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. 1991 Sep;173(18):5800–5807. doi: 10.1128/jb.173.18.5800-5807.1991

Integration host factor binds specifically to multiple sites in the ompB promoter of Escherichia coli and inhibits transcription.

P Tsui 1, L Huang 1, M Freundlich 1
PMCID: PMC208313  PMID: 1885551

Abstract

Escherichia coli integration host factor (IHF) is a DNA-binding protein that participates in gene regulation, site-specific recombination, and other processes in E. coli and some of its bacteriophages and plasmids. In the present study, we showed that IHF is a direct negative effector of the ompB operon of E. coli. Gel retardation experiments and DNase I footprinting studies revealed that IHF binds to three sites in the ompB promoter region. In vitro transcription from ompB promoter fragments was specifically blocked by IHF. In vivo experiments showed that IHF is a negative effector of ompB expression in growing cells. Analysis of IHF binding site mutations strongly suggested that IHF binding in the ompB promoter region is necessary for the negative effects seen in vivo.

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Selected References

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  1. Aiba H., Mizuno T. Phosphorylation of a bacterial activator protein, OmpR, by a protein kinase, EnvZ, stimulates the transcription of the ompF and ompC genes in Escherichia coli. FEBS Lett. 1990 Feb 12;261(1):19–22. doi: 10.1016/0014-5793(90)80626-t. [DOI] [PubMed] [Google Scholar]
  2. Berman M. L., Jackson D. E. Selection of lac gene fusions in vivo: ompR-lacZ fusions that define a functional domain of the ompR gene product. J Bacteriol. 1984 Aug;159(2):750–756. doi: 10.1128/jb.159.2.750-756.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Cannon W. V., Kreutzer R., Kent H. M., Morett E., Buck M. Activation of the Klebsiella pneumoniae nifU promoter: identification of multiple and overlapping upstream NifA binding sites. Nucleic Acids Res. 1990 Apr 11;18(7):1693–1701. doi: 10.1093/nar/18.7.1693. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Coleman J., Green P. J., Inouye M. The use of RNAs complementary to specific mRNAs to regulate the expression of individual bacterial genes. Cell. 1984 Jun;37(2):429–436. doi: 10.1016/0092-8674(84)90373-8. [DOI] [PubMed] [Google Scholar]
  5. Comeau D. E., Ikenaka K., Tsung K. L., Inouye M. Primary characterization of the protein products of the Escherichia coli ompB locus: structure and regulation of synthesis of the OmpR and EnvZ proteins. J Bacteriol. 1985 Nov;164(2):578–584. doi: 10.1128/jb.164.2.578-584.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Craig N. L., Nash H. A. E. coli integration host factor binds to specific sites in DNA. Cell. 1984 Dec;39(3 Pt 2):707–716. doi: 10.1016/0092-8674(84)90478-1. [DOI] [PubMed] [Google Scholar]
  7. Drlica K., Rouviere-Yaniv J. Histonelike proteins of bacteria. Microbiol Rev. 1987 Sep;51(3):301–319. doi: 10.1128/mr.51.3.301-319.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Forst S., Comeau D., Norioka S., Inouye M. Localization and membrane topology of EnvZ, a protein involved in osmoregulation of OmpF and OmpC in Escherichia coli. J Biol Chem. 1987 Dec 5;262(34):16433–16438. [PubMed] [Google Scholar]
  9. Fried M., Crothers D. M. Equilibria and kinetics of lac repressor-operator interactions by polyacrylamide gel electrophoresis. Nucleic Acids Res. 1981 Dec 11;9(23):6505–6525. doi: 10.1093/nar/9.23.6505. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Friedman D. I. Integration host factor: a protein for all reasons. Cell. 1988 Nov 18;55(4):545–554. doi: 10.1016/0092-8674(88)90213-9. [DOI] [PubMed] [Google Scholar]
  11. Galas D. J., Schmitz A. DNAse footprinting: a simple method for the detection of protein-DNA binding specificity. Nucleic Acids Res. 1978 Sep;5(9):3157–3170. doi: 10.1093/nar/5.9.3157. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Garner M. M., Revzin A. A gel electrophoresis method for quantifying the binding of proteins to specific DNA regions: application to components of the Escherichia coli lactose operon regulatory system. Nucleic Acids Res. 1981 Jul 10;9(13):3047–3060. doi: 10.1093/nar/9.13.3047. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Giladi H., Gottesman M., Oppenheim A. B. Integration host factor stimulates the phage lambda pL promoter. J Mol Biol. 1990 May 5;213(1):109–121. doi: 10.1016/S0022-2836(05)80124-X. [DOI] [PubMed] [Google Scholar]
