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. 1986 Nov;168(2):494–504. doi: 10.1128/jb.168.2.494-504.1986

DNA replication termination in Escherichia coli parB (a dnaG allele), parA, and gyrB mutants affected in DNA distribution.

V Norris, T Alliotte, A Jaffé, R D'Ari
PMCID: PMC213509  PMID: 3536848

Abstract

We investigated the Escherichia coli mutants carrying the parB, parA, and gyrB mutations, all of which display faulty chromosome partitioning at the nonpermissive temperature, to see whether their phenotype reflected a defect in the termination of DNA replication. In the parB strain DNA synthesis slowed down at 42 degrees C and the SOS response was induced, whereas in the parA strain DNA synthesis continued normally for 120 min and there was no SOS induction. To see whether replication forks accumulated in the vicinity of terC at the nonpermissive temperature, the mutants were incubated for 60 min at 42 degrees C and then returned to low temperature and pulse-labeled with [3H]thymidine. In all cases the restriction pattern of the labeled DNA was incompatible with that of the terC region, suggesting that replication termination was normal. In the parA mutant no DNA sequences were preferentially labeled, whereas in the parB and gyrB strains there was specific labeling of sequences whose restriction pattern resembled that of oriC. In the case of parB this was confirmed by DNA-DNA hybridization with appropriate probes. This test further revealed that the parB mutant over initiates at oriC after the return to the permissive temperature. Like dna(Ts) strains, the parB mutant formed filaments at 42 degrees C in the absence of SOS-associated division inhibition, accompanied by the appearance of anucleate cells of nearly normal size (28% of the population after 3 h), as revealed by autoradiography. The DNA in the filaments was either centrally located or distributed throughout. The parB mutation lies at 67 min, and the ParB- phenotype is corrected by a cloned dnaG gene or by a plasmid primase, strongly suggesting that parB is an allele of dnaG, the structural gene of the E. coli primase. It is thus likely that the parB mutant possesses an altered primase which does not affect replication termination but causes a partial defect in replication initiation and elongation and in chromosome distribution.

