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. 1989 Aug;171(8):4121–4129. doi: 10.1128/jb.171.8.4121-4129.1989

Identification and characterization of genes controlled by the sporulation-regulatory gene spo0H in Bacillus subtilis.

K J Jaacks 1, J Healy 1, R Losick 1, A D Grossman 1
PMCID: PMC210181  PMID: 2502532

Abstract

We describe a general strategy for the identification of genes that are controlled by a specific regulatory factor in vivo and the use of this strategy to identify genes in Bacillus subtilis that are controlled by spo0H, a regulatory gene required for the initiation of sporulation. The general strategy makes use of a cloned regulatory gene fused to an inducible promoter to control expression of the regulatory gene and random gene fusions to a reporter gene to monitor expression in the presence and absence of the regulatory gene product. spo0H encodes a sigma factor of RNA polymerase, sigma H, and is required for the extensive reprograming of gene expression during the transition from growth to stationary phase and during the initiation of sporulation. We identified 18 genes that are controlled by sigma H (csh genes) in vivo by monitoring expression of random gene fusions to lacZ, made by insertion mutagenesis with the transposon Tn917lac, in the presence and absence of sigma H. These genes had lower levels of expression in the absence of sigma H than in the presence of sigma H. Patterns of expression of the csh genes during growth and sporulation in wild-type and spo0H mutant cells indicated that other regulatory factors are probably involved in controlling expression of some of these genes. Three of the csh::Tn917lac insertion mutations caused noticeable phenotypes. One caused a defect in vegetative growth, but only in combination with a spo0H mutation. Two others caused a partial defect in sporulation. One of these also caused a defect in the development of genetic competence. Detailed characterization of some of the csh genes and their regulatory regions should help define the role of spo0H in the regulation of gene expression during the transition from growth to stationary phase and during the initiation of sporulation.

