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.

  • Article
  • Published:

Global mapping of protein-DNA interactions in vivo by digital genomic footprinting

Abstract

The orchestrated binding of transcriptional activators and repressors to specific DNA sequences in the context of chromatin defines the regulatory program of eukaryotic genomes. We developed a digital approach to assay regulatory protein occupancy on genomic DNA in vivo by dense mapping of individual DNase I cleavages from intact nuclei using massively parallel DNA sequencing. Analysis of >23 million cleavages across the Saccharomyces cerevisiae genome revealed thousands of protected regulatory protein footprints, enabling de novo derivation of factor binding motifs and the identification of hundreds of new binding sites for major regulators. We observed striking correspondence between single-nucleotide resolution DNase I cleavage patterns and protein-DNA interactions determined by crystallography. The data also yielded a detailed view of larger chromatin features including positioned nucleosomes flanking factor binding regions. Digital genomic footprinting should be a powerful approach to delineate the cis-regulatory framework of any organism with an available genome sequence.

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

Access options

Rent or buy this article

Prices vary by article type

from$1.95

to$39.95

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

Figure 1: Digital DNase I analysis of yeast chromatin structure from chromosomal to nucleotide resolution.
Figure 2: Detection of footprints and corresponding sequence motifs.
Figure 3: Mean nucleotide-level accessibility parallels protein-DNA interactions.
Figure 4: Individual yeast regulatory regions and factor binding sites.
Figure 5: Higher-order patterns of DNA accessibility.

Similar content being viewed by others

References

  1. Maniatis, T. & Ptashne, M. Structure of the lambda operators. Nature 246, 133–136 (1973).

    Article  CAS  PubMed  Google Scholar 

  2. Gilbert, W. in Polymerization in Biological Systems 245–259 (Elsevier, North-Holland, Amsterdam, 1972).

    Google Scholar 

  3. Galas, D.J. & Schmitz, A. DNAse footprinting: a simple method for the detection of protein-DNA binding specificity. Nucleic Acids Res. 5, 3157–3170 (1978).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Ren, B. et al. Genome-wide location and function of DNA binding proteins. Science 290, 2306–2309 (2000).

    Article  CAS  PubMed  Google Scholar 

  5. Johnson, D.S., Mortazavi, A., Myers, R.M. & Wold, B. Genome-wide mapping of in vivo protein-DNA interactions. Science 316, 1497–1502 (2007).

    Article  CAS  PubMed  Google Scholar 

  6. Wei, C.L. et al. A global map of p53 transcription-factor binding sites in the human genome. Cell 124, 207–219 (2006).

    Article  CAS  PubMed  Google Scholar 

  7. Gross, D.S. & Garrard, W.T. Nuclease hypersensitive sites in chromatin. Annu. Rev. Biochem. 57, 159–197 (1988).

    Article  CAS  PubMed  Google Scholar 

  8. Sabo, P.J. et al. Genome-scale mapping of DNase I sensitivity in vivo using tiling DNA microarrays. Nat. Methods 3, 511–518 (2006).

    Article  CAS  PubMed  Google Scholar 

  9. Bailey, T.L. & Elkan, C. Fitting a mixture model by expectation maximization to discover motifs in biopolymers. Proc. Int. Conf. Intell. Syst. Mol. Biol. 2, 28–36 (1994).

    CAS  PubMed  Google Scholar 

  10. MacIsaac, K.D. et al. An improved map of conserved regulatory sites for Saccharomyces cerevisiae. BMC Bioinformatics 7, 113 (2006).

    Article  PubMed  PubMed Central  Google Scholar 

  11. Siepel, A. et al. Evolutionarily conserved elements in vertebrate, insect, worm, and yeast genomes. Genome Res. 15, 1034–1050 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Harbison, C.T. et al. Transcriptional regulatory code of a eukaryotic genome. Nature 431, 99–104 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Cliften, P. et al. Finding functional features in Saccharomyces genomes by phylogenetic footprinting. Science 301, 71–76 (2003).

    Article  CAS  PubMed  Google Scholar 

  14. Kellis, M., Patterson, N., Endrizzi, M., Birren, B. & Lander, E.S. Sequencing and comparison of yeast species to identify genes and regulatory elements. Nature 423, 241–254 (2003).

