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. 1993 Dec 25;21(25):5972–5977. doi: 10.1093/nar/21.25.5972

Control of ColE2 DNA replication: in vitro binding of the antisense RNA to the Rep mRNA.

T Sugiyama 1, T Itoh 1
PMCID: PMC310483  PMID: 7507236

Abstract

Expression of the Rep protein of colicin E2 plasmid is negatively controlled at a post-transcriptional step by a plasmid-coded RNA (RNAI). RNAI is complementary to the 5' nontranslated region of the Rep mRNA and can be folded into two stem-loop structures. Several cop mutations that increased the copy number of the plasmid have been mapped in the RNAI coding region. Here we demonstrated that RNAI and the Rep mRNA rapidly form a stable complex under near physiological conditions. The cop mutations drastically reduced the binding rate. The reduction correlated to the in vivo phenotypes of the cop mutant plasmids, indicating that binding of RNAI to the Rep mRNA is involved in the regulation of the Rep protein expression. Binding properties of RNAI and the Rep mRNA with the cop mutations and those with various extents of deletion suggested that the initial interaction of the two RNAs occurs between the loop portions of the larger of the two stem-loop structures. RNAI does not completely sequester the putative S/D region, even when the two RNAs form a complete duplex. Possible mechanisms of the inhibition of the rep gene expression by binding of RNAI to the Rep mRNA under such a situation were discussed.

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Selected References

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

  1. Asano K., Moriwaki H., Mizobuchi K. An induced mRNA secondary structure enhances repZ translation in plasmid ColIb-P9. J Biol Chem. 1991 Dec 25;266(36):24549–24556. [PubMed] [Google Scholar]
  2. Burgess R. R., Jendrisak J. J. A procedure for the rapid, large-scall purification of Escherichia coli DNA-dependent RNA polymerase involving Polymin P precipitation and DNA-cellulose chromatography. Biochemistry. 1975 Oct 21;14(21):4634–4638. doi: 10.1021/bi00692a011. [DOI] [PubMed] [Google Scholar]
  3. 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]
  4. Gold L. Posttranscriptional regulatory mechanisms in Escherichia coli. Annu Rev Biochem. 1988;57:199–233. doi: 10.1146/annurev.bi.57.070188.001215. [DOI] [PubMed] [Google Scholar]
  5. Gonzalez N., Wiggs J., Chamberlin M. J. A simple procedure for resolution of Escherichia coli RNA polymerase holoenzyme from core polymerase. Arch Biochem Biophys. 1977 Aug;182(2):404–408. doi: 10.1016/0003-9861(77)90521-5. [DOI] [PubMed] [Google Scholar]
  6. Hjalt T., Wagner E. G. The effect of loop size in antisense and target RNAs on the efficiency of antisense RNA control. Nucleic Acids Res. 1992 Dec 25;20(24):6723–6732. doi: 10.1093/nar/20.24.6723. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Horii T., Itoh T. Replication of ColE2 and ColE3 plasmids: the regions sufficient for autonomous replication. Mol Gen Genet. 1988 May;212(2):225–231. doi: 10.1007/BF00334689. [DOI] [PubMed] [Google Scholar]
  8. Itoh T., Horii T. Replication of ColE2 and ColE3 plasmids: in vitro replication dependent on plasmid-coded proteins. Mol Gen Genet. 1989 Oct;219(1-2):249–255. doi: 10.1007/BF00261184. [DOI] [PubMed] [Google Scholar]
  9. Kido M., Yasueda H., Itoh T. Identification of a plasmid-coded protein required for initiation of ColE2 DNA replication. Nucleic Acids Res. 1991 Jun 11;19(11):2875–2880. doi: 10.1093/nar/19.11.2875. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Kittle J. D., Simons R. W., Lee J., Kleckner N. Insertion sequence IS10 anti-sense pairing initiates by an interaction between the 5' end of the target RNA and a loop in the anti-sense RNA. J Mol Biol. 1989 Dec 5;210(3):561–572. doi: 10.1016/0022-2836(89)90132-0. [DOI] [PubMed] [Google Scholar]
  11. Krinke L., Wulff D. L. OOP RNA, produced from multicopy plasmids, inhibits lambda cII gene expression through an RNase III-dependent mechanism. Genes Dev. 1987 Nov;1(9):1005–1013. doi: 10.1101/gad.1.9.1005. [DOI] [PubMed] [Google Scholar]
  12. Ma C., Simons R. W. The IS10 antisense RNA blocks ribosome binding at the transposase translation initiation site. EMBO J. 1990 Apr;9(4):1267–1274. doi: 10.1002/j.1460-2075.1990.tb08235.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. McCarthy J. E., Gualerzi C. Translational control of prokaryotic gene expression. Trends Genet. 1990 Mar;6(3):78–85. doi: 10.1016/0168-9525(90)90098-q. [DOI] [PubMed] [Google Scholar]
  14. Novick R. P., Iordanescu S., Projan S. J., Kornblum J., Edelman I. pT181 plasmid replication is regulated by a countertranscript-driven transcriptional attenuator. Cell. 1989 Oct 20;59(2):395–404. doi: 10.1016/0092-8674(89)90300-0. [DOI] [PubMed] [Google Scholar]
  15. Persson C., Wagner E. G., Nordström K. Control of replication of plasmid R1: kinetics of in vitro interaction between the antisense RNA, CopA, and its target, CopT. EMBO J. 1988 Oct;7(10):3279–3288. doi: 10.1002/j.1460-2075.1988.tb03195.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Persson C., Wagner E. G., Nordström K. Control of replication of plasmid R1: structures and sequences of the antisense RNA, CopA, required for its binding to the target RNA, CopT. EMBO J. 1990 Nov;9(11):3767–3775. doi: 10.1002/j.1460-2075.1990.tb07590.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. 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]
  18. Tajima Y., Horii T., Itoh T. Replication of ColE2 and ColE3 plasmids: two ColE2 incompatibility functions. Mol Gen Genet. 1988 Nov;214(3):451–455. doi: 10.1007/BF00330479. [DOI] [PubMed] [Google Scholar]
  19. Tomizawa J. Control of ColE1 plasmid replication: binding of RNA I to RNA II and inhibition of primer formation. Cell. 1986 Oct 10;47(1):89–97. doi: 10.1016/0092-8674(86)90369-7. [DOI] [PubMed] [Google Scholar]
  20. Tomizawa J. Control of ColE1 plasmid replication: the process of binding of RNA I to the primer transcript. Cell. 1984 Oct;38(3):861–870. doi: 10.1016/0092-8674(84)90281-2. [DOI] [PubMed] [Google Scholar]
  21. Tomizawa J., Itoh T. Plasmid ColE1 incompatibility determined by interaction of RNA I with primer transcript. Proc Natl Acad Sci U S A. 1981 Oct;78(10):6096–6100. doi: 10.1073/pnas.78.10.6096. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Tomizawa J., Som T. Control of ColE1 plasmid replication: enhancement of binding of RNA I to the primer transcript by the Rom protein. Cell. 1984 Oct;38(3):871–878. doi: 10.1016/0092-8674(84)90282-4. [DOI] [PubMed] [Google Scholar]
  23. Yasueda H., Horii T., Itoh T. Structural and functional organization of ColE2 and ColE3 replicons. Mol Gen Genet. 1989 Jan;215(2):209–216. doi: 10.1007/BF00339719. [DOI] [PubMed] [Google Scholar]

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