Skip to main content
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1982 Nov;152(2):829–839. doi: 10.1128/jb.152.2.829-839.1982

The incompatibility product of IncFII R plasmid NR1 controls gene expression in the plasmid replication region.

A M Easton, R H Rownd
PMCID: PMC221537  PMID: 6290455

Abstract

The incompatibility properties of IncFII R plasmid NR1 were compared with those of two of its copy number mutants, pRR12 and pRR21. pRR12 produced an altered incompatibility product and also had an altered incompatibility target site. The target site appeared to be located within the incompatibility gene, which is located more than 1,200 base pairs from the plasmid origin of replication. The incompatibility properties of pRR21 were indistinguishable from those of NR1. Lambda phages have been constructed which contain the incompatibility region of NR1 or of one of its copy mutants fused to the lacZ gene. In lysogens constructed with these phages, beta-galactosidase was produced under the control of a promoter located within the plasmid incompatibility region. Lysogens containing prophages with the incompatibility regions from pRR12 and pRR21 produced higher levels of beta-galactosidase than did lysogens containing prophages with the incompatibility region from the wild-type NR1. The introduction into these inc-lac lysogens of pBR322 plasmids carrying the incompatibility regions of the wild-type or mutant plasmids resulted in decreased levels of beta-galactosidase production. For a given lysogen, the decrease was greater when the pBR322 derivative expressed a stronger incompatibility toward the plasmid from which the fragment in the prophage was derived. This suggested that the incompatibility product acts on its target to repress gene expression in the plasmid replication region.

