Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Research Article
  • Published:

Quantitative whole-genome analysis of DNA-protein interactions by in vivo methylase protection in E. coli

Abstract

A global methylation-based technique was used to identify, display, and quantitate the in vivo occupancy of numerous protein-binding sites within the Escherichia coli genome. The protein occupancy profiles of these sites showed variation across different growth conditions and genetic backgrounds. Of the 25 sites identified in this study, 24 occurred within 5′ noncoding regions. Protein occupancy at 13 of these sites was supported by independent biochemical and genetic evidence. Most of the remaining 12 sites fell upstream of genes with no previously known function. A multivariate statistical analysis was utilized to group such uncharacterized genes with well-characterized ones, providing insights into their function based on a common pattern of transcriptional regulation.

This is a preview of subscription content, access via your institution

Access options

Buy this article

Prices may be subject to local taxes which are calculated during checkout

Similar content being viewed by others

References

  1. Cartwright, I.L., and Kelly, S.E. 1991. Probing the nature of chromosomal DNA-protein contacts by in vivofootprinting. Biotechniques 11: 188–203.

    CAS  PubMed  Google Scholar 

  2. Barras, F., and Marinus, M.G. 1989. The Great GATC:DNA methylation in E. coli. Trends Genet. 5: 139–143.

    Article  CAS  Google Scholar 

  3. Rinquist, S., and Smith, C.L. 1992. The Escherichia coli chromosome contains specific, unmethylated dam and dcm sites. Proc. Natl. Acad. Sci. USA 89: 4539–4543.

    Article  Google Scholar 

  4. Wang, M.X., and Church, G.M. 1992. A whole genome approach to in vivo DNA-protein interactions in E. coli. Nature 360: 606–610.

    Article  CAS  Google Scholar 

  5. Hale, W.B., van der Woude M.W., and Low D.A. 1994. Analysis of Nonmethylated GATC sites in the Escherichia coli chromosome and identification of sites that are differentially methylated in response to environmental stimuli. J. Bact. 176: 3438–3441.

    Article  CAS  Google Scholar 

  6. Prashar, Y., and Weissman, S.M. 1996. Analysis of differential gene expression by display of 3′ end restriction fragments of cDNAs. Proc. Natl. Acad. Sci. USA 93: 659–663.

    Article  CAS  Google Scholar 

  7. Sabourin, D., and Beckwith, J. 1975. Deletion of Escherichia coli crp gene. J. Bacteriol. 122: 338–340.

    CAS  PubMed  PubMed Central  Google Scholar 

  8. Kolb, A., Busby, X., Buc, H., Garges, S., and Adhya, S. 1993. Transcriptional regulation by cAMP and its receptor protein. Annu. Rev. Biochem. 62: 749–795.

    Article  CAS  Google Scholar 

  9. Renbaum, P., Abrahamove, D., Fainsod, A., Wilson, G.G., Rottem, S., and Razin, A. 1990. Cloning, characterization, and expression in Escherichia coli of the gene coding for the CpG DNA methylase from Spiroplasma sp. strain MQ1 (M.SssI). Nucleic Acids Res. 18: 1145–1152.

    Article  CAS  Google Scholar 

  10. Zhang, X., and Mathews, C.K. 1994. Effect of DNA cytosine methylation upon deamination-induced mutagenesis in a natural target sequence in duplex DNA. J. Biol. Chem. 269: 7066–7069.

    CAS  PubMed  Google Scholar 

  11. Kelleher, J., and Raleigh, E.A. 1991. A novel activity in Escherichia coli K-12 that directs restriction of DNA modified at CG dinucleotides. J. Bacteriol. 173: 5220–5223.

    Article  CAS  Google Scholar 

  12. Hall, B.G., and Xu, L. 1992. Nucleotide sequence, function, activation, and evolution of the cryptic asc operon of Escherichia coli K12. Mol. Biol. Evol. 9: 688–706.

    CAS  PubMed  Google Scholar 

  13. Irani, M.H., Orosz, L.M., and Adhya, S. 1983. A control element within a structural gene: The gal operon of Escherichia coli. Cell 32: 783–788.

    CAS  PubMed  Google Scholar 

  14. Weinstein, J.N., Myers, T.G., O'Connor, P.M., Friend, S.H., Fornace, A.J., Jr Kohn, K.W. et al. 1997. An information-intensive approach to the molecular pharmacology of cancer. Science 275: 343–349.

    Article  CAS  Google Scholar 

  15. Kendall, M. 1980. Multivariate analysis. Macmillan, New York.

    Google Scholar 

  16. Norregaard-Madsen, M., Mygind, B., Pedersen, R., Valentin-Hansen, P., and Sogaard-Anderson, L. 1994. The gene encoding the periplasmic cyclophilin homologue, PPIase A, in Escherichia coli, is expressed from four promoters, three of which are activated by the cAMP-CRP complex and negatively regulated by the CytR repressor. Mol. Microbiol. 14: 989–997.

