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. 1987 Apr;6(4):1115–1119. doi: 10.1002/j.1460-2075.1987.tb04866.x

Transcriptional termination at a fully rho-independent site in Escherichia coli is prevented by uninterrupted translation of the nascent RNA.

J J Wright, R S Hayward
PMCID: PMC553510  PMID: 3036492

Abstract

We have examined the possibility that translation reading through a fully rho-independent transcriptional terminator in Escherichia coli might prevent termination, as already established for rho-dependent terminators. Plasmids were constructed with and without interposition of the rho-independent coliphage T7 'early' terminator between a promoter and galK. Our constructions ensured either that there was no upstream translation, or that translation (initiated at the galE ribosome binding site) stopped upstream of, or at the normal position (the T7 gene 1.3 stop codon) with respect to, the transcriptional terminator; or else downstream of both this stop codon and the terminator. Our galactokinase enzyme and mRNA measurements on strains harbouring these plasmids indicate that 'readthrough translation' eliminates transcriptional termination at the T7 site. This effect is suppressed if the rate of ribosome movement is reduced with fusidic acid.

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Selected References

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

  1. Adams C. W., Hatfield G. W. Effects of promoter strengths and growth conditions on copy number of transcription-fusion vectors. J Biol Chem. 1984 Jun 25;259(12):7399–7403. [PubMed] [Google Scholar]
  2. Adhya S., Gottesman M. Control of transcription termination. Annu Rev Biochem. 1978;47:967–996. doi: 10.1146/annurev.bi.47.070178.004535. [DOI] [PubMed] [Google Scholar]
  3. Bedwell D., Davis G., Gosink M., Post L., Nomura M., Kestler H., Zengel J. M., Lindahl L. Nucleotide sequence of the alpha ribosomal protein operon of Escherichia coli. Nucleic Acids Res. 1985 Jun 11;13(11):3891–3903. doi: 10.1093/nar/13.11.3891. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bennett P. M., Maaloe O. The effects of fusidic acid on growth, ribosome synthesis and RNA metabolism in Escherichia coli. J Mol Biol. 1974 Dec 15;90(3):541–561. doi: 10.1016/0022-2836(74)90234-4. [DOI] [PubMed] [Google Scholar]
  5. Biggin M. D., Gibson T. J., Hong G. F. Buffer gradient gels and 35S label as an aid to rapid DNA sequence determination. Proc Natl Acad Sci U S A. 1983 Jul;80(13):3963–3965. doi: 10.1073/pnas.80.13.3963. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Bonekamp F., Clemmesen K., Karlström O., Jensen K. F. Mechanism of UTP-modulated attenuation at the pyrE gene of Escherichia coli: an example of operon polarity control through the coupling of translation to transcription. EMBO J. 1984 Dec 1;3(12):2857–2861. doi: 10.1002/j.1460-2075.1984.tb02220.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Brendel V., Hamm G. H., Trifonov E. N. Terminators of transcription with RNA polymerase from Escherichia coli: what they look like and how to find them. J Biomol Struct Dyn. 1986 Feb;3(4):705–723. doi: 10.1080/07391102.1986.10508457. [DOI] [PubMed] [Google Scholar]
  8. Briat J. F., Chamberlin M. J. Identification and characterization of a new transcriptional termination factor from Escherichia coli. Proc Natl Acad Sci U S A. 1984 Dec;81(23):7373–7377. doi: 10.1073/pnas.81.23.7373. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Dunn J. J., Studier F. W. Complete nucleotide sequence of bacteriophage T7 DNA and the locations of T7 genetic elements. J Mol Biol. 1983 Jun 5;166(4):477–535. doi: 10.1016/s0022-2836(83)80282-4. [DOI] [PubMed] [Google Scholar]
  10. Feinberg A. P., Vogelstein B. "A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity". Addendum. Anal Biochem. 1984 Feb;137(1):266–267. doi: 10.1016/0003-2697(84)90381-6. [DOI] [PubMed] [Google Scholar]
  11. Garner I., Cromie K. D., Marson E. A., Hayward R. S. Transcription termination regions of coliphage T7 DNA: the effects of nusA1. Mol Gen Genet. 1985;200(2):295–301. doi: 10.1007/BF00425439. [DOI] [PubMed] [Google Scholar]
  12. Jenkins A. J., March J. B., Oliver I. R., Masters M. A DNA fragment containing the groE genes can suppress mutations in the Escherichia coli dnaA gene. Mol Gen Genet. 1986 Mar;202(3):446–454. doi: 10.1007/BF00333275. [DOI] [PubMed] [Google Scholar]
  13. Johnston H. M., Roth J. R. DNA sequence changes of mutations altering attenuation control of the histidine operon of Salmonella typhimurium. J Mol Biol. 1981 Feb 5;145(4):735–756. doi: 10.1016/0022-2836(81)90312-0. [DOI] [PubMed] [Google Scholar]
  14. Kassavetis G. A., Chamberlin M. J. Pausing and termination of transcription within the early region of bacteriophage T7 DNA in vitro. J Biol Chem. 1981 Mar 25;256(6):2777–2786. [PubMed] [Google Scholar]