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Selected References

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  1. Arai K., Kornberg A. Unique primed start of phage phi X174 DNA replication and mobility of the primosome in a direction opposite chain synthesis. Proc Natl Acad Sci U S A. 1981 Jan;78(1):69–73. doi: 10.1073/pnas.78.1.69. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Botta G. A., Park J. T. Evidence for involvement of penicillin-binding protein 3 in murein synthesis during septation but not during cell elongation. J Bacteriol. 1981 Jan;145(1):333–340. doi: 10.1128/jb.145.1.333-340.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bouché J. P., Gélugne J. P., Louarn J., Louarn J. M., Kaiser K. Relationships between the physical and genetic maps of a 470 x 10(3) base-pair region around the terminus of Escherichia coli K12 DNA replication. J Mol Biol. 1982 Jan 5;154(1):21–32. doi: 10.1016/0022-2836(82)90414-4. [DOI] [PubMed] [Google Scholar]
  4. Bouché J. P. Physical map of a 470 x 10(3) base-pair region flanking the terminus of DNA replication in the Escherichia coli K12 genome. J Mol Biol. 1982 Jan 5;154(1):1–20. doi: 10.1016/0022-2836(82)90413-2. [DOI] [PubMed] [Google Scholar]
  5. Burton P., Holland I. B. Two pathways of division inhibition in UV-irradiated E. coli. Mol Gen Genet. 1983;190(2):309–314. doi: 10.1007/BF00330656. [DOI] [PubMed] [Google Scholar]
  6. Béjar S., Bouché J. P. Molecular cloning of the region of the terminus of Escherichia coli K-12 DNA replication. J Bacteriol. 1983 Feb;153(2):604–609. doi: 10.1128/jb.153.2.604-609.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Casaregola S., D'Ari R., Huisman O. Role of DNA replication in the induction and turn-off of the SOS response in Escherichia coli. Mol Gen Genet. 1982;185(3):440–444. doi: 10.1007/BF00334136. [DOI] [PubMed] [Google Scholar]
  8. Chen P. L., Carl P. L. Genetic map location of the Escherichia coli dnaG gene. J Bacteriol. 1975 Dec;124(3):1613–1614. doi: 10.1128/jb.124.3.1613-1614.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Clarke L., Carbon J. A colony bank containing synthetic Col El hybrid plasmids representative of the entire E. coli genome. Cell. 1976 Sep;9(1):91–99. doi: 10.1016/0092-8674(76)90055-6. [DOI] [PubMed] [Google Scholar]
  10. D'Ari R., Huisman O. Novel mechanism of cell division inhibition associated with the SOS response in Escherichia coli. J Bacteriol. 1983 Oct;156(1):243–250. doi: 10.1128/jb.156.1.243-250.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Filutowicz M., Jonczyk P. The gyrB gene product functions in both initiation and chain polymerization of Escherichia coli chromosome replication: suppression of the initiation deficiency in gyrB-ts mutants by a class of rpoB mutations. Mol Gen Genet. 1983;191(2):282–287. doi: 10.1007/BF00334827. [DOI] [PubMed] [Google Scholar]
  12. George J., Castellazzi M., Buttin G. Prophage induction and cell division in E. coli. III. Mutations sfiA and sfiB restore division in tif and lon strains and permit the expression of mutator properties of tif. Mol Gen Genet. 1975 Oct 22;140(4):309–332. [PubMed] [Google Scholar]
  13. Hansen F. G., von Meyenburg K. Characterization of the dnaA, gyrB and other genes in the dnaA region of the Escherichia coli chromosome on specialized transducing phages lambda tna. Mol Gen Genet. 1979 Sep;175(2):135–144. doi: 10.1007/BF00425529. [DOI] [PubMed] [Google Scholar]
  14. Helmstetter C. E. DNA synthesis during the division cycle of rapidly growing Escherichia coli B/r. J Mol Biol. 1968 Feb 14;31(3):507–518. doi: 10.1016/0022-2836(68)90424-5. [DOI] [PubMed] [Google Scholar]
  15. Henson J. M., Kuempel P. L. Deletion of the terminus region (340 kilobase pairs of DNA) from the chromosome of Escherichia coli. Proc Natl Acad Sci U S A. 1985 Jun;82(11):3766–3770. doi: 10.1073/pnas.82.11.3766. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Hirota Y., Ryter A., Jacob F. Thermosensitive mutants of E. coli affected in the processes of DNA synthesis and cellular division. Cold Spring Harb Symp Quant Biol. 1968;33:677–693. doi: 10.1101/sqb.1968.033.01.077. [DOI] [PubMed] [Google Scholar]
  17. Hu J. C., Gross C. A. Mutations in the sigma subunit of E. coli RNA polymerase which affect positive control of transcription. Mol Gen Genet. 1985;199(1):7–13. doi: 10.1007/BF00327502. [DOI] [PubMed] [Google Scholar]
  18. Huisman O., D'Ari R. An inducible DNA replication-cell division coupling mechanism in E. coli. Nature. 1981 Apr 30;290(5809):797–799. doi: 10.1038/290797a0. [DOI] [PubMed] [Google Scholar]
  19. Huisman O., D'Ari R. Effect of suppressors of SOS-mediated filamentation on sfiA operon expression in Escherichia coli. J Bacteriol. 1983 Jan;153(1):169–175. doi: 10.1128/jb.153.1.169-175.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Huisman O., D'Ari R., George J. Inducible sfi dependent division inhibition in Escherichia coli. Mol Gen Genet. 1980;177(4):629–636. doi: 10.1007/BF00272673. [DOI] [PubMed] [Google Scholar]
  21. Huisman O., D'Ari R., Gottesman S. Cell-division control in Escherichia coli: specific induction of the SOS function SfiA protein is sufficient to block septation. Proc Natl Acad Sci U S A. 1984 Jul;81(14):4490–4494. doi: 10.1073/pnas.81.14.4490. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Jaffé A., D'Ari R., Norris V. SOS-independent coupling between DNA replication and cell division in Escherichia coli. J Bacteriol. 1986 Jan;165(1):66–71. doi: 10.1128/jb.165.1.66-71.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Jaffé A., D'Ari R. Regulation of chromosome segregation in Escherichia coli. Ann Inst Pasteur Microbiol. 1985 Jan-Feb;136A(1):159–164. doi: 10.1016/s0769-2609(85)80036-3. [DOI] [PubMed] [Google Scholar]
  24. Jones N. C., Donachie W. D. Chromosome replication, transcription and control of cell division in Escherichia coli. Nat New Biol. 1973 May 23;243(125):100–103. [PubMed] [Google Scholar]
  25. Kingsbury D. T., Helinski D. R. DNA polymerase as a requirement for the maintenance of the bacterial plasmid colicinogenic factor E1. Biochem Biophys Res Commun. 1970 Dec 24;41(6):1538–1544. doi: 10.1016/0006-291x(70)90562-0. [DOI] [PubMed] [Google Scholar]
  26. Kourilsky P., Leidner J., Tremblay G. Y. DNA-DNA hybridization on filters at low temperature in the presence of formamide or urea. Biochimie. 1971;53(10):1111–1114. doi: 10.1016/s0300-9084(71)80201-8. [DOI] [PubMed] [Google Scholar]
  27. Kuempel P. L., Duerr S. A., Seeley N. R. Terminus region of the chromosome in Escherichia coli inhibits replication forks. Proc Natl Acad Sci U S A. 1977 Sep;74(9):3927–3931. doi: 10.1073/pnas.74.9.3927. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. LaVerne L. S., Ray D. S. Site-specific integration of an F' lac pro factor in the region of the replication origin (oriC) of E. coli. Mol Gen Genet. 1980;179(2):437–446. doi: 10.1007/BF00425474. [DOI] [PubMed] [Google Scholar]
  29. 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]
  30. Louarn J., Patte J., Louarn J. M. Evidence for a fixed termination site of chromosome replication in Escherichia coli K12. J Mol Biol. 1977 Sep 25;115(3):295–314. doi: 10.1016/0022-2836(77)90156-5. [DOI] [PubMed] [Google Scholar]
  31. Lupski J. R., Godson G. N. The rpsU-dnaG-rpoD macromolecular synthesis operon of E. coli. Cell. 1984 Dec;39(2 Pt 1):251–252. doi: 10.1016/0092-8674(84)90001-1. [DOI] [PubMed] [Google Scholar]
  32. Marsh R. C., Hepburn M. L. Inititation and termination of chromosome replication in Escherichia coli subjected to amino acid starvation. J Bacteriol. 1980 Apr;142(1):236–242. doi: 10.1128/jb.142.1.236-242.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Marsh R. C. Map location of the Escherichia coli origin of replication. Mol Gen Genet. 1978 Nov 9;166(3):299–304. doi: 10.1007/BF00267622. [DOI] [PubMed] [Google Scholar]