  14. Gober J. W., Shapiro L. Integration host factor is required for the activation of developmentally regulated genes in Caulobacter. Genes Dev. 1990 Sep;4(9):1494–1504. doi: 10.1101/gad.4.9.1494. [DOI] [PubMed] [Google Scholar]
  15. Goodman S. D., Nash H. A. Functional replacement of a protein-induced bend in a DNA recombination site. Nature. 1989 Sep 21;341(6239):251–254. doi: 10.1038/341251a0. [DOI] [PubMed] [Google Scholar]
  16. Goodrich J. A., Schwartz M. L., McClure W. R. Searching for and predicting the activity of sites for DNA binding proteins: compilation and analysis of the binding sites for Escherichia coli integration host factor (IHF). Nucleic Acids Res. 1990 Sep 11;18(17):4993–5000. doi: 10.1093/nar/18.17.4993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Griffo G., Oppenheim A. B., Gottesman M. E. Repression of the lambda pcin promoter by integrative host factor. J Mol Biol. 1989 Sep 5;209(1):55–64. doi: 10.1016/0022-2836(89)90169-1. [DOI] [PubMed] [Google Scholar]
  18. Hall M. N., Silhavy T. J. Genetic analysis of the ompB locus in Escherichia coli K-12. J Mol Biol. 1981 Sep 5;151(1):1–15. doi: 10.1016/0022-2836(81)90218-7. [DOI] [PubMed] [Google Scholar]
  19. Hall M. N., Silhavy T. J. The ompB locus and the regulation of the major outer membrane porin proteins of Escherichia coli K12. J Mol Biol. 1981 Feb 15;146(1):23–43. doi: 10.1016/0022-2836(81)90364-8. [DOI] [PubMed] [Google Scholar]
  20. Higgins N. P., Collier D. A., Kilpatrick M. W., Krause H. M. Supercoiling and integration host factor change the DNA conformation and alter the flow of convergent transcription in phage Mu. J Biol Chem. 1989 Feb 15;264(5):3035–3042. [PubMed] [Google Scholar]
  21. Hoover T. R., Santero E., Porter S., Kustu S. The integration host factor stimulates interaction of RNA polymerase with NIFA, the transcriptional activator for nitrogen fixation operons. Cell. 1990 Oct 5;63(1):11–22. doi: 10.1016/0092-8674(90)90284-l. [DOI] [PubMed] [Google Scholar]
  22. Huang L., Tsui P., Freundlich M. Integration host factor is a negative effector of in vivo and in vitro expression of ompC in Escherichia coli. J Bacteriol. 1990 Sep;172(9):5293–5298. doi: 10.1128/jb.172.9.5293-5298.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Igo M. M., Ninfa A. J., Silhavy T. J. A bacterial environmental sensor that functions as a protein kinase and stimulates transcriptional activation. Genes Dev. 1989 May;3(5):598–605. doi: 10.1101/gad.3.5.598. [DOI] [PubMed] [Google Scholar]
  24. Igo M. M., Ninfa A. J., Stock J. B., Silhavy T. J. Phosphorylation and dephosphorylation of a bacterial transcriptional activator by a transmembrane receptor. Genes Dev. 1989 Nov;3(11):1725–1734. doi: 10.1101/gad.3.11.1725. [DOI] [PubMed] [Google Scholar]
  25. Jo Y. L., Nara F., Ichihara S., Mizuno T., Mizushima S. Purification and characterization of the OmpR protein, a positive regulator involved in osmoregulatory expression of the ompF and ompC genes in Escherichia coli. J Biol Chem. 1986 Nov 15;261(32):15252–15256. [PubMed] [Google Scholar]
  26. Krause H. M., Higgins N. P. Positive and negative regulation of the Mu operator by Mu repressor and Escherichia coli integration host factor. J Biol Chem. 1986 Mar 15;261(8):3744–3752. [PubMed] [Google Scholar]
  27. Kur J., Hasan N., Szybalski W. Physical and biological consequences of interactions between integration host factor (IHF) and coliphage lambda late p'R promoter and its mutants. Gene. 1989 Sep 1;81(1):1–15. doi: 10.1016/0378-1119(89)90331-4. [DOI] [PubMed] [Google Scholar]
  28. Leong J. M., Nunes-Düby S., Lesser C. F., Youderian P., Susskind M. M., Landy A. The phi 80 and P22 attachment sites. Primary structure and interaction with Escherichia coli integration host factor. J Biol Chem. 1985 Apr 10;260(7):4468–4477. [PubMed] [Google Scholar]
  29. Mahajna J., Oppenheim A. B., Rattray A., Gottesman M. Translation initiation of bacteriophage lambda gene cII requires integration host factor. J Bacteriol. 1986 Jan;165(1):167–174. doi: 10.1128/jb.165.1.167-174.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Mitchell P. J., Tjian R. Transcriptional regulation in mammalian cells by sequence-specific DNA binding proteins. Science. 1989 Jul 28;245(4916):371–378. doi: 10.1126/science.2667136. [DOI] [PubMed] [Google Scholar]
  31. Nash H. A., Robertson C. A., Flamm E., Weisberg R. A., Miller H. I. Overproduction of Escherichia coli integration host factor, a protein with nonidentical subunits. J Bacteriol. 1987 Sep;169(9):4124–4127. doi: 10.1128/jb.169.9.4124-4127.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Nash H. A., Robertson C. A. Purification and properties of the Escherichia coli protein factor required for lambda integrative recombination. J Biol Chem. 1981 Sep 10;256(17):9246–9253. [PubMed] [Google Scholar]
  33. Norioka S., Ramakrishnan G., Ikenaka K., Inouye M. Interaction of a transcriptional activator, OmpR, with reciprocally osmoregulated genes, ompF and ompC, of Escherichia coli. J Biol Chem. 1986 Dec 25;261(36):17113–17119. [PubMed] [Google Scholar]
  34. Okamoto K., Freundlich M. Mechanism for the autogenous control of the crp operon: transcriptional inhibition by a divergent RNA transcript. Proc Natl Acad Sci U S A. 1986 Jul;83(14):5000–5004. doi: 10.1073/pnas.83.14.5000. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Pereira R. F., Ortuno M. J., Lawther R. P. Binding of integration host factor (IHF) to the ilvGp1 promoter of the ilvGMEDA operon of Escherichia coli K12. Nucleic Acids Res. 1988 Jul 11;16(13):5973–5989. doi: 10.1093/nar/16.13.5973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Prentki P., Chandler M., Galas D. J. Escherichia coli integration host factor bends the DNA at the ends of IS1 and in an insertion hotspot with multiple IHF binding sites. EMBO J. 1987 Aug;6(8):2479–2487. doi: 10.1002/j.1460-2075.1987.tb02529.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Robertson C. A., Nash H. A. Bending of the bacteriophage lambda attachment site by Escherichia coli integration host factor. J Biol Chem. 1988 Mar 15;263(8):3554–3557. [PubMed] [Google Scholar]
  38. Santero E., Hoover T., Keener J., Kustu S. In vitro activity of the nitrogen fixation regulatory protein NIFA. Proc Natl Acad Sci U S A. 1989 Oct;86(19):7346–7350. doi: 10.1073/pnas.86.19.7346. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. 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]
  40. Stenzel T. T., Patel P., Bastia D. The integration host factor of Escherichia coli binds to bent DNA at the origin of replication of the plasmid pSC101. Cell. 1987 Jun 5;49(5):709–717. doi: 10.1016/0092-8674(87)90547-2. [DOI] [PubMed] [Google Scholar]
  41. Straney S. B., Crothers D. M. Lac repressor is a transient gene-activating protein. Cell. 1987 Dec 4;51(5):699–707. doi: 10.1016/0092-8674(87)90093-6. [DOI] [PubMed] [Google Scholar]
  42. Thompson J. F., Landy A. Empirical estimation of protein-induced DNA bending angles: applications to lambda site-specific recombination complexes. Nucleic Acids Res. 1988 Oct 25;16(20):9687–9705. doi: 10.1093/nar/16.20.9687. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Thompson J. F., Waechter-Brulla D., Gumport R. I., Gardner J. F., Moitoso de Vargas L., Landy A. Mutations in an integration host factor-binding site: effect on lambda site-specific recombination and regulatory implications. J Bacteriol. 1986 Dec;168(3):1343–1351. doi: 10.1128/jb.168.3.1343-1351.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Thompson R. J., Mosig G. Integration host factor (IHF) represses a Chlamydomonas chloroplast promoter in E. coli. Nucleic Acids Res. 1988 Apr 25;16(8):3313–3326. doi: 10.1093/nar/16.8.3313. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Tsui P., Freundlich M. Integration host factor bends the DNA in the Escherichia coli ilvBN promoter region. Mol Gen Genet. 1990 Sep;223(2):349–352. doi: 10.1007/BF00265076. [DOI] [PubMed] [Google Scholar]
  46. Tsui P., Freundlich M. Integration host factor binds specifically to sites in the ilvGMEDA operon in Escherichia coli. J Mol Biol. 1988 Oct 5;203(3):817–820. doi: 10.1016/0022-2836(88)90212-4. [DOI] [PubMed] [Google Scholar]
  47. Tsui P., Helu V., Freundlich M. Altered osmoregulation of ompF in integration host factor mutants of Escherichia coli. J Bacteriol. 1988 Oct;170(10):4950–4953. doi: 10.1128/jb.170.10.4950-4953.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Vershon A. K., Liao S. M., McClure W. R., Sauer R. T. Interaction of the bacteriophage P22 Arc repressor with operator DNA. J Mol Biol. 1987 May 20;195(2):323–331. doi: 10.1016/0022-2836(87)90653-x. [DOI] [PubMed] [Google Scholar]
  49. Wiater L. A., Grindley N. D. Gamma delta transposase and integration host factor bind cooperatively at both ends of gamma delta. EMBO J. 1988 Jun;7(6):1907–1911. doi: 10.1002/j.1460-2075.1988.tb03024.x. [DOI] [PMC free article] [PubMed] [Google Scholar]

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