Skip to main content
NCBI home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1993 Mar;175(6):1705–1716. doi: 10.1128/jb.175.6.1705-1716.1993

Cloning and characterization of a gene required for assembly of the Bacillus subtilis spore coat.

B Beall 1, A Driks 1, R Losick 1, C P Moran Jr 1
PMCID: PMC203965  PMID: 8449878

Abstract

During endospore formation in Bacillus subtilis, approximately a dozen proteins are synthesized and assembled around the prespore to form a protective coat. Little is known about the assembly process, but several of the genes encoding these coat proteins are expressed in the mother cell compartment, where the proteins accumulate on the outer side of the developing endospore. Transcription of these genes is directed by the mother cell-specific sigma factor, sigma K, during the later stages of endospore development. sigma E may direct expression of the genes that encode proteins that function in the earliest stages of coat assembly. By screening for sigma E-dependent promoters, we cloned a gene, designated spoVID, required for assembly of a normal spore coat. Expression of spoVID was initiated at about the second hour of sporulation and continued throughout development from a sigma E-dependent promoter. The spoVID gene was located on the B. subtilis chromosome just downstream of the previously characterized hemAXCDBL operon and is predicted to encode an extremely acidic protein with 575 residues. Insertion mutants of spoVID produced refractile spores that were resistant to heat and to chloroform but were sensitive to lysozyme. Electron microscopic examination of sporulating spoVID mutant cells revealed normal morphological development up to about the third hour of sporulation. However, during the later stages of development the coat proteins assembled into aberrant structures that occurred freely in the mother cell cytoplasm and that consisted of reiterations of the single inner and outer layers that normally make up the spore coat.

Full text

PDF
1705

Images in this article

1711Image
on p.1711
1713Image
on p.1713
1714Image
on p.1714

Selected References

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

  1. Aronson A. I., Song H. Y., Bourne N. Gene structure and precursor processing of a novel Bacillus subtilis spore coat protein. Mol Microbiol. 1989 Mar;3(3):437–444. doi: 10.1111/j.1365-2958.1989.tb00189.x. [DOI] [PubMed] [Google Scholar]
  2. Bourne N., FitzJames P. C., Aronson A. I. Structural and germination defects of Bacillus subtilis spores with altered contents of a spore coat protein. J Bacteriol. 1991 Oct;173(20):6618–6625. doi: 10.1128/jb.173.20.6618-6625.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Cutting S., Driks A., Schmidt R., Kunkel B., Losick R. Forespore-specific transcription of a gene in the signal transduction pathway that governs Pro-sigma K processing in Bacillus subtilis. Genes Dev. 1991 Mar;5(3):456–466. doi: 10.1101/gad.5.3.456. [DOI] [PubMed] [Google Scholar]
  4. 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]
  5. Cutting S., Zheng L. B., Losick R. Gene encoding two alkali-soluble components of the spore coat from Bacillus subtilis. J Bacteriol. 1991 May;173(9):2915–2919. doi: 10.1128/jb.173.9.2915-2919.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Diederich B., Tatti K. M., Jones C. H., Beall B., Moran C. P., Jr Genetic suppression analysis of sigma E interaction with three promoters in sporulating Bacillus subtilis. Gene. 1992 Nov 2;121(1):63–69. doi: 10.1016/0378-1119(92)90162-i. [DOI] [PubMed] [Google Scholar]
  7. 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]
  8. Driks A., Losick R. Compartmentalized expression of a gene under the control of sporulation transcription factor sigma E in Bacillus subtilis. Proc Natl Acad Sci U S A. 1991 Nov 15;88(22):9934–9938. doi: 10.1073/pnas.88.22.9934. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Errington J. A general method for fusion of the Escherichia coli lacZ gene to chromosomal genes in Bacillus subtilis. J Gen Microbiol. 1986 Nov;132(11):2953–2966. doi: 10.1099/00221287-132-11-2953. [DOI] [PubMed] [Google Scholar]
  10. Errington J., Illing N. Establishment of cell-specific transcription during sporulation in Bacillus subtilis. Mol Microbiol. 1992 Mar;6(6):689–695. doi: 10.1111/j.1365-2958.1992.tb01517.x. [DOI] [PubMed] [Google Scholar]
  11. 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]
  12. Hansson M., Rutberg L., Schröder I., Hederstedt L. The Bacillus subtilis hemAXCDBL gene cluster, which encodes enzymes of the biosynthetic pathway from glutamate to uroporphyrinogen III. J Bacteriol. 1991 Apr;173(8):2590–2599. doi: 10.1128/jb.173.8.2590-2599.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Jenkinson H. F. Altered arrangement of proteins in the spore coat of a germination mutant of Bacillus subtilis. J Gen Microbiol. 1983 Jun;129(6):1945–1958. doi: 10.1099/00221287-129-6-1945. [DOI] [PubMed] [Google Scholar]
  14. 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]
  15. 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]
  16. Kunkel B., Kroos L., Poth H., Youngman P., Losick R. Temporal and spatial control of the mother-cell regulatory gene spoIIID of Bacillus subtilis. Genes Dev. 1989 Nov;3(11):1735–1744. doi: 10.1101/gad.3.11.1735. [DOI] [PubMed] [Google Scholar]
  17. Kunkel B., Losick R., Stragier P. The Bacillus subtilis gene for the development transcription factor sigma K is generated by excision of a dispensable DNA element containing a sporulation recombinase gene. Genes Dev. 1990 Apr;4(4):525–535. doi: 10.1101/gad.4.4.525. [DOI] [PubMed] [Google Scholar]
  18. Losick R., Stragier P. Crisscross regulation of cell-type-specific gene expression during development in B. subtilis. Nature. 1992 Feb 13;355(6361):601–604. doi: 10.1038/355601a0. [DOI] [PubMed] [Google Scholar]
  19. Lu S., Halberg R., Kroos L. Processing of the mother-cell sigma factor, sigma K, may depend on events occurring in the forespore during Bacillus subtilis development. Proc Natl Acad Sci U S A. 1990 Dec;87(24):9722–9726. doi: 10.1073/pnas.87.24.9722. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. 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]
  21. Messing J. New M13 vectors for cloning. Methods Enzymol. 1983;101:20–78. doi: 10.1016/0076-6879(83)01005-8. [DOI] [PubMed] [Google Scholar]
  22. Moir A. Germination properties of a spore coat-defective mutant of Bacillus subtilis. J Bacteriol. 1981 Jun;146(3):1106–1116. doi: 10.1128/jb.146.3.1106-1116.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. 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]
  24. Rather P. N., Coppolecchia R., DeGrazia H., Moran C. P., Jr Negative regulator of sigma G-controlled gene expression in stationary-phase Bacillus subtilis. J Bacteriol. 1990 Feb;172(2):709–715. doi: 10.1128/jb.172.2.709-715.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Roels S., Driks A., Losick R. Characterization of spoIVA, a sporulation gene involved in coat morphogenesis in Bacillus subtilis. J Bacteriol. 1992 Jan;174(2):575–585. doi: 10.1128/jb.174.2.575-585.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Sandman K., Kroos L., Cutting S., Youngman P., Losick R. Identification of the promoter for a spore coat protein gene in Bacillus subtilis and studies on the regulation of its induction at a late stage of sporulation. J Mol Biol. 1988 Apr 5;200(3):461–473. doi: 10.1016/0022-2836(88)90536-0. [DOI] [PubMed] [Google Scholar]
  27. 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]
  28. Stevens C. M., Daniel R., Illing N., Errington J. Characterization of a sporulation gene, spoIVA, involved in spore coat morphogenesis in Bacillus subtilis. J Bacteriol. 1992 Jan;174(2):586–594. doi: 10.1128/jb.174.2.586-594.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Stragier P., Kunkel B., Kroos L., Losick R. Chromosomal rearrangement generating a composite gene for a developmental transcription factor. Science. 1989 Jan 27;243(4890):507–512. doi: 10.1126/science.2536191. [DOI] [PubMed] [Google Scholar]
  30. 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]
  31. Zheng L. B., Donovan W. P., Fitz-James P. C., Losick R. Gene encoding a morphogenic protein required in the assembly of the outer coat of the Bacillus subtilis endospore. Genes Dev. 1988 Aug;2(8):1047–1054. doi: 10.1101/gad.2.8.1047. [DOI] [PubMed] [Google Scholar]
  32. Zheng L. B., Losick R. Cascade regulation of spore coat gene expression in Bacillus subtilis. J Mol Biol. 1990 Apr 20;212(4):645–660. doi: 10.1016/0022-2836(90)90227-d. [DOI] [PubMed] [Google Scholar]
  33. Zheng L., Halberg R., Roels S., Ichikawa H., Kroos L., Losick R. Sporulation regulatory protein GerE from Bacillus subtilis binds to and can activate or repress transcription from promoters for mother-cell-specific genes. J Mol Biol. 1992 Aug 20;226(4):1037–1050. doi: 10.1016/0022-2836(92)91051-p. [DOI] [PubMed] [Google Scholar]
  34. 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