  34. Marsh R. C., Worcel A. A DNA fragment containing the origin of replication of the Escherichia coli chromosome. Proc Natl Acad Sci U S A. 1977 Jul;74(7):2720–2724. doi: 10.1073/pnas.74.7.2720. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. McCorkle G. M., Leathers T. D., Umbarger H. E. Physical organization of the ilvEDAC genes of Escherichia coli strain K-12. Proc Natl Acad Sci U S A. 1978 Jan;75(1):89–93. doi: 10.1073/pnas.75.1.89. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Monk M., Gross J. Induction of prophage lambda in a mutant of E. coli K12 defective in initiation of DNA replication at high temperature. Mol Gen Genet. 1971;110(4):299–306. doi: 10.1007/BF00438272. [DOI] [PubMed] [Google Scholar]
  37. Murakami Y., Nagata T., Schwarz W., Wada C., Yura T. Novel dnaG mutation in a dnaP mutant of Escherichia coli. J Bacteriol. 1985 May;162(2):830–832. doi: 10.1128/jb.162.2.830-832.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Nishimura Y., Takeda Y., Nishimura A., Suzuki H., Inouye M., Hirota Y. Synthetic ColE1 plasmids carrying genes for cell division in Escherichia coli. Plasmid. 1977 Nov;1(1):67–77. doi: 10.1016/0147-619x(77)90009-9. [DOI] [PubMed] [Google Scholar]
  39. Orr E., Fairweather N. F., Holland I. B., Pritchard R. H. Isolation and characterisation of a strain carrying a conditional lethal mutation in the cou gene of Escherichia coli K12. Mol Gen Genet. 1979;177(1):103–112. doi: 10.1007/BF00267259. [DOI] [PubMed] [Google Scholar]
  40. Plumbridge J. A., Howe J. G., Springer M., Touati-Schwartz D., Hershey J. W., Grunberg-Manago M. Cloning and mapping of a gene for translational initiation factor IF2 in Escherichia coli. Proc Natl Acad Sci U S A. 1982 Aug;79(16):5033–5037. doi: 10.1073/pnas.79.16.5033. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Sasakawa C., Yoshikawa M. Requirements for suppression of a dnaG mutation by an I-type plasmid. J Bacteriol. 1978 Feb;133(2):485–491. doi: 10.1128/jb.133.2.485-491.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Steck T. R., Drlica K. Bacterial chromosome segregation: evidence for DNA gyrase involvement in decatenation. Cell. 1984 Apr;36(4):1081–1088. doi: 10.1016/0092-8674(84)90058-8. [DOI] [PubMed] [Google Scholar]
  43. Subrahmanyam C. S., Noti J. D., Umbarger H. E. Regulation of ilvEDA expression occurs upstream of ilvG in Escherichia coli: additional evidence for an ilvGEDA operon. J Bacteriol. 1980 Oct;144(1):279–290. doi: 10.1128/jb.144.1.279-290.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Taylor W. E., Straus D. B., Grossman A. D., Burton Z. F., Gross C. A., Burgess R. R. Transcription from a heat-inducible promoter causes heat shock regulation of the sigma subunit of E. coli RNA polymerase. Cell. 1984 Sep;38(2):371–381. doi: 10.1016/0092-8674(84)90492-6. [DOI] [PubMed] [Google Scholar]
  45. Utsumi R., Nakamoto Y., Kawamukai M., Himeno M., Komano T. Involvement of cyclic AMP and its receptor protein in filamentation of an Escherichia coli fic mutant. J Bacteriol. 1982 Aug;151(2):807–812. doi: 10.1128/jb.151.2.807-812.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Uzan M., Favre R., Gallay E., Caro L. Genetical and structural analysis of a group of lambda ilv and lambda rho transducing phages. Mol Gen Genet. 1981;182(3):462–470. doi: 10.1007/BF00293936. [DOI] [PubMed] [Google Scholar]
  47. Valkenburg J. A., Woldringh C. L., Brakenhoff G. J., van der Voort H. T., Nanninga N. Confocal scanning light microscopy of the Escherichia coli nucleoid: comparison with phase-contrast and electron microscope images. J Bacteriol. 1985 Feb;161(2):478–483. doi: 10.1128/jb.161.2.478-483.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Wada C., Yura T. Phenethyl alcohol resistance in Escherichia coli. 3. A temperature-sensitive mutation(dnaP) affecting DNA replication. Genetics. 1974 Jun;77(2):199–220. doi: 10.1093/genetics/77.2.199. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. 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]
  50. Worcel A. Induction of chromosome re-initiations in a thermosensitive DNA mutant of Escherichiacoli. J Mol Biol. 1970 Sep 14;52(2):371–386. doi: 10.1016/0022-2836(70)90037-9. [DOI] [PubMed] [Google Scholar]
  51. de Massy B., Patte J., Louarn J. M., Bouché J. P. oriX: a new replication origin in E. coli. Cell. 1984 Jan;36(1):221–227. doi: 10.1016/0092-8674(84)90092-8. [DOI] [PubMed] [Google Scholar]

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