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. 1988 Jun;170(6):2549–2554. doi: 10.1128/jb.170.6.2549-2554.1988

Perturbed chromosomal replication in recA mutants of Escherichia coli.

K Skarstad 1, E Boye 1
PMCID: PMC211170  PMID: 3286612

Abstract

When initiation of DNA replication is inhibited in wild-type Escherichia coli cells by rifampin or chloramphenicol, completion of ongoing rounds of replication (runout of replication) leads to cells containing two, four, or eight fully replicated chromosomes, as measured by flow cytometry. In recombination-deficient recA strains, a high frequency of cells with three, five, six, or seven fully replicated chromosomes was observed in addition to cells with two, four, or eight chromosomes. recA mutants affected only in the protease-stimulating function behaved like wild-type cells. Thus, in the absence of the recombinase function of RecA protein, the frequency of productive initiations was significantly reduced compared with that in its presence. DNA degradation during runout of replication in the presence of rifampin was about 15%. The DNA degradation necessary to account for the whole effect described above was in this range or even lower. However, a model involving selective and complete degradation of partially replicated chromosomes is considered unlikely. It is suggested that the lack of RecA protein causes initiations or newly formed replication forks to stall but remain reactivatable for a period of time by functional RecA protein.

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Selected References

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

  1. Atlung T., Løbner-Olesen A., Hansen F. G. Overproduction of DnaA protein stimulates initiation of chromosome and minichromosome replication in Escherichia coli. Mol Gen Genet. 1987 Jan;206(1):51–59. doi: 10.1007/BF00326535. [DOI] [PubMed] [Google Scholar]
  2. Bianchi M., Riboli B., Magni G. E. coli recA protein possesses a strand separating activity on short duplex DNAs. EMBO J. 1985 Nov;4(11):3025–3030. doi: 10.1002/j.1460-2075.1985.tb04039.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Blanco M., Herrera G., Collado P., Rebollo J. E., Botella L. M. Influence of RecA protein on induced mutagenesis. Biochimie. 1982 Aug-Sep;64(8-9):633–636. doi: 10.1016/s0300-9084(82)80102-8. [DOI] [PubMed] [Google Scholar]
  4. Boye E., Steen H. B., Skarstad K. Flow cytometry of bacteria: a promising tool in experimental and clinical microbiology. J Gen Microbiol. 1983 Apr;129(4):973–980. doi: 10.1099/00221287-129-4-973. [DOI] [PubMed] [Google Scholar]
  5. Clark A. J. Recombination deficient mutants of E. coli and other bacteria. Annu Rev Genet. 1973;7:67–86. doi: 10.1146/annurev.ge.07.120173.000435. [DOI] [PubMed] [Google Scholar]
  6. Cooper S., Helmstetter C. E. Chromosome replication and the division cycle of Escherichia coli B/r. J Mol Biol. 1968 Feb 14;31(3):519–540. doi: 10.1016/0022-2836(68)90425-7. [DOI] [PubMed] [Google Scholar]
  7. Donachie W. D. Relationship between cell size and time of initiation of DNA replication. Nature. 1968 Sep 7;219(5158):1077–1079. doi: 10.1038/2191077a0. [DOI] [PubMed] [Google Scholar]
  8. Glickman B. W., Radman M. Escherichia coli mutator mutants deficient in methylation-instructed DNA mismatch correction. Proc Natl Acad Sci U S A. 1980 Feb;77(2):1063–1067. doi: 10.1073/pnas.77.2.1063. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Kogoma T. RNase H-defective mutants of Escherichia coli. J Bacteriol. 1986 May;166(2):361–363. doi: 10.1128/jb.166.2.361-363.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Kogoma T., Skarstad K., Boye E., von Meyenburg K., Steen H. B. RecA protein acts at the initiation of stable DNA replication in rnh mutants of Escherichia coli K-12. J Bacteriol. 1985 Aug;163(2):439–444. doi: 10.1128/jb.163.2.439-444.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Koppes L. J. OriC plasmids do not affect timing of chromosome replication in Escherichia coli K12. Mol Gen Genet. 1987 Aug;209(1):188–192. doi: 10.1007/BF00329857. [DOI] [PubMed] [Google Scholar]
  12. Lark K. G. Evidence for the direct involvement of RNA in the initiation of DNA replication in Escherichia coli 15T. J Mol Biol. 1972 Feb 28;64(1):47–60. doi: 10.1016/0022-2836(72)90320-8. [DOI] [PubMed] [Google Scholar]
  13. Little J. W., Mount D. W. The SOS regulatory system of Escherichia coli. Cell. 1982 May;29(1):11–22. doi: 10.1016/0092-8674(82)90085-x. [DOI] [PubMed] [Google Scholar]
