Skip to main content
PMC home page
Nucleic Acids Research logoLink to Nucleic Acids Research
. 1987 Feb 25;15(4):1521–1542. doi: 10.1093/nar/15.4.1521

Effect of the deletion of upstream DNA sequences on expression from the ilvGp2 promoter of the ilvGMEDA operon of Escherichia coli K-12.

M J Ortuno, R P Lawther
PMCID: PMC340565  PMID: 3547337

Abstract

Transcription in vitro of the regulatory region of the ilvGMEDA operon yields two attenuated RNAs initiated from the tandem promoters ilvGp1 and ilvGp2. Both S1 nuclease analysis and the fusion of ilvGp1 to galK indicate that transcription is not initiated in vivo from ilvGp1. However deletion of DNA sequences 150 to 100 bp upstream of ilvGp2 drastically reduces expression in vivo from ilvGp2. Both the distance separating ilvGp2 from the upstream DNA sequences and their relative orientation to each other on the DNA helix affect expression from ilvGp2. Deletion of DNA sequences approximately 400 bp upstream of ilvGp2 increases expression in vivo from this promoter. Analysis of products of transcription in vitro indicates that the effects observed in vivo are probably not due to DNA conformation or interactions of RNA polymerase.

Full text

PDF
1526

Images in this article

1528Image
on p.1528
1534Image
on p.1534

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. Adams C. W., Rosenberg M., Hatfield G. W. Analysis of in vivo RNA transcription products of the ilvGEDA attenuator region of Escherichia coli K12. J Biol Chem. 1985 Jul 15;260(14):8538–8544. [PubMed] [Google Scholar]
  3. Altman S., Smith J. D. Tyrosine tRNA precursor molecule polynucleotide sequence. Nat New Biol. 1971 Sep 8;233(36):35–39. doi: 10.1038/newbio233035a0. [DOI] [PubMed] [Google Scholar]
  4. Bachmann B. J., Low K. B. Linkage map of Escherichia coli K-12, edition 6. Microbiol Rev. 1980 Mar;44(1):1–56. doi: 10.1128/mr.44.1.1-56.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Berg C. M., Shaw K. J., Vender J., Borucka-Mankiewicz M. Physiological characterization of polar Tn5-induced isoleucine-valine auxotrophs in Escherichia coli K.12: evidence for an internal promoter in the ilvOGEDA operon. Genetics. 1979 Oct;93(2):308–319. [PMC free article] [PubMed] [Google Scholar]
  6. 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]
  7. Bossi L., Smith D. M. Conformational change in the DNA associated with an unusual promoter mutation in a tRNA operon of Salmonella. Cell. 1984 Dec;39(3 Pt 2):643–652. doi: 10.1016/0092-8674(84)90471-9. [DOI] [PubMed] [Google Scholar]
  8. Dunn T. M., Hahn S., Ogden S., Schleif R. F. An operator at -280 base pairs that is required for repression of araBAD operon promoter: addition of DNA helical turns between the operator and promoter cyclically hinders repression. Proc Natl Acad Sci U S A. 1984 Aug;81(16):5017–5020. doi: 10.1073/pnas.81.16.5017. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Friden P., Voelkel K., Sternglanz R., Freundlich M. Reduced expression of the isoleucine and valine enzymes in integration host factor mutants of Escherichia coli. J Mol Biol. 1984 Feb 5;172(4):573–579. doi: 10.1016/s0022-2836(84)80024-8. [DOI] [PubMed] [Google Scholar]
