb.169.7.3062-3067.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Donovan W., Zheng L. B., Sandman K., Losick R. Genes encoding spore coat polypeptides from Bacillus subtilis. J Mol Biol. 1987 Jul 5;196(1):1–10. doi: 10.1016/0022-2836(87)90506-7. [DOI] [PubMed] [Google Scholar]
  9. Dower W. J., Miller J. F., Ragsdale C. W. High efficiency transformation of E. coli by high voltage electroporation. Nucleic Acids Res. 1988 Jul 11;16(13):6127–6145. doi: 10.1093/nar/16.13.6127. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Dubnau D., Davidoff-Abelson R. Fate of transforming DNA following uptake by competent Bacillus subtilis. I. Formation and properties of the donor-recipient complex. J Mol Biol. 1971 Mar 14;56(2):209–221. doi: 10.1016/0022-2836(71)90460-8. [DOI] [PubMed] [Google Scholar]
  11. Fortnagel P., Freese E. Analysis of sporulation mutants. II. Mutants blocked in the citric acid cycle. J Bacteriol. 1968 Apr;95(4):1431–1438. doi: 10.1128/jb.95.4.1431-1438.1968. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Fortnagel P., Freese E. Inhibition of aconitase by chelation of transition metals causing inhibition of sporulation in Bacillus subtilis. J Biol Chem. 1968 Oct 25;243(20):5289–5295. [PubMed] [Google Scholar]
  13. Fouet A., Sonenshein A. L. A target for carbon source-dependent negative regulation of the citB promoter of Bacillus subtilis. J Bacteriol. 1990 Feb;172(2):835–844. doi: 10.1128/jb.172.2.835-844.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Grossman A. D. Integration of developmental signals and the initiation of sporulation in B. subtilis. Cell. 1991 Apr 5;65(1):5–8. doi: 10.1016/0092-8674(91)90353-z. [DOI] [PubMed] [Google Scholar]
  15. He M., Kaderbhai M. A., Adcock I., Austen B. M. An improved and rapid procedure for isolating RNA-free Escherichia coli plasmid DNA. Genet Anal Tech Appl. 1991 May;8(3):107–110. doi: 10.1016/1050-3862(91)90045-s. [DOI] [PubMed] [Google Scholar]
  16. Hentze M. W., Kühn L. C. Molecular control of vertebrate iron metabolism: mRNA-based regulatory circuits operated by iron, nitric oxide, and oxidative stress. Proc Natl Acad Sci U S A. 1996 Aug 6;93(16):8175–8182. doi: 10.1073/pnas.93.16.8175. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Hoch J. A. Regulation of the phosphorelay and the initiation of sporulation in Bacillus subtilis. Annu Rev Microbiol. 1993;47:441–465. doi: 10.1146/annurev.mi.47.100193.002301. [DOI] [PubMed] [Google Scholar]
  18. Hoch J. A., Trach K., Kawamura F., Saito H. Identification of the transcriptional suppressor sof-1 as an alteration in the spo0A protein. J Bacteriol. 1985 Feb;161(2):552–555. doi: 10.1128/jb.161.2.552-555.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Ireton K., Grossman A. D. A developmental checkpoint couples the initiation of sporulation to DNA replication in Bacillus subtilis. EMBO J. 1994 Apr 1;13(7):1566–1573. doi: 10.1002/j.1460-2075.1994.tb06419.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Ireton K., Jin S., Grossman A. D., Sonenshein A. L. Krebs cycle function is required for activation of the Spo0A transcription factor in Bacillus subtilis. Proc Natl Acad Sci U S A. 1995 Mar 28;92(7):2845–2849. doi: 10.1073/pnas.92.7.2845. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Jin S., Levin P. A., Matsuno K., Grossman A. D., Sonenshein A. L. Deletion of the Bacillus subtilis isocitrate dehydrogenase gene causes a block at stage I of sporulation. J Bacteriol. 1997 Aug;179(15):4725–4732. doi: 10.1128/jb.179.15.4725-4732.1997. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Jin S., Sonenshein A. L. Identification of two distinct Bacillus subtilis citrate synthase genes. J Bacteriol. 