    Article  CAS  PubMed  Google Scholar 

  15. Borneman, A.R. et al. Divergence of transcription factor binding sites across related yeast species. Science 317, 815–819 (2007).

    Article  CAS  PubMed  Google Scholar 

  16. Tan, S. & Richmond, T.J. Crystal structure of the yeast MATalpha2/MCM1/DNA ternary complex. Nature 391, 660–666 (1998).

    Article  CAS  PubMed  Google Scholar 

  17. Ferre-D'Amare, A.R., Pognonec, P., Roeder, R.G. & Burley, S.K. Structure and function of the b/HLH/Z domain of USF. EMBO J. 13, 180–189 (1994).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Acton, T.B., Zhong, H. & Vershon, A.K. DNA-binding specificity of Mcm1: operator mutations that alter DNA-bending and transcriptional activities by a MADS box protein. Mol. Cell. Biol. 17, 1881–1889 (1997).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Wang, K.L. & Warner, J.R. Positive and negative autoregulation of REB1 transcription in Saccharomyces cerevisiae. Mol. Cell. Biol. 18, 4368–4376 (1998).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Planta, R.J., Goncalves, P.M. & Mager, W.H. Global regulators of ribosome biosynthesis in yeast. Biochem. Cell Biol. 73, 825–834 (1995).

    Article  CAS  PubMed  Google Scholar 

  21. Boeger, H., Griesenbeck, J. & Kornberg, R.D. Nucleosome retention and the stochastic nature of promoter chromatin remodeling for transcription. Cell 133, 716–726 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Mavrich, T.N. et al. A barrier nucleosome model for statistical positioning of nucleosomes throughout the yeast genome. Genome Res. 18, 1073–1083 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Lee, W. et al. A high-resolution atlas of nucleosome occupancy in yeast. Nat. Genet. 39, 1235–1244 (2007).

    Article  CAS  PubMed  Google Scholar 

  24. Shivaswamy, S. et al. Dynamic remodeling of individual nucleosomes across a eukaryotic genome in response to transcriptional perturbation. PLoS Biol. 6, e65 (2008).

    Article  PubMed  PubMed Central  Google Scholar 

  25. Yuan, G.C. et al. Genome-scale identification of nucleosome positions in S. cerevisiae. Science 309, 626–630 (2005).

    Article  CAS  PubMed  Google Scholar 

  26. Raisner, R.M. et al. Histone variant H2A.Z marks the 5′ ends of both active and inactive genes in euchromatin. Cell 123, 233–248 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Jakobsen, B.K. & Pelham, H.R. Constitutive binding of yeast heat shock factor to DNA in vivo. Mol. Cell. Biol. 8, 5040–5042 (1988).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Strauss, E.C. & Orkin, S.H. In vivo protein-DNA interactions at hypersensitive site 3 of the human beta-globin locus control region. Proc. Natl. Acad. Sci. USA 89, 5809–5813 (1992).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. David, L. et al. A high-resolution map of transcription in the yeast genome. Proc. Natl. Acad. Sci. USA 103, 5320–5325 (2006).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

We thank the staff of the University of Washington Genome Sciences High-Throughput Genomics Unit for technical assistance with Illumina-Solexa sequencing, and members of the Stamatoyannopoulos and Fields laboratories for many helpful discussions. This work was supported by US National Institutes of Health grants R01GM071923 and U54HG004592 to J.A.S., and P41RR11823 to S.F. and W.S.N.; X.C. was supported by a fellowship from the Natural Sciences and Engineering Research Council of Canada (NSERC PGS D3).

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to John A Stamatoyannopoulos.

Ethics declarations

Competing interests

Z.Z. is presently an employee of Illumina, Inc.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–7, Supplementary Tables 1–4, Supplementary Methods (PDF 2438 kb)

Supplementary Software

Footprint detection software. (ZIP 9 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Hesselberth, J., Chen, X., Zhang, Z. et al. Global mapping of protein-DNA interactions in vivo by digital genomic footprinting. Nat Methods 6, 283–289 (2009). https://doi.org/10.1038/nmeth.1313

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nmeth.1313

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