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. 1980 Dec;144(3):923–932. doi: 10.1128/jb.144.3.923-932.1980

Location of two relaxation nick sites in R6K and single sites in pSC101 and RSF1010 close to origins of vegetative replication: implication for conjugal transfer of plasmid deoxyribonucleic acid.

A Nordheim, T Hashimoto-Gotoh, K N Timmis
PMCID: PMC294754  PMID: 6254952

Abstract

A nick-labeling method has been used to localize the relaxation complex nick sites in three plasmids (pSC101, RSF1010, and R6K) that differ markedly in their host range, deoxyribonucleic acid replication, and conjugal transfer properties. Single specific relaxation sites were located in pSC101 and RSF1010, but surprisingly two distinct sites could be identified in the bi-origin plasmid R6K. In all cases, relaxation nick sites, which are thought to be origins of plasmid conjugal transfer, were shown to be located near origins of vegetative replication. This result suggests a functional interaction between these two types of deoxyribonucleic acid loci, and we speculate here that application events initiated at origins of replication may constitute an integral part of the process of conjugal transfer of small plasmids among bacteria. Consistent with this proposal is the finding that inhibition of vegetative replication of the pSC101 and ColE1 plasmids results in a severe inhibition of their conjugal transfer ability.

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Selected References

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

  1. Achtman M., Kusećek B., Timmis K. N. Tra cistrons and proteins encoded by the Escherichia coli antibiotic resistance plasmid R6-5. Mol Gen Genet. 1978 Jul 11;163(2):169–179. doi: 10.1007/BF00267407. [DOI] [PubMed] [Google Scholar]
  2. Andrés I., Slocombe P. M., Cabello F., Timmis J. K., Lurz R., Burkardt H. J., Timmis K. N. Plasmid replication functions. II. Cloning analysis of the repA replication region of antibiotic resistance plasmid R6-5. Mol Gen Genet. 1979 Jan 5;168(1):1–25. doi: 10.1007/BF00267929. [DOI] [PubMed] [Google Scholar]
  3. Bagdasarian M. M., Izakowska M., Bagdasarian M. Suppression of the DnaA phenotype by mutations in the rpoB cistron of ribonucleic acid polymerase in Salmonella typhimurium and Escherichia coli. J Bacteriol. 1977 May;130(2):577–582. doi: 10.1128/jb.130.2.577-582.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bastia D. Determination of restriction sites and the nucleotide sequence surrounding the relaxation site of ColE1. J Mol Biol. 1978 Oct 5;124(4):601–639. doi: 10.1016/0022-2836(78)90174-2. [DOI] [PubMed] [Google Scholar]
  5. Boulnois G. J., Wilkins B. M. A novel priming system for conjugal synthesis of an IncI alpha plasmid in recipients. Mol Gen Genet. 1979 Oct 1;175(3):275–279. doi: 10.1007/BF00397227. [DOI] [PubMed] [Google Scholar]
  6. Cabello F., Timmis K., Cohen S. N. Replication control in a composite plasmid constructed by in vitro linkage of two distinct replicons. Nature. 1976 Jan 29;259(5541):285–290. doi: 10.1038/259285a0. [DOI] [PubMed] [Google Scholar]
  7. Clewell D. B., Helinski D. R. Supercoiled circular DNA-protein complex in Escherichia coli: purification and induced conversion to an opern circular DNA form. Proc Natl Acad Sci U S A. 1969 Apr;62(4):1159–1166. doi: 10.1073/pnas.62.4.1159. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Cohen S. N., Chang A. C. Revised interpretation of the origin of the pSC101 plasmid. J Bacteriol. 1977 Nov;132(2):734–737. doi: 10.1128/jb.132.2.734-737.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Crisona N. J., Clark A. J. Increase in conjugational transmission frequency of nonconjugative plasmids. Science. 1977 Apr 8;196(4286):186–187. doi: 10.1126/science.322280. [DOI] [PubMed] [Google Scholar]
  10. Crosa J. H., Luttropp L. K., Falkow S. Mode of replication of the conjugative R-plasmid RSF1040 in Escherichia coli. J Bacteriol. 1976 Apr;126(1):454–466. doi: 10.1128/jb.126.1.454-466.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Crosa J. H., Luttropp L. K., Falkow S. Molecular cloning of replication and incompatibility regions from the R-plasmid R6K. J Mol Biol. 1978 Sep 25;124(3):443–468. doi: 10.1016/0022-2836(78)90181-x. [DOI] [PubMed] [Google Scholar]
  12. Dougan G., Sherratt D. The transposon Tn1 as a probe for studying ColE1 structure and function. Mol Gen Genet. 1977 Mar 7;151(2):151–160. doi: 10.1007/BF00338689. [DOI] [PubMed] [Google Scholar]
  13. Eisenberg S., Griffith J., Kornberg A. phiX174 cistron A protein is a multifunctional enzyme in DNA replication. Proc Natl Acad Sci U S A. 1977 Aug;74(8):3198–3202. doi: 10.1073/pnas.74.8.3198. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Guiney D. G., Helinski D. R. Relaxation complexes of poasmid DNA and protein. III. Association of protein with the 5' terminus of the broken DNA strand in the relaxed complex of plasmid ColE1. J Biol Chem. 1975 Nov 25;250(22):8796–8803. [PubMed] [Google Scholar]
  15. Guiney D. G., Helinski D. R. The DNA-protein relaxation complex of the plasmid RK2: location of the site-specific nick in the region of the proposed origin of transfer. Mol Gen Genet. 1979 Oct 3;176(2):183–189. doi: 10.1007/BF00273212. [DOI] [PubMed] [Google Scholar]
  16. Hashimoto-Gotoh T., Inselburg J. ColE1 plasmid incompatibility: localization and analysis of mutations affecting incompatibility. J Bacteriol. 1979 Aug;139(2):608–619. doi: 10.1128/jb.139.2.608-619.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Hashimoto-Gotoh T., Sekiguchi M. Mutations of temperature sensitivity in R plasmid pSC101. J Bacteriol. 1977 Aug;131(2):405–412. doi: 10.1128/jb.131.2.405-412.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Hasunuma K., Sekiguchi M. Replication of plasmid pSC101 in Escherichia coli K12: requirement for dnaA function. Mol Gen Genet. 1977 Sep 9;154(3):225–230. doi: 10.1007/BF00571277. [DOI] [PubMed] [Google Scholar]
  19. Helinski D. R. Plasmid DNA replication. Fed Proc. 1976 Jul;35(9):2026–2030. [PubMed] [Google Scholar]
  20. Inselburg J. Studies of colicin E1 plasmid functions by analysis of deletions and TnA insertions of the plasmid. J Bacteriol. 1977 Oct;132(1):332–340. doi: 10.1128/jb.132.1.332-340.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Kingsbury D. T., Helinski D. R. Temperature-sensitive mutants for the replication of plasmids in Escherichia coli: requirement for deoxyribonucleic acid polymerase I in the replication of the plasmid ColE 1 . J Bacteriol. 1973 Jun;114(3):1116–1124. doi: 10.1128/jb.114.3.1116-1124.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Kingsman A., Willetts N. The requirements for conjugal DNA synthesis in the donor strain during flac transfer. J Mol Biol. 1978 Jul 5;122(3):287–300. doi: 10.1016/0022-2836(78)90191-2. [DOI] [PubMed] [Google Scholar]
  23. Kolter R., Helinski D. R. Construction of plasmid R6K derivatives in vitro: characterization of the R6K replication region. Plasmid. 1978 Sep;1(4):571–580. doi: 10.1016/0147-619x(78)90014-8. [DOI] [PubMed] [Google Scholar]
  24. Kupersztoch-Portnoy Y. M., Lovett M. A., Helinski D. R. Strand and site specificity of the relaxation event for the relaxation complex of the antibiotic resistance plasmid R6K. Biochemistry. 1974 Dec 31;13(27):5484–5490. doi: 10.1021/bi00724a005. [DOI] [PubMed] [Google Scholar]
  25. Lanka E., Scherzinger E., Günther E., Schuster H. A DNA primase specified by I-like plasmids. Proc Natl Acad Sci U S A. 1979 Aug;76(8):3632–3636. doi: 10.1073/pnas.76.8.3632. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Lovett M. A., Guiney D. G., Helinski D. R. Relaxation complexes of plasmids ColE1 and ColE2: unique site of the nick in the open circular DNA of the relaxed complexes. Proc Natl Acad Sci U S A. 1974 Oct;71(10):3854–3857. doi: 10.1073/pnas.71.10.3854. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Lovett M. A., Helinski D. R. Relaxation complexes of plasmid DNA and protein. II. Characterization of the proteins associated with the unrelaxed and relaxed complexes of plasmid ColE1. J Biol Chem. 1975 Nov 25;250(22):8790–8795. [PubMed] [Google Scholar]
  28. Lovett M. A., Sparks R. B., Helinski D. R. Bidirectional replication of plasmid R6K DNA in Escherichia coli; correspondence between origin of replication and position of single-strand break in relaxed complex. Proc Natl Acad Sci U S A. 1975 Aug;72(8):2905–2909. doi: 10.1073/pnas.72.8.2905. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Meacock P. A., Cohen S. N. Genetic analysis of the inter-relationship between plasmid replication and incompatibility. Mol Gen Genet. 1979 Jul 13;174(2):135–147. doi: 10.1007/BF00268351. [DOI] [PubMed] [Google Scholar]
  30. Ohki M., Tomizawa J. Asymmetric transfer of DNA strands in bacterial conjugation. Cold Spring Harb Symp Quant Biol. 1968;33:651–658. doi: 10.1101/sqb.1968.033.01.074. [DOI] [PubMed] [Google Scholar]
  31. Rupp W. D., Ihler G. Strand selection during bacterial mating. Cold Spring Harb Symp Quant Biol. 1968;33:647–650. doi: 10.1101/sqb.1968.033.01.073. [DOI] [PubMed] [Google Scholar]
  32. Staudenbauer W. L. REPLICAtion of small plasmids in extracts of Escherichia coli: requirement for both DNA polymerases I and II. Mol Gen Genet. 1976 Dec 8;149(2):151–158. doi: 10.1007/BF00332883. [DOI] [PubMed] [Google Scholar]
  33. Synenki R. M., Nordheim A., Timmis K. N. Plasmid replication functions. III. Origin and direction of replication of a "mini" plasmid derived from R6-5. Mol Gen Genet. 1979 Jan 5;168(1):27–36. doi: 10.1007/BF00267930. [DOI] [PubMed] [Google Scholar]
  34. Tait R. C., Boyer H. W. On the nature of tetracycline resistance controlled by the plasmid pSC101. Cell. 1978 Jan;13(1):73–81. doi: 10.1016/0092-8674(78)90139-3. [DOI] [PubMed] [Google Scholar]
  35. Timmis K. N., Cabello F., Cohen S. N. Cloning and characterization of EcoRI and HindIII restriction endonuclease-generated fragments of antibiotic resistance plasmids R6-5 and R6. Mol Gen Genet. 1978 Jun 14;162(2):121–137. doi: 10.1007/BF00267869. [DOI] [PubMed] [Google Scholar]
  36. Timmis K., Cabello F., Cohen S. N. Cloning, isolation, and characterization of replication regions of complex plasmid genomes. Proc Natl Acad Sci U S A. 1975 Jun;72(6):2242–2246. doi: 10.1073/pnas.72.6.2242. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Warren G. J., Twigg A. J., Sherratt D. J. ColE1 plasmid mobility and relaxation complex. Nature. 1978 Jul 20;274(5668):259–261. doi: 10.1038/274259a0. [DOI] [PubMed] [Google Scholar]
  38. de Graaff J., Crosa J. H., Heffron F., Falkow S. Replication of the nonconjugative plasmid RSF1010 in Escherichia coli K-12. J Bacteriol. 1978 Jun;134(3):1117–1122. doi: 10.1128/jb.134.3.1117-1122.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]

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