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Selected References

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  1. Albano M., Hahn J., Dubnau D. Expression of competence genes in Bacillus subtilis. J Bacteriol. 1987 Jul;169(7):3110–3117. doi: 10.1128/jb.169.7.3110-3117.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Banner C. D., Moran C. P., Jr, Losick R. Deletion analysis of a complex promoter for a developmentally regulated gene from Bacillus subtilis. J Mol Biol. 1983 Aug 5;168(2):351–365. doi: 10.1016/s0022-2836(83)80023-0. [DOI] [PubMed] [Google Scholar]
  3. Carter H. L., 3rd, Moran C. P., Jr New RNA polymerase sigma factor under spo0 control in Bacillus subtilis. Proc Natl Acad Sci U S A. 1986 Dec;83(24):9438–9442. doi: 10.1073/pnas.83.24.9438. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Carter H. L., 3rd, Wang L. F., Doi R. H., Moran C. P., Jr rpoD operon promoter used by sigma H-RNA polymerase in Bacillus subtilis. J Bacteriol. 1988 Apr;170(4):1617–1621. doi: 10.1128/jb.170.4.1617-1621.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Connors M. J., Mason J. M., Setlow P. Cloning and nucleotide sequencing of genes for three small, acid-soluble proteins from Bacillus subtilis spores. J Bacteriol. 1986 May;166(2):417–425. doi: 10.1128/jb.166.2.417-425.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Connors M. J., Setlow P. Cloning of a small, acid-soluble spore protein gene from Bacillus subtilis and determination of its complete nucleotide sequence. J Bacteriol. 1985 Jan;161(1):333–339. doi: 10.1128/jb.161.1.333-339.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Dedonder R. A., Lepesant J. A., Lepesant-Kejzlarová J., Billault A., Steinmetz M., Kunst F. Construction of a kit of reference strains for rapid genetic mapping in Bacillus subtilis 168. Appl Environ Microbiol. 1977 Apr;33(4):989–993. doi: 10.1128/aem.33.4.989-993.1977. [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. 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]
  10. Dubnau E., Weir J., Nair G., Carter L., 3rd, Moran C., Jr, Smith I. Bacillus sporulation gene spo0H codes for sigma 30 (sigma H). J Bacteriol. 1988 Mar;170(3):1054–1062. doi: 10.1128/jb.170.3.1054-1062.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Feavers I. M., Price V., Moir A. The regulation of the fumarase (citG) gene of Bacillus subtilis 168. Mol Gen Genet. 1988 Mar;211(3):465–471. doi: 10.1007/BF00425702. [DOI] [PubMed] [Google Scholar]
  12. Ferrari F. A., Nguyen A., Lang D., Hoch J. A. Construction and properties of an integrable plasmid for Bacillus subtilis. J Bacteriol. 1983 Jun;154(3):1513–1515. doi: 10.1128/jb.154.3.1513-1515.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Freese E., Heinze J. E., Galliers E. M. Partial purine deprivation causes sporulation of Bacillus subtilis in the presence of excess ammonia, glucose and phosphate. J Gen Microbiol. 1979 Nov;115(1):193–205. doi: 10.1099/00221287-115-1-193. [DOI] [PubMed] [Google Scholar]
  14. Grossman A. D., Losick R. Extracellular control of spore formation in Bacillus subtilis. Proc Natl Acad Sci U S A. 1988 Jun;85(12):4369–4373. doi: 10.1073/pnas.85.12.4369. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Guzmán P., Westpheling J., Youngman P. Characterization of the promoter region of the Bacillus subtilis spoIIE operon. J Bacteriol. 1988 Apr;170(4):1598–1609. doi: 10.1128/jb.170.4.1598-1609.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Hahn J., Albano M., Dubnau D. Isolation and characterization of Tn917lac-generated competence mutants of Bacillus subtilis. J Bacteriol. 1987 Jul;169(7):3104–3109. doi: 10.1128/jb.169.7.3104-3109.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Hoch J. A. Genetic analysis of pleiotropic negative sporulation mutants in Bacillus subtilis. J Bacteriol. 1971 Mar;105(3):896–901. doi: 10.1128/jb.105.3.896-901.1971. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Kenney T. J., Kirchman P. A., Moran C. P., Jr Gene encoding sigma E is transcribed from a sigma A-like promoter in Bacillus subtilis. J Bacteriol. 1988 Jul;170(7):3058–3064. doi: 10.1128/jb.170.7.3058-3064.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. 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]
  20. Lopez J. M., Dromerick A., Freese E. Response of guanosine 5'-triphosphate concentration to nutritional changes and its significance for Bacillus subtilis sporulation. J Bacteriol. 1981 May;146(2):605–613. doi: 10.1128/jb.146.2.605-613.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Lopez J. M., Marks C. L., Freese E. The decrease of guanine nucleotides initiates sporulation of Bacillus subtilis. Biochim Biophys Acta. 1979 Oct 4;587(2):238–252. doi: 10.1016/0304-4165(79)90357-x. [DOI] [PubMed] [Google Scholar]
  22. Losick R., Youngman P., Piggot P. J. Genetics of endospore formation in Bacillus subtilis. Annu Rev Genet. 1986;20:625–669. doi: 10.1146/annurev.ge.20.120186.003205. [DOI] [PubMed] [Google Scholar]
  23. Love P. E., Lyle M. J., Yasbin R. E. DNA-damage-inducible (din) loci are transcriptionally activated in competent Bacillus subtilis. Proc Natl Acad Sci U S A. 1985 Sep;82(18):6201–6205. doi: 10.1073/pnas.82.18.6201. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Mitani T., Heinze J. E., Freese E. Induction of sporulation in Bacillus subtilis by decoyinine or hadacidin. Biochem Biophys Res Commun. 1977 Aug 8;77(3):1118–1125. doi: 10.1016/s0006-291x(77)80094-6. [DOI] [PubMed] [Google Scholar]
  25. Ochi K., Ohsawa S. Initiation of antibiotic production by the stringent response of Bacillus subtilis Marburg. J Gen Microbiol. 1984 Oct;130(10):2473–2482. doi: 10.1099/00221287-130-10-2473. [DOI] [PubMed] [Google Scholar]
  26. Perkins J. B., Youngman P. J. Construction and properties of Tn917-lac, a transposon derivative that mediates transcriptional gene fusions in Bacillus subtilis. Proc Natl Acad Sci U S A. 1986 Jan;83(1):140–144. doi: 10.1073/pnas.83.1.140. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Piggot P. J., Coote J. G. Genetic aspects of bacterial endospore formation. Bacteriol Rev. 1976 Dec;40(4):908–962. doi: 10.1128/br.40.4.908-962.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Piggot P. J., Hoch J. A. Revised genetic linkage map of Bacillus subtilis. Microbiol Rev. 1985 Jun;49(2):158–179. doi: 10.1128/mr.49.2.158-179.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Sandman K., Losick R., Youngman P. Genetic analysis of Bacillus subtilis spo mutations generated by Tn917-mediated insertional mutagenesis. Genetics. 1987 Dec;117(4):603–617. doi: 10.1093/genetics/117.4.603. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Schaeffer P., Millet J., Aubert J. P. Catabolic repression of bacterial sporulation. Proc Natl Acad Sci U S A. 1965 Sep;54(3):704–711. doi: 10.1073/pnas.54.3.704. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Segall J., Losick R. Cloned Bacillus subtilis DNA containing a gene that is activated early during sporulation. Cell. 1977 Aug;11(4):751–761. doi: 10.1016/0092-8674(77)90289-6. [DOI] [PubMed] [Google Scholar]
  32. Vasantha N., Freese E. Enzyme changes during Bacillus subtilis sporulation caused by deprivation of guanine nucleotides. J Bacteriol. 1980 Dec;144(3):1119–1125. doi: 10.1128/jb.144.3.1119-1125.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Vasantha N., Freese E. The role of manganese in growth and sporulation of Bacillus subtilis. J Gen Microbiol. 1979 Jun;112(2):329–336. doi: 10.1099/00221287-112-2-329. [DOI] [PubMed] [Google Scholar]
  34. Wu J. J., Howard M. G., Piggot P. J. Regulation of transcription of the Bacillus subtilis spoIIA locus. J Bacteriol. 1989 Feb;171(2):692–698. doi: 10.1128/jb.171.2.692-698.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Yamashita S., Yoshikawa H., Kawamura F., Takahashi H., Yamamoto T., Kobayashi Y., Saito H. The effect of spo0 mutations on the expression of spo0A- and spo0F-lacZ fusions. Mol Gen Genet. 1986 Oct;205(1):28–33. doi: 10.1007/BF02428029. [DOI] [PubMed] [Google Scholar]
  36. Yansura D. G., Henner D. J. Use of the Escherichia coli lac repressor and operator to control gene expression in Bacillus subtilis. Proc Natl Acad Sci U S A. 1984 Jan;81(2):439–443. doi: 10.1073/pnas.81.2.439. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Youngman P., Perkins J. B., Losick R. Construction of a cloning site near one end of Tn917 into which foreign DNA may be inserted without affecting transposition in Bacillus subtilis or expression of the transposon-borne erm gene. Plasmid. 1984 Jul;12(1):1–9. doi: 10.1016/0147-619x(84)90061-1. [DOI] [PubMed] [Google Scholar]
  38. Youngman P., Zuber P., Perkins J. B., Sandman K., Igo M., Losick R. New ways to study developmental genes in spore-forming bacteria. Science. 1985 Apr 19;228(4697):285–291. doi: 10.1126/science.228.4697.285. [DOI] [PubMed] [Google Scholar]
  39. Zuber P., Healy J. M., Losick R. Effects of plasmid propagation of a sporulation promoter on promoter utilization and sporulation in Bacillus subtilis. J Bacteriol. 1987 Feb;169(2):461–469. doi: 10.1128/jb.169.2.461-469.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Zuber P., Healy J., Carter H. L., 3rd, Cutting S., Moran C. P., Jr, Losick R. Mutation changing the specificity of an RNA polymerase sigma factor. J Mol Biol. 1989 Apr 20;206(4):605–614. doi: 10.1016/0022-2836(89)90569-x. [DOI] [PubMed] [Google Scholar]
  41. 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]
  42. Zuber P., Losick R. Use of a lacZ fusion to study the role of the spoO genes of Bacillus subtilis in developmental regulation. Cell. 1983 Nov;35(1):275–283. doi: 10.1016/0092-8674(83)90230-1. [DOI] [PubMed] [Google Scholar]

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