    Article  CAS  Google Scholar 

  17. Ramseier, T.M., and Saier, M.H., Jr. 1995. cAMP-cAMP receptor protein complex: five binding sites in the control region of the Escherichia coli mannitol operon. Microbiology 141: 1901–1907.

    CAS  PubMed  Google Scholar 

  18. Blattner, F.R., Plunkett, G., Bloch, C.A., Perna, N.T., Burland, V., Riley, M. et al. 1997. The complete sequence of Eschericia coli K-12. Science 277: 1453–1462.

    Article  CAS  Google Scholar 

  19. Barras, F., and Marinus, M.G. 1988. Arrangement of Dam methylation sites (GATC) in the Escherichia coli chromosome. Nucleic Acids Res. 16: 9821–9838.

    Article  CAS  Google Scholar 

  20. Charlier, D., Gigot, D., Huysveld, N., Roovers, M., Pierard, A., and Glansdorff, N. 1995. Pyrimidine regulation of the Escherichia coli and Salmonella typhimurium carAB operon: CarP and Intergration Host Factor (IHF) modulate the methylation status of a GATC site present in the control region. J. Mol. Biol. 250: 391–391.

    Google Scholar 

  21. Jovanovic, G., Weiner, L., and Model, P. 1996. Identification, nucleotide sequence, and characterization of PspF, the transcriptional activator of the Escherichia coli stress-induced psp operon. J. Bacteriol. 178: 1936–1945.

    Article  CAS  Google Scholar 

  22. Hoist, B., Sogaard-Anderson, L., Pedersen, H., and Valentin-Hansen, P. 1992. The cAMP-CRP/CytR nucleoprotein complex in Escherichia coli: two pairs of closely linked binding sites for the cAMP-CRP activator complex are involved in combinatorial regulation of the odd promoter. EMBO J. 11: 3635–3643.

    Article  Google Scholar 

  23. Shea, C.M. and Mclntosh, M.A. 1991. Nucleotide sequence and genetic organization of the ferric enterobactin transport system: homology to other periplasmic binding protein-dependent systems in Escherichia coli. Mol. Microbiol. 5: 1 415–1428.

    Article  Google Scholar 

  24. Yamada, M., Asaoka, S., Saier, M.H., and Yamada, Y. 1993. Characterization of the gcd gene from Escherichia coli K-12 W3110 and regulation of its expression. J. Bacteriol. 175: 568–571.

    Article  CAS  Google Scholar 

  25. Kutsukake, K., Ohya, Y. and lino, T. 1989. Transcriptional analysis of the flagellar regulon of Salmonella typhimurium. J. Bacteriol. 177: 741–747.

    Google Scholar 

  26. Andrews, S.C., Harrison, P.M., and Guest, J.R. 1991. A molecular analysis of the 53.4 minute of the Escherichia coli linkage map. J. Gen. Microbiol. 137: 361–367.

    Article  CAS  Google Scholar 

  27. Yamada, M., Saier, M.H., and Yamada, Y. 1988. Positive and negative regulators for glucitol (gut) operon expression in Escherichia coli. J. Mol. Biol. 203: 569–583.

    Article  CAS  Google Scholar 

  28. Xu, J., and Johnson, R.C. 1995. Fis activates the RpoS-dependent stationary-phase expression of proP in Escherichia coli. J. Bacteriol. 177: 5222–5231.

    Article  CAS  Google Scholar 

  29. Weiner, L., Brissette, J.L., Ramani, N., and Model, P. 1995. Analysis of the proteins and c/s-acting elements regulating the stress-induced phage shock protein operon. Nucleic Acid Res. 23: 2030–2036.

    Article  CAS  Google Scholar 

  30. Goodrich, J.A., Schwartz, M.L., and McClure W.R. 1990. 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. 18: 4993–5000.

    Article  CAS  Google Scholar 

  31. Lockhart, D.J., Dong, H.L., Byrne, M.C., Follettie, M.T., Gallo, M.V., Chee, M.S. et al. 1996. Expression monitoring by hybridizatiion to high-density oligonucleotide arrays. Nature Biotechnology 14: 1675–1680.

    Article  CAS  Google Scholar 

  32. DeRisi, J.L., Iyer, V.R., and Brown, P.O. 1997. Exploring the metabolic and genetic control of gene expression on a genomic scale. Science 278: 680–686.

    Article  CAS  Google Scholar 

  33. Gottshcling, D.E. 1992. Telomere-proximal DNA in Saccharomyces cerevisiae is refractory to methyltransferase activity in vivo. Proc. Natl. Acad. Sci. USA 89: 4062–4065.

    Article  Google Scholar 

  34. Ausubel, F.M., Brent, R., Kingston, R.E., Moore, D.D., Seidman, J.G., Smith, J.A. et al. 1989. Current protocols in molecular biology. John Wiley and Sons, New York.

Download references

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to George M. Church.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Tavazoie, S., Church, G. Quantitative whole-genome analysis of DNA-protein interactions by in vivo methylase protection in E. coli. Nat Biotechnol 16, 566–571 (1998). https://doi.org/10.1038/nbt0698-566

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nbt0698-566

This article is cited by

Search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing