Genomics Protocols pp 131-145

Part of the Methods in Molecular Biology™ book series (MIMB, volume 439) | Cite as

Genomewide Identification of Protein Binding Locations Using Chromatin Immunoprecipitation Coupled with Microarray

  • Byung-Kwan Cho
  • Eric M. Knight
  • Bernhard Ø. Palsson

Abstract

Interactions between cis-acting elements and proteins play a key role in transcriptional regulation of all known organisms. To better understand these interactions, researchers developed a method that couples chromatin immunoprecipitation with microarrays (also known as ChIP-chip), which is capable of providing a whole-genome map of protein-DNA interactions. This versatile and high-throughput strategy is initiated by formaldehyde-mediated cross-linking of DNA and proteins, followed by cell lysis, DNA fragmentation, and immunopurification. The immunoprecipitated DNA fragments are then purified from the proteins by reverse-cross-linking followed by amplification, labeling, and hybridization to a whole-genome tiling microarray against a reference sample. The enriched signals obtained from the microarray then are normalized by the reference sample and used to generate the whole-genome map of protein-DNA interactions. The protocol described here has been used for discovering the genomewide distribution of RNA polymerase and several transcription factors of Escherichia coli.

Keywords

ChIP-chip chromatin immunoprecipitation microarray RNA polymerase transcription factor transcription factor binding 