  15. Kiefer M., Neff N., Chamberlin M. J. Transcriptional termination at the end of the early region of bacteriophages T3 and T7 is not affected by polarity suppressors. J Virol. 1977 May;22(2):548–552. doi: 10.1128/jvi.22.2.548-552.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Kieny M. P., Lathe R., Lecocq J. P. New versatile cloning and sequencing vectors based on bacteriophage M13. Gene. 1983 Dec;26(1):91–99. doi: 10.1016/0378-1119(83)90039-2. [DOI] [PubMed] [Google Scholar]
  17. Kolter R., Yanofsky C. Attenuation in amino acid biosynthetic operons. Annu Rev Genet. 1982;16:113–134. doi: 10.1146/annurev.ge.16.120182.000553. [DOI] [PubMed] [Google Scholar]
  18. Landick R., Carey J., Yanofsky C. Translation activates the paused transcription complex and restores transcription of the trp operon leader region. Proc Natl Acad Sci U S A. 1985 Jul;82(14):4663–4667. doi: 10.1073/pnas.82.14.4663. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. McKenney K., Shimatake H., Court D., Schmeissner U., Brady C., Rosenberg M. A system to study promoter and terminator signals recognized by Escherichia coli RNA polymerase. Gene Amplif Anal. 1981;2:383–415. [PubMed] [Google Scholar]
  20. Messing J., Vieira J. A new pair of M13 vectors for selecting either DNA strand of double-digest restriction fragments. Gene. 1982 Oct;19(3):269–276. doi: 10.1016/0378-1119(82)90016-6. [DOI] [PubMed] [Google Scholar]
  21. Morgan W. D., Bear D. G., Litchman B. L., von Hippel P. H. RNA sequence and secondary structure requirements for rho-dependent transcription termination. Nucleic Acids Res. 1985 May 24;13(10):3739–3754. doi: 10.1093/nar/13.10.3739. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Newman A. J., Ma J. C., Howe K. M., Garner I., Hayward R. S. Evidence that rifampicin can stimulate readthrough of transcriptional terminators in Escherichia coli, including the attenuator of the rpoBC operon. Nucleic Acids Res. 1982 Nov 25;10(22):7409–7424. doi: 10.1093/nar/10.22.7409. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Norrander J., Kempe T., Messing J. Construction of improved M13 vectors using oligodeoxynucleotide-directed mutagenesis. Gene. 1983 Dec;26(1):101–106. doi: 10.1016/0378-1119(83)90040-9. [DOI] [PubMed] [Google Scholar]
  24. O'Hare K. M., Hayward R. S. Termination of transcription of the coliphage T7 "early" operon in vitro: slowness of enzyme release, and lack of any role for sigma. Nucleic Acids Res. 1981 Sep 25;9(18):4689–4707. doi: 10.1093/nar/9.18.4689. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Olson E. R., Tomich C. S., Friedman D. I. The nusA recognition site. Alteration in its sequence or position relative to upstream translation interferes with the action of the N antitermination function of phage lambda. J Mol Biol. 1984 Dec 25;180(4):1053–1063. doi: 10.1016/0022-2836(84)90270-5. [DOI] [PubMed] [Google Scholar]
  26. Platt T., Squires C., Yanofsky C. Ribosome-protected regions in the leader-trpE sequence of Escherichia coli tryptophan operon messenger RNA. J Mol Biol. 1976 May 15;103(2):411–420. doi: 10.1016/0022-2836(76)90320-x. [DOI] [PubMed] [Google Scholar]
  27. Platt T. Transcription termination and the regulation of gene expression. Annu Rev Biochem. 1986;55:339–372. doi: 10.1146/annurev.bi.55.070186.002011. [DOI] [PubMed] [Google Scholar]
  28. Poulsen P., Bonekamp F., Jensen K. F. Structure of the Escherichia coli pyrE operon and control of pyrE expression by a UTP modulated intercistronic attentuation. EMBO J. 1984 Aug;3(8):1783–1790. doi: 10.1002/j.1460-2075.1984.tb02046.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Robinson A. C., Kenan D. J., Sweeney J., Donachie W. D. Further evidence for overlapping transcriptional units in an Escherichia coli cell envelope-cell division gene cluster: DNA sequence and transcriptional organization of the ddl ftsQ region. J Bacteriol. 1986 Sep;167(3):809–817. doi: 10.1128/jb.167.3.809-817.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Roland K. L., Powell F. E., Turnbough C. L., Jr Role of translation and attenuation in the control of pyrBI operon expression in Escherichia coli K-12. J Bacteriol. 1985 Sep;163(3):991–999. doi: 10.1128/jb.163.3.991-999.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Stanssens P., Remaut E., Fiers W. Inefficient translation initiation causes premature transcription termination in the lacZ gene. Cell. 1986 Mar 14;44(5):711–718. doi: 10.1016/0092-8674(86)90837-8. [DOI] [PubMed] [Google Scholar]
  33. Warren F., Das A. Formation of termination-resistant transcription complex at phage lambda nut locus: effects of altered translation and a ribosomal mutation. Proc Natl Acad Sci U S A. 1984 Jun;81(12):3612–3616. doi: 10.1073/pnas.81.12.3612. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Zalkin H., Yanofsky C., Squires C. L. Regulated in vitro synthesis of Escherichia coli tryptophan operon messenger ribonucleic acid and enzymes. J Biol Chem. 1974 Jan 25;249(2):465–475. [PubMed] [Google Scholar]
  35. Zengel J. M., Mueckl D., Lindahl L. Protein L4 of the E. coli ribosome regulates an eleven gene r protein operon. Cell. 1980 Sep;21(2):523–535. doi: 10.1016/0092-8674(80)90490-0. [DOI] [PubMed] [Google Scholar]

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