  10. Friedman D. I., Olson E. J., Carver D., Gellert M. Synergistic effect of himA and gyrB mutations: evidence that him functions control expression of ilv and xyl genes. J Bacteriol. 1984 Feb;157(2):484–489. doi: 10.1128/jb.157.2.484-489.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Gilmour D. S., Lis J. T. Detecting protein-DNA interactions in vivo: distribution of RNA polymerase on specific bacterial genes. Proc Natl Acad Sci U S A. 1984 Jul;81(14):4275–4279. doi: 10.1073/pnas.81.14.4275. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Haughn G. W., Squires C. H., De Felice M., Largo C. T., Calvo J. M. Unusual organization of the ilvIH promoter of Escherichia coli. J Bacteriol. 1985 Jul;163(1):186–198. doi: 10.1128/jb.163.1.186-198.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Haughn G. W., Wessler S. R., Gemmill R. M., Calvo J. M. High A + T content conserved in DNA sequences upstream of leuABCD in Escherichia coli and Salmonella typhimurium. J Bacteriol. 1986 Jun;166(3):1113–1117. doi: 10.1128/jb.166.3.1113-1117.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Hawley D. K., McClure W. R. Compilation and analysis of Escherichia coli promoter DNA sequences. Nucleic Acids Res. 1983 Apr 25;11(8):2237–2255. doi: 10.1093/nar/11.8.2237. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Hochschild A., Ptashne M. Cooperative binding of lambda repressors to sites separated by integral turns of the DNA helix. Cell. 1986 Mar 14;44(5):681–687. doi: 10.1016/0092-8674(86)90833-0. [DOI] [PubMed] [Google Scholar]
  16. Horn G. T., Wells R. D. The leftward promoter of bacteriophage lambda. Structure, biological activity, and influence by adjacent regions. J Biol Chem. 1981 Feb 25;256(4):2003–2009. [PubMed] [Google Scholar]
  17. Johnson D. I., Somerville R. L. New regulatory genes involved in the control of transcription initiation at the thr and ilv promoters of Escherichia coli K-12. Mol Gen Genet. 1984;195(1-2):70–76. doi: 10.1007/BF00332726. [DOI] [PubMed] [Google Scholar]
  18. Jones H. M., Brajkovich C. M., Gunsalus R. P. In vivo 5' terminus and length of the mRNA for the proton-translocating ATPase (unc) operon of Escherichia coli. J Bacteriol. 1983 Sep;155(3):1279–1287. doi: 10.1128/jb.155.3.1279-1287.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Küpper H., Contreras R., Landy A., Khorana H. G. Promoter-dependent transcription of tRNAITyr genes using DNA fragments produced by restriction enzymes. Proc Natl Acad Sci U S A. 1975 Dec;72(12):4754–4758. doi: 10.1073/pnas.72.12.4754. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Lamond A. I., Travers A. A. Requirement for an upstream element for optimal transcription of a bacterial tRNA gene. Nature. 1983 Sep 15;305(5931):248–250. doi: 10.1038/305248a0. [DOI] [PubMed] [Google Scholar]
  21. Lawther R. P., Hatfield G. W. Multivalent translational control of transcription termination at attenuator of ilvGEDA operon of Escherichia coli K-12. Proc Natl Acad Sci U S A. 1980 Apr;77(4):1862–1866. doi: 10.1073/pnas.77.4.1862. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Lawther R. P., Nichols B., Zurawski G., Hatfield G. W. The nucleotide sequence preceding and including the beginning of the ilvE gene of the ilvGEDA operon of Escherichia coli K12. Nucleic Acids Res. 1979 Dec 20;7(8):2289–2301. doi: 10.1093/nar/7.8.2289. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Lopes J. M., Lawther R. P. Analysis and comparison of the internal promoter, pE, of the ilvGMEDA operons from Escherichia coli K-12 and Salmonella typhimurium. Nucleic Acids Res. 1986 Mar 25;14(6):2779–2798. doi: 10.1093/nar/14.6.2779. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Maxam A. M., Gilbert W. Sequencing end-labeled DNA with base-specific chemical cleavages. Methods Enzymol. 1980;65(1):499–560. doi: 10.1016/s0076-6879(80)65059-9. [DOI] [PubMed] [Google Scholar]
  25. 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]