  14. Lloyd R. G., Low B., Godson G. N., Birge E. A. Isolation and characterization of an Escherichia coli K-12 mutant with a temperature-sensitive recA- phenotype. J Bacteriol. 1974 Oct;120(1):407–415. doi: 10.1128/jb.120.1.407-415.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. MAALOE O., HANAWALT P. C. Thymine deficiency and the normal DNA replication cycle. I. J Mol Biol. 1961 Apr;3:144–155. doi: 10.1016/s0022-2836(61)80041-7. [DOI] [PubMed] [Google Scholar]
  16. Messer W. Initiation of deoxyribonucleic acid replication in Escherichia coli B-r: chronology of events and transcriptional control of initiation. J Bacteriol. 1972 Oct;112(1):7–12. doi: 10.1128/jb.112.1.7-12.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Radding C. M. Homologous pairing and strand exchange in genetic recombination. Annu Rev Genet. 1982;16:405–437. doi: 10.1146/annurev.ge.16.120182.002201. [DOI] [PubMed] [Google Scholar]
  18. Roberts J. W., Roberts C. W. Two mutations that alter the regulatory activity of E. coli recA protein. Nature. 1981 Apr 2;290(5805):422–424. doi: 10.1038/290422a0. [DOI] [PubMed] [Google Scholar]
  19. Schmidt L. S., Botta G., Park J. T. Effects of furazlocillin, a beta-lactam antibiotic which binds selectively to penicillin-binding protein 3, on Escherichia coli mutants deficient in other penicillin-binding proteins. J Bacteriol. 1981 Jan;145(1):632–637. doi: 10.1128/jb.145.1.632-637.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Skarstad K., Boye E., Steen H. B. Timing of initiation of chromosome replication in individual Escherichia coli cells. EMBO J. 1986 Jul;5(7):1711–1717. doi: 10.1002/j.1460-2075.1986.tb04415.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Skarstad K., Steen H. B., Boye E. Cell cycle parameters of slowly growing Escherichia coli B/r studied by flow cytometry. J Bacteriol. 1983 May;154(2):656–662. doi: 10.1128/jb.154.2.656-662.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Skarstad K., Steen H. B., Boye E. Escherichia coli DNA distributions measured by flow cytometry and compared with theoretical computer simulations. J Bacteriol. 1985 Aug;163(2):661–668. doi: 10.1128/jb.163.2.661-668.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Skarstad K., von Meyenburg K., Hansen F. G., Boye E. Coordination of chromosome replication initiation in Escherichia coli: effects of different dnaA alleles. J Bacteriol. 1988 Feb;170(2):852–858. doi: 10.1128/jb.170.2.852-858.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Slilaty S. N., Little J. W. Lysine-156 and serine-119 are required for LexA repressor cleavage: a possible mechanism. Proc Natl Acad Sci U S A. 1987 Jun;84(12):3987–3991. doi: 10.1073/pnas.84.12.3987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Slilaty S. N., Rupley J. A., Little J. W. Intramolecular cleavage of LexA and phage lambda repressors: dependence of kinetics on repressor concentration, pH, temperature, and solvent. Biochemistry. 1986 Nov 4;25(22):6866–6875. doi: 10.1021/bi00370a020. [DOI] [PubMed] [Google Scholar]
  26. Steen H. B. A microscope-based flow cytophotometer. Histochem J. 1983 Feb;15(2):147–160. doi: 10.1007/BF01042283. [DOI] [PubMed] [Google Scholar]
  27. Steen H. B., Lindmo T. Flow cytometry: a high-resolution instrument for everyone. Science. 1979 Apr 27;204(4391):403–404. doi: 10.1126/science.441727. [DOI] [PubMed] [Google Scholar]
  28. Torrey T. A., Kogoma T. Suppressor mutations (rin) that specifically suppress the recA+ dependence of stable DNA replication in Escherichia coliK-12. Mol Gen Genet. 1982;187(2):225–230. doi: 10.1007/BF00331121. [DOI] [PubMed] [Google Scholar]
  29. Willetts N. S., Clark A. J. Characteristics of some multiply recombination-deficient strains of Escherichia coli. J Bacteriol. 1969 Oct;100(1):231–239. doi: 10.1128/jb.100.1.231-239.1969. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Williams J. G., Shibata T., Radding C. M. Escherichia coli recA protein protects single-stranded DNA or gapped duplex DNA from degradation by RecBC DNase. J Biol Chem. 1981 Jul 25;256(14):7573–7582. [PubMed] [Google Scholar]
  31. Witkin E. M. Ultraviolet mutagenesis and inducible DNA repair in Escherichia coli. Bacteriol Rev. 1976 Dec;40(4):869–907. doi: 10.1128/br.40.4.869-907.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. von Meyenburg K., Boye E., Skarstad K., Koppes L., Kogoma T. Mode of initiation of constitutive stable DNA replication in RNase H-defective mutants of Escherichia coli K-12. J Bacteriol. 1987 Jun;169(6):2650–2658. doi: 10.1128/jb.169.6.2650-2658.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]

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