1994 Aug;176(15):4669–4679. doi: 10.1128/jb.176.15.4669-4679.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Jin S., Sonenshein A. L. Transcriptional regulation of Bacillus subtilis citrate synthase genes. J Bacteriol. 1994 Aug;176(15):4680–4690. doi: 10.1128/jb.176.15.4680-4690.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Karmazyn-Campelli C., Bonamy C., Savelli B., Stragier P. Tandem genes encoding sigma-factors for consecutive steps of development in Bacillus subtilis. Genes Dev. 1989 Feb;3(2):150–157. doi: 10.1101/gad.3.2.150. [DOI] [PubMed] [Google Scholar]
  25. Kenney T. J., Moran C. P., Jr Organization and regulation of an operon that encodes a sporulation-essential sigma factor in Bacillus subtilis. J Bacteriol. 1987 Jul;169(7):3329–3339. doi: 10.1128/jb.169.7.3329-3339.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Knecht D. A., Dimond R. L. Visualization of antigenic proteins on Western blots. Anal Biochem. 1984 Jan;136(1):180–184. doi: 10.1016/0003-2697(84)90321-x. [DOI] [PubMed] [Google Scholar]
  27. Kobayashi K., Shoji K., Shimizu T., Nakano K., Sato T., Kobayashi Y. Analysis of a suppressor mutation ssb (kinC) of sur0B20 (spo0A) mutation in Bacillus subtilis reveals that kinC encodes a histidine protein kinase. J Bacteriol. 1995 Jan;177(1):176–182. doi: 10.1128/jb.177.1.176-182.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. 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]
  29. LeDeaux J. R., Grossman A. D. Isolation and characterization of kinC, a gene that encodes a sensor kinase homologous to the sporulation sensor kinases KinA and KinB in Bacillus subtilis. J Bacteriol. 1995 Jan;177(1):166–175. doi: 10.1128/jb.177.1.166-175.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Leyva-Vazquez M. A., Setlow P. Cloning and nucleotide sequences of the genes encoding triose phosphate isomerase, phosphoglycerate mutase, and enolase from Bacillus subtilis. J Bacteriol. 1994 Jul;176(13):3903–3910. doi: 10.1128/jb.176.13.3903-3910.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Londoño-Vallejo J. A., Fréhel C., Stragier P. SpoIIQ, a forespore-expressed gene required for engulfment in Bacillus subtilis. Mol Microbiol. 1997 Apr;24(1):29–39. doi: 10.1046/j.1365-2958.1997.3181680.x. [DOI] [PubMed] [Google Scholar]
  32. Mason J. M., Hackett R. H., Setlow P. Regulation of expression of genes coding for small, acid-soluble proteins of Bacillus subtilis spores: studies using lacZ gene fusions. J Bacteriol. 1988 Jan;170(1):239–244. doi: 10.1128/jb.170.1.239-244.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Mueller J. P., Bukusoglu G., Sonenshein A. L. Transcriptional regulation of Bacillus subtilis glucose starvation-inducible genes: control of gsiA by the ComP-ComA signal transduction system. J Bacteriol. 1992 Jul;174(13):4361–4373. doi: 10.1128/jb.174.13.4361-4373.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Oh Y. K., Freese E. Manganese requirement of phosphoglycerate phosphomutase and its consequences for growth and sporulation of Bacillus subtilis. J Bacteriol. 1976 Aug;127(2):739–746. doi: 10.1128/jb.127.2.739-746.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Ohné M. Regulation of aconitase synthesis in Bacillus subtilis: induction, feedback repression, and catabolite repression. J Bacteriol. 1974 Mar;117(3):1295–1305. doi: 10.1128/jb.117.3.1295-1305.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Perego M., Hoch J. A. Cell-cell communication regulates the effects of protein aspartate phosphatases on the phosphorelay controlling development in Bacillus subtilis. Proc Natl Acad Sci U S A. 1996 Feb 20;93(4):1549–1553. doi: 10.1073/pnas.93.4.1549. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Perego M., Spiegelman G. B., Hoch J. A. Structure of the gene for the transition state regulator, abrB: regulator synthesis is controlled by the spo0A sporulation gene in Bacillus subtilis. Mol Microbiol. 1988 Nov;2(6):689–699. doi: 10.1111/j.1365-2958.1988.tb00079.x. [DOI] [PubMed] [Google Scholar]