  26. Miller K. G., Sollner-Webb B. Transcription of mouse rRNA genes by RNA polymerase I: in vitro and in vivo initiation and processing sites. Cell. 1981 Nov;27(1 Pt 2):165–174. doi: 10.1016/0092-8674(81)90370-6. [DOI] [PubMed] [Google Scholar]
  27. Moyed H. S., Nguyen T. T., Bertrand K. P. Multicopy Tn10 tet plasmids confer sensitivity to induction of tet gene expression. J Bacteriol. 1983 Aug;155(2):549–556. doi: 10.1128/jb.155.2.549-556.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Nakamura K., Inouye M. DNA sequence of the gene for the outer membrane lipoprotein of E. coli: an extremely AT-rich promoter. Cell. 1979 Dec;18(4):1109–1117. doi: 10.1016/0092-8674(79)90224-1. [DOI] [PubMed] [Google Scholar]
  29. Nargang F. E., Subrahmanyam C. S., Umbarger H. E. Nucleotide sequence of ilvGEDA operon attenuator region of Escherichia coli. Proc Natl Acad Sci U S A. 1980 Apr;77(4):1823–1827. doi: 10.1073/pnas.77.4.1823. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Nick H., Gilbert W. Detection in vivo of protein-DNA interactions within the lac operon of Escherichia coli. 1985 Feb 28-Mar 6Nature. 313(6005):795–798. doi: 10.1038/313795a0. [DOI] [PubMed] [Google Scholar]
  31. Nishi T., Itoh S. Enhancement of transcriptional activity of the Escherichia coli trp promoter by upstream A + T-rich regions. Gene. 1986;44(1):29–36. doi: 10.1016/0378-1119(86)90039-9. [DOI] [PubMed] [Google Scholar]
  32. Reitzer L. J., Magasanik B. Transcription of glnA in E. coli is stimulated by activator bound to sites far from the promoter. Cell. 1986 Jun 20;45(6):785–792. doi: 10.1016/0092-8674(86)90553-2. [DOI] [PubMed] [Google Scholar]
  33. Rosenberg M., Court D. Regulatory sequences involved in the promotion and termination of RNA transcription. Annu Rev Genet. 1979;13:319–353. doi: 10.1146/annurev.ge.13.120179.001535. [DOI] [PubMed] [Google Scholar]
  34. Taillon M. P., Gotto D. A., Lawther R. P. The DNA sequence of the promoter-attenuator of the ilvGEDA operon of Salmonella typhimurium. Nucleic Acids Res. 1981 Jul 24;9(14):3419–3432. doi: 10.1093/nar/9.14.3419. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Travers A. A., Lamond A. I., Mace H. A., Berman M. L. RNA polymerase interactions with the upstream region of the E. coli tyrT promoter. Cell. 1983 Nov;35(1):265–273. doi: 10.1016/0092-8674(83)90229-5. [DOI] [PubMed] [Google Scholar]
  36. Tuggle C. K., Fuchs J. A. Regulation of the operon encoding ribonucleotide reductase in Escherichia coli: evidence for both positive and negative control. EMBO J. 1986 May;5(5):1077–1085. doi: 10.1002/j.1460-2075.1986.tb04325.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Vieira J., Messing J. The pUC plasmids, an M13mp7-derived system for insertion mutagenesis and sequencing with synthetic universal primers. Gene. 1982 Oct;19(3):259–268. doi: 10.1016/0378-1119(82)90015-4. [DOI] [PubMed] [Google Scholar]
  38. Wek R. C., Hatfield G. W. Examination of the internal promoter, PE, in the ilvGMEDA operon of E. coli K-12. Nucleic Acids Res. 1986 Mar 25;14(6):2763–2777. doi: 10.1093/nar/14.6.2763. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Wek R. C., Hauser C. A., Hatfield G. W. The nucleotide sequence of the ilvBN operon of Escherichia coli: sequence homologies of the acetohydroxy acid synthase isozymes. Nucleic Acids Res. 1985 Jun 11;13(11):3995–4010. doi: 10.1093/nar/13.11.3995. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Nucleic Acids Research are provided here courtesy of Oxford University Press

RESOURCES