Full text

PDF
829

Images in this article

Selected References

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

  1. Alton N. K., Vapnek D. Nucleotide sequence analysis of the chloramphenicol resistance transposon Tn9. Nature. 1979 Dec 20;282(5741):864–869. doi: 10.1038/282864a0. [DOI] [PubMed] [Google Scholar]
  2. Barton C. R., Warren R. L., Jezo P., Easton A. M., Rownd R. H. Sa/I restriction endonuclease maps of FII incompatibility group R plasmids. Plasmid. 1979 Jan;2(1):150–154. doi: 10.1016/0147-619x(79)90013-1. [DOI] [PubMed] [Google Scholar]
  3. Blattner F. R., Blechl A. E., Denniston-Thompson K., Faber H. E., Richards J. E., Slightom J. L., Tucker P. W., Smithies O. Cloning human fetal gamma globin and mouse alpha-type globin DNA: preparation and screening of shotgun collections. Science. 1978 Dec 22;202(4374):1279–1284. doi: 10.1126/science.725603. [DOI] [PubMed] [Google Scholar]
  4. Bolivar F., Rodriguez R. L., Greene P. J., Betlach M. C., Heyneker H. L., Boyer H. W., Crosa J. H., Falkow S. Construction and characterization of new cloning vehicles. II. A multipurpose cloning system. Gene. 1977;2(2):95–113. [PubMed] [Google Scholar]
  5. 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]
  6. Conrad S. E., Campbell J. L. Role of plasmid-coded RNA and ribonuclease III in plasmid DNA replication. Cell. 1979 Sep;18(1):61–71. doi: 10.1016/0092-8674(79)90354-4. [DOI] [PubMed] [Google Scholar]
  7. Danbara H., Brady G., Timmis J. K., Timmis K. N. Regulation of DNA replication: "target" determinant of the replication control elements of plasmid R6-5 lies within a control element gene. Proc Natl Acad Sci U S A. 1981 Aug;78(8):4699–4703. doi: 10.1073/pnas.78.8.4699. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Guarente L., Lauer G., Roberts T. M., Ptashne M. Improved methods for maximizing expression of a cloned gene: a bacterium that synthesizes rabbit beta-globin. Cell. 1980 Jun;20(2):543–553. doi: 10.1016/0092-8674(80)90640-6. [DOI] [PubMed] [Google Scholar]
  9. Kollek R., Oertel W., Goebel W. Isolation and characterization of the minimal fragment required for autonomous replication ("basic replicon") of a copy mutant (pKN102) of the antibiotic resistance factor R1. Mol Gen Genet. 1978 Jun 1;162(1):51–57. doi: 10.1007/BF00333850. [DOI] [PubMed] [Google Scholar]
  10. Kolter R. Replication properties of plasmic R6K. Plasmid. 1981 Jan;5(1):2–9. doi: 10.1016/0147-619x(81)90073-1. [DOI] [PubMed] [Google Scholar]
  11. LENNOX E. S. Transduction of linked genetic characters of the host by bacteriophage P1. Virology. 1955 Jul;1(2):190–206. doi: 10.1016/0042-6822(55)90016-7. [DOI] [PubMed] [Google Scholar]
  12. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  13. Light J., Molin S. Replication control functions of plasmid R1 act as inhibitors of expression of a gene required for replication. Mol Gen Genet. 1981;184(1):56–61. doi: 10.1007/BF00271195. [DOI] [PubMed] [Google Scholar]
  14. Mickel S., Bauer W. Isolation, by tetracycline selection, of small plasmids derived from R-factor R12 in Escherichia coli K-12. J Bacteriol. 1976 Jul;127(1):644–655. doi: 10.1128/jb.127.1.644-655.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Miki T., Easton A. M., Rownd R. H. Cloning of replication, incompatibility, and stability functions of R plasmid NR1. J Bacteriol. 1980 Jan;141(1):87–99. doi: 10.1128/jb.141.1.87-99.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Miki T., Easton A. M., Rownd R. H. Mapping of the resistance genes of the R plasmid NR1. Mol Gen Genet. 1978 Jan 17;158(3):217–224. doi: 10.1007/BF00267192. [DOI] [PubMed] [Google Scholar]
  17. Molin S., Nordström K. Control of plasmid R1 replication: functions involved in replication, copy number control, incompatibility, and switch-off of replication. J Bacteriol. 1980 Jan;141(1):111–120. doi: 10.1128/jb.141.1.111-120.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Morris C. F., Hashimoto H., Mickel S., Rownd R. Round of replication mutant of a drug resistance factor. J Bacteriol. 1974 Jun;118(3):855–866. doi: 10.1128/jb.118.3.855-866.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Muesing M., Tamm J., Shepard H. M., Polisky B. A single base-pair alteration is responsible for the DNA overproduction phenotype of a plasmid copy-number mutant. Cell. 1981 Apr;24(1):235–242. doi: 10.1016/0092-8674(81)90519-5. [DOI] [PubMed] [Google Scholar]
  20. Novick R. P., Hoppensteadt F. C. On plasmid incompatibility. Plasmid. 1978 Sep;1(4):421–434. doi: 10.1016/0147-619x(78)90001-x. [DOI] [PubMed] [Google Scholar]
  21. Ohtsubo E., Feingold J., Ohtsubo H., Mickel S., Bauer W. Unidirectional replication in Escherichia coli of three small plasmids derived from R factor R12. Plasmid. 1977 Nov;1(1):8–18. doi: 10.1016/0147-619x(77)90004-x. [DOI] [PubMed] [Google Scholar]
  22. Rosen J., Ryder T., Inokuchi H., Ohtsubo H., Ohtsubo E. Genes and sites involved in replication and incompatibility of an R100 plasmid derivative based on nucleotide sequence analysis. Mol Gen Genet. 1980;179(3):527–537. doi: 10.1007/BF00271742. [DOI] [PubMed] [Google Scholar]
  23. Rosen J., Ryder T., Ohtsubo H., Ohtsubo E. Role of RNA transcripts in replication incompatibility and copy number control in antibiotic resistance plasmid derivatives. Nature. 1981 Apr 30;290(5809):794–797. doi: 10.1038/290794a0. [DOI] [PubMed] [Google Scholar]
  24. Sharp P. A., Cohen S. N., Davidson N. Electron microscope heteroduplex studies of sequence relations among plasmids of Escherichia coli. II. Structure of drug resistance (R) factors and F factors. J Mol Biol. 1973 Apr 5;75(2):235–255. doi: 10.1016/0022-2836(73)90018-1. [DOI] [PubMed] [Google Scholar]
  25. Shaw W. V. Chloramphenicol acetyltransferase from chloramphenicol-resistant bacteria. Methods Enzymol. 1975;43:737–755. doi: 10.1016/0076-6879(75)43141-x. [DOI] [PubMed] [Google Scholar]
  26. Shepard H. M., Gelfand D. H., Polisky B. Analysis of a recessive plasmid copy number mutant: evidence for negative control of Col E1 replication. Cell. 1979 Oct;18(2):267–275. doi: 10.1016/0092-8674(79)90046-1. [DOI] [PubMed] [Google Scholar]
  27. Stougaard P., Molin S., Nordström K. RNAs involved in copy-number control and incompatibility of plasmid R1. Proc Natl Acad Sci U S A. 1981 Oct;78(10):6008–6012. doi: 10.1073/pnas.78.10.6008. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. 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]
  29. Tanak N., Cramer J. H., Rownd R. H. EcoRI restriction endonuclease map of the composite R plasmid NR1. J Bacteriol. 1976 Jul;127(1):619–636. doi: 10.1128/jb.127.1.619-636.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Taylor D. P., Cohen S. N. Structural and functional analysis of cloned DNA segments containing the replication and incompatibility regions of a miniplasmid derived from a copy number mutant of NR1. J Bacteriol. 1979 Jan;137(1):92–104. doi: 10.1128/jb.137.1.92-104.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Taylor D. P., Greenberg J., Rownd R. H. Generation of miniplasmids from copy number mutants of the R plasmid NR1. J Bacteriol. 1977 Dec;132(3):986–995. doi: 10.1128/jb.132.3.986-995.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Timmis K. N., Andrés I., Slocombe P. M. Plasmid incompatibility: cloning analysis of an incFII determinant of R6-5. Nature. 1978 May 4;273(5657):27–32. doi: 10.1038/273027a0. [DOI] [PubMed] [Google Scholar]
  33. Tomizawa J., Itoh T., Selzer G., Som T. Inhibition of ColE1 RNA primer formation by a plasmid-specified small RNA. Proc Natl Acad Sci U S A. 1981 Mar;78(3):1421–1425. doi: 10.1073/pnas.78.3.1421. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Uhlin B. E., Nordström K. Plasmid incompatibility and control of replication: copy mutants of the R-factor R1 in Escherichia coli K-12. J Bacteriol. 1975 Nov;124(2):641–649. doi: 10.1128/jb.124.2.641-649.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Womble D. D., Taylor D. P., Rownd R. H. Method for obtaining more-accurate covalently closed circular plasmid-to-chromosome ratios from bacterial lysates by dye-buoyant density centrifugation. J Bacteriol. 1977 Apr;130(1):148–153. doi: 10.1128/jb.130.1.148-153.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Journal of Bacteriology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES