Skip to main content
PMC home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1993 Jul;175(14):4405–4413. doi: 10.1128/jb.175.14.4405-4413.1993

The pcnB gene of Escherichia coli, which is required for ColE1 copy number maintenance, is dispensable.

M Masters 1, M D Colloms 1, I R Oliver 1, L He 1, E J Macnaughton 1, Y Charters 1
PMCID: PMC204881  PMID: 8331071

Abstract

The pcnB gene product of Escherchia coli is required for copy number maintenance of plasmids related to ColE1 and also for that of the IncFII plasmid R1. Because PcnB is similar to the tRNA-binding protein tRNA nucleotidyltransferase, we have suggested that the protein would be required only for processes in which an RNA is a prominent regulatory component. This appears to be so; strains deleted for pcnB, although defective in ColE1 and R1 plasmid maintenance, maintain the iteron-regulated plasmids F and P1 normally. We also find that strains deleted for pcnB grow normally, demonstrating that PcnB has no essential cellular role under the conditions tested and suggesting that regulation by antisense RNAs similar to RNAI has no critical role in any essential host process. We confirm by immunological tests that PcnB is likely to be the commercially available enzyme poly(A) polymerase.

Full text

PDF
4406

Images in this article

4407Image
on p.4407
4411Image
on p.4411

Selected References

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

  1. Blomberg P., Wagner E. G., Nordström K. Control of replication of plasmid R1: the duplex between the antisense RNA, CopA, and its target, CopT, is processed specifically in vivo and in vitro by RNase III. EMBO J. 1990 Jul;9(7):2331–2340. doi: 10.1002/j.1460-2075.1990.tb07405.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bolivar F. Construction and characterization of new cloning vehicles. III. Derivatives of plasmid pBR322 carrying unique Eco RI sites for selection of Eco RI generated recombinant DNA molecules. Gene. 1978 Oct;4(2):121–136. doi: 10.1016/0378-1119(78)90025-2. [DOI] [PubMed] [Google Scholar]
  3. Cao G. J., Sarkar N. Identification of the gene for an Escherichia coli poly(A) polymerase. Proc Natl Acad Sci U S A. 1992 Nov 1;89(21):10380–10384. doi: 10.1073/pnas.89.21.10380. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Eguchi Y., Itoh T., Tomizawa J. Antisense RNA. Annu Rev Biochem. 1991;60:631–652. doi: 10.1146/annurev.bi.60.070191.003215. [DOI] [PubMed] [Google Scholar]
  5. Green P. J., Pines O., Inouye M. The role of antisense RNA in gene regulation. Annu Rev Biochem. 1986;55:569–597. doi: 10.1146/annurev.bi.55.070186.003033. [DOI] [PubMed] [Google Scholar]
  6. Gross-Bellard M., Oudet P., Chambon P. Isolation of high-molecular-weight DNA from mammalian cells. Eur J Biochem. 1973 Jul 2;36(1):32–38. doi: 10.1111/j.1432-1033.1973.tb02881.x. [DOI] [PubMed] [Google Scholar]
  7. Grunstein M., Hogness D. S. Colony hybridization: a method for the isolation of cloned DNAs that contain a specific gene. Proc Natl Acad Sci U S A. 1975 Oct;72(10):3961–3965. doi: 10.1073/pnas.72.10.3961. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. 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]
  9. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  10. Liu J. D., Parkinson J. S. Genetics and sequence analysis of the pcnB locus, an Escherichia coli gene involved in plasmid copy number control. J Bacteriol. 1989 Mar;171(3):1254–1261. doi: 10.1128/jb.171.3.1254-1261.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Lopilato J., Bortner S., Beckwith J. Mutations in a new chromosomal gene of Escherichia coli K-12, pcnB, reduce plasmid copy number of pBR322 and its derivatives. Mol Gen Genet. 1986 Nov;205(2):285–290. doi: 10.1007/BF00430440. [DOI] [PubMed] [Google Scholar]
  12. March J. B., Colloms M. D., Hart-Davis D., Oliver I. R., Masters M. Cloning and characterization of an Escherichia coli gene, pcnB, affecting plasmid copy number. Mol Microbiol. 1989 Jul;3(7):903–910. doi: 10.1111/j.1365-2958.1989.tb00239.x. [DOI] [PubMed] [Google Scholar]
  13. Masters M., March J. B., Oliver I. R., Collins J. F. A possible role for the pcnB gene product of Escherichia coli in modulating RNA: RNA interactions. Mol Gen Genet. 1990 Jan;220(2):341–344. doi: 10.1007/BF00260507. [DOI] [PubMed] [Google Scholar]
  14. Masters M., Paterson T., Popplewell A. G., Owen-Hughes T., Pringle J. H., Begg K. J. The effect of DnaA protein levels and the rate of initiation at oriC on transcription originating in the ftsQ and ftsA genes: in vivo experiments. Mol Gen Genet. 1989 Apr;216(2-3):475–483. doi: 10.1007/BF00334393. [DOI] [PubMed] [Google Scholar]
  15. McLennan N. F., Girshovich A. S., Lissin N. M., Charters Y., Masters M. The strongly conserved carboxyl-terminus glycine-methionine motif of the Escherichia coli GroEL chaperonin is dispensable. Mol Microbiol. 1993 Jan;7(1):49–58. doi: 10.1111/j.1365-2958.1993.tb01096.x. [DOI] [PubMed] [Google Scholar]
  16. Minden J. S., Marians K. J. Replication of pBR322 DNA in vitro with purified proteins. Requirement for topoisomerase I in the maintenance of template specificity. J Biol Chem. 1985 Aug 5;260(16):9316–9325. [PubMed] [Google Scholar]
  17. Moir P. D., Spiegelberg R., Oliver I. R., Pringle J. H., Masters M. Proteins encoded by the Escherichia coli replication terminus region. J Bacteriol. 1992 Apr;174(7):2102–2110. doi: 10.1128/jb.174.7.2102-2110.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Nordström K. Control of plasmid replication--how do DNA iterons set the replication frequency? Cell. 1990 Dec 21;63(6):1121–1124. doi: 10.1016/0092-8674(90)90405-4. [DOI] [PubMed] [Google Scholar]
  19. Ogura T., Hiraga S. Mini-F plasmid genes that couple host cell division to plasmid proliferation. Proc Natl Acad Sci U S A. 1983 Aug;80(15):4784–4788. doi: 10.1073/pnas.80.15.4784. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Ogura T., Niki H., Kano Y., Imamoto F., Hiraga S. Maintenance of plasmids in HU and IHF mutants of Escherichia coli. Mol Gen Genet. 1990 Jan;220(2):197–203. doi: 10.1007/BF00260482. [DOI] [PubMed] [Google Scholar]
  21. Russell C. B., Thaler D. S., Dahlquist F. W. Chromosomal transformation of Escherichia coli recD strains with linearized plasmids. J Bacteriol. 1989 May;171(5):2609–2613. doi: 10.1128/jb.171.5.2609-2613.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Simons R. W., Kleckner N. Biological regulation by antisense RNA in prokaryotes. Annu Rev Genet. 1988;22:567–600. doi: 10.1146/annurev.ge.22.120188.003031. [DOI] [PubMed] [Google Scholar]
  23. Sippel A. E. Purification and characterization of adenosine triphosphate: ribonucleic acid adenyltransferase from Escherichia coli. Eur J Biochem. 1973 Aug 1;37(1):31–40. doi: 10.1111/j.1432-1033.1973.tb02953.x. [DOI] [PubMed] [Google Scholar]
  24. Smith D. B., Davern K. M., Board P. G., Tiu W. U., Garcia E. G., Mitchell G. F. Mr 26,000 antigen of Schistosoma japonicum recognized by resistant WEHI 129/J mice is a parasite glutathione S-transferase. Proc Natl Acad Sci U S A. 1986 Nov;83(22):8703–8707. doi: 10.1073/pnas.83.22.8703. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Stirling C. J., Colloms S. D., Collins J. F., Szatmari G., Sherratt D. J. xerB, an Escherichia coli gene required for plasmid ColE1 site-specific recombination, is identical to pepA, encoding aminopeptidase A, a protein with substantial similarity to bovine lens leucine aminopeptidase. EMBO J. 1989 May;8(5):1623–1627. doi: 10.1002/j.1460-2075.1989.tb03547.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Stirling C. J., Szatmari G., Stewart G., Smith M. C., Sherratt D. J. The arginine repressor is essential for plasmid-stabilizing site-specific recombination at the ColE1 cer locus. EMBO J. 1988 Dec 20;7(13):4389–4395. doi: 10.1002/j.1460-2075.1988.tb03338.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. 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]
  28. Whittaker P. A., Campbell A. J., Southern E. M., Murray N. E. Enhanced recovery and restriction mapping of DNA fragments cloned in a new lambda vector. Nucleic Acids Res. 1988 Jul 25;16(14B):6725–6736. doi: 10.1093/nar/16.14.6725. [DOI] [PMC free article] [PubMed] [Google Scholar]

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

RESOURCES