  38. Perego M., Wu J. J., Spiegelman G. B., Hoch J. A. Mutational dissociation of the positive and negative regulatory properties of the Spo0A sporulation transcription factor of Bacillus subtilis. Gene. 1991 Apr;100:207–212. doi: 10.1016/0378-1119(91)90368-l. [DOI] [PubMed] [Google Scholar]
  39. Rose M., Entian K. D. New genes in the 170 degrees region of the Bacillus subtilis genome encode DNA gyrase subunits, a thioredoxin, a xylanase and an amino acid transporter. Microbiology. 1996 Nov;142(Pt 11):3097–3101. doi: 10.1099/13500872-142-11-3097. [DOI] [PubMed] [Google Scholar]
  40. Rosenkrantz M. S., Dingman D. W., Sonenshein A. L. Bacillus subtilis citB gene is regulated synergistically by glucose and glutamine. J Bacteriol. 1985 Oct;164(1):155–164. doi: 10.1128/jb.164.1.155-164.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Rouault T. A., Klausner R. D. Iron-sulfur clusters as biosensors of oxidants and iron. Trends Biochem Sci. 1996 May;21(5):174–177. [PubMed] [Google Scholar]
  42. Satola S. W., Baldus J. M., Moran C. P., Jr Binding of Spo0A stimulates spoIIG promoter activity in Bacillus subtilis. J Bacteriol. 1992 Mar;174(5):1448–1453. doi: 10.1128/jb.174.5.1448-1453.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Satola S., Kirchman P. A., Moran C. P., Jr Spo0A binds to a promoter used by sigma A RNA polymerase during sporulation in Bacillus subtilis. Proc Natl Acad Sci U S A. 1991 May 15;88(10):4533–4537. doi: 10.1073/pnas.88.10.4533. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Setlow P. Small, acid-soluble spore proteins of Bacillus species: structure, synthesis, genetics, function, and degradation. Annu Rev Microbiol. 1988;42:319–338. doi: 10.1146/annurev.mi.42.100188.001535. [DOI] [PubMed] [Google Scholar]
  45. Sterlini J. M., Mandelstam J. Commitment to sporulation in Bacillus subtilis and its relationship to development of actinomycin resistance. Biochem J. 1969 Jun;113(1):29–37. doi: 10.1042/bj1130029. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Strauch M., Webb V., Spiegelman G., Hoch J. A. The SpoOA protein of Bacillus subtilis is a repressor of the abrB gene. Proc Natl Acad Sci U S A. 1990 Mar;87(5):1801–1805. doi: 10.1073/pnas.87.5.1801. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. York K., Kenney T. J., Satola S., Moran C. P., Jr, Poth H., Youngman P. Spo0A controls the sigma A-dependent activation of Bacillus subtilis sporulation-specific transcription unit spoIIE. J Bacteriol. 1992 Apr;174(8):2648–2658. doi: 10.1128/jb.174.8.2648-2658.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Yousten A. A., Hanson R. S. Sporulation of tricarboxylic acid cycle mutants of Bacillus subtilis. J Bacteriol. 1972 Feb;109(2):886–894. doi: 10.1128/jb.109.2.886-894.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Zuber P., Losick R. Role of AbrB in Spo0A- and Spo0B-dependent utilization of a sporulation promoter in Bacillus subtilis. J Bacteriol. 1987 May;169(5):2223–2230. doi: 10.1128/jb.169.5.2223-2230.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]

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

RESOURCES