References

  1. 1.
    1. Kim TH, Barrera LO, Zheng M, Qu C, Singer MA, Richmond TA, Wu Y, Green RD, Ren B (2005) A high-resolution map of active promoters in the human genome. Nature 436:876–880PubMedCrossRefGoogle Scholar
  2. 2.
    2. Ren B, Robert R, Wyrick JJ, Aparicio O, Jennings EG, Simon I. Zeitlinger J, Schreiber J, Hannett,N, Kanin E, Volkert TL, Wilson CJ, Bell SP, Young RA (2000) Genome-wide resolution and function of DNA binding proteins. Science 290:2306–2309PubMedCrossRefGoogle Scholar
  3. 3.
    3. Herring CD, Raffaelle M, Allen TE, Kanin EI, Landick R, Ansari AZ, Palsson BO (2005) Immobilization of Escherichia coli RNA polymerase and location of binding sites by use of chromatin immunoprecipitation and microarray. J Bacteriol 187:6166–6174PubMedCrossRefGoogle Scholar
  4. 4.
    4. Lee TI, Rinaldi NJ, Robert F, Odom DT, Bar-Joseph Z, Gerber GK, Hannett NM, Harbison CT, Thompson CM, Simon I, Zeitlinger J, Jennings EG, Murray HL, Gordon DB, Ren B, Wyrick JJ, Tagne J, Volkert TL, Fraenkel E, Gifford DK, Young RA (2002) Transcriptional regulatory networks in Saccharomyces cerevisiae. Science 298:799–804PubMedCrossRefGoogle Scholar
  5. 5.
    5. Wardle FC, Odom DT, Bell GW, Yuan B, Danford TW, Wiellette EL, Herbolsheimer E, Sive HL, Young RA, Smith JC (2006) Zebrafish promoter microarrays identify actively transcribed embryonic genes. Genome Biol 7:R71PubMedCrossRefGoogle Scholar
  6. 6.
    6. Gilchrist M, Thorsson V, Li B, Rust AG, Korb M, Kennedy K, Hai T, Bolouri H, Aderem A (2006) Systems biology approaches identify ATF3 as a negative regulator of Toll-like receptor 4. Nature 441:173–178PubMedCrossRefGoogle Scholar
  7. 7.
    7. Orian A, Steensel BV, Delrow J, Bussemaker HJ, Li L, Sawado T, Williams E, Loo LWM, Cowley SM, Yost C, Pierce S, Edgar BA, Parkhurst,SM, Eisenman RN (2003) Genomic binding by the Drosophila Myc, Max, Mad/Mnt transcription factor network. Genes Dev. 17, 1101–1114.PubMedCrossRefGoogle Scholar
  8. 8.
    8. Chua YL, Mott E, Brown AP, MacLean D, Gray JC (2004) Microarray analysis of chromatin-immunoprecipitated DNA identifies specific regions of tobacco genes associated with acetylated histones. Plant J 37:789–800PubMedCrossRefGoogle Scholar
  9. 9.
    9. Grainger DC, Hurd D, Harrison M, Holdstock J, Busby JW (2005) Studies of the distribution of Escherichia coli cAMP-receptor protein and RNA polymerase along the E. coli chromosome. Proc Natl Acad Sci USA 102:17693–17698PubMedCrossRefGoogle Scholar
  10. 10.
    10. Carter NP, Vetrie D (2004) Applications of genomic microarrays to explore human chromosome structure and function. Hum Mol Genet 13:R297–R302PubMedCrossRefGoogle Scholar
  11. 11.
    11. MacAlpine DM, Bell SP (2005) A genomic view of eukaryotic DNA replication. Chromosome Res 13:309–326PubMedCrossRefGoogle Scholar
  12. 12.
    12. Workman CT, Mak HC, McCuine S, Tagne JB, Agarwal M, Ozier O, Begley TJ, Samson LD, Ideker T (2006) A systems approach to mapping DNA damage response pathways. Science 312:1054–1059PubMedCrossRefGoogle Scholar
  13. 13.
    Wade JT, Roa DC, Grainger DC, Hurd D, Busby JW, Struhl K, Nudler E (2006) Extensive functional overlap between σ factors in Escherichia coli. Nat Struct Mol Biol. doi: 10.1038/ nsmb1130Google Scholar
  14. 14.
    14. Hall DA, Zhu H, Zhu X, Royce T, Gerstein M, Snyder M (2004) Regulation of gene expression by a metabolic enzyme. Science 306:482–484PubMedCrossRefGoogle Scholar
  15. 15.
    15. Kim TH, Ren B (2006) Genome-wide analysis of protein-DNA interactions. Annu Rev Genomics Hum Genet 7:81–102PubMedCrossRefGoogle Scholar
  16. 16.
    16. Lee TI, Johnstone SE, Young RA (2006) Chromatin immunoprecipitation and microarray-based analysis of protein location. Nat Protocols 1:729–748CrossRefGoogle Scholar
  17. 17.
    17. Buck MJ, Lieb JD (2004) ChIP-chip: Consideration for the design, analysis, and application of genome-wide chromatin immunoprecipitation experiments. Genomics 83:349–360PubMedCrossRefGoogle Scholar
  18. 18.
    18. Negre N, Lavrov S, Hennetin J, Bellis M, Cavalli G (2006) Mapping the distribution of chromatin proteins by ChIP on Chip. Methods Enzymol 410:316–341PubMedCrossRefGoogle Scholar
  19. 19.
    19. Hecht A, Grunstein M (1999) Mapping DNA interaction sites of chromosomal proteins using immunoprecipitation and polymerase chain reaction. Methods Enzymol 304:399–414PubMedCrossRefGoogle Scholar
  20. 20.
    20. Ren B,d Dynlacht BD (2004) Use of chromatin immunoprecipitation assays in genome-wide location analysis of mammalian transcription factors. Methods Enzymol 376:304–315PubMedCrossRefGoogle Scholar
  21. 21.
    21. Cho BK, Knight EM, Palsson BØ (2006) PCR-based tandem epitope tagging system for Escherichia coli genome engineering. Biotechniques 40:67–72PubMedCrossRefGoogle Scholar
  22. 22.
    22. Iyer VR, Horak CE, Scafe CS, Botstein D, Snyder M, Brown PO (2001) Genomic binding sites of the yeast cell-cycle transcription factors SBF and MBF. Nature 409:533–538PubMedCrossRefGoogle Scholar
  23. 23.
    23. Buck MJ, Nobel AB, Lieb JD (2005) ChIPOTle: A user-friendly tool for the analysis of ChIP-chip data. Genome Biol 6:R97PubMedCrossRefGoogle Scholar
  24. 24.
    24. Bieda M, Xu X, Singer MA, Green R, Farnham PJ (2006) Unbiased location analysis of E2F1-binding sites suggests a widespread role for E2F1 in the human genome. Genome Res 16:595–605PubMedCrossRefGoogle Scholar
  25. 25.
    25. Ji H, Wong WH (2005) TileMap: Create chromosomal map of tiling array hybridizations. Bioinformatics 21:3629–3636PubMedCrossRefGoogle Scholar
  26. 26.
    26. Qi Y, Rolfe A, MacIsaac KD, Gerber GK, Pokholok D, Zeitlinger J, Danford T, Dowell RD, Fraenkel E, Jaakkola TS, Young RA, Gifford DK (2006) High-resolution computational models of genome binding events. Nat Biotechnol 24:963–970PubMedCrossRefGoogle Scholar
  27. 27.
    27. Johnson WE, Li W, Meyer CA, Gottardo R, Carroll JS, Brown M, Liu XS (2006) Modelbased analysis of tiling-arrays for ChIP-chip. Proc Natl Acad Sci USA 103:12457–12462PubMedCrossRefGoogle Scholar
  28. 28.
    28. Gibbons FD, Proft M, Struhl K, Roth FP (2005) Chipper: Discovering transcription-factor targets from chromatin immunoprecipitation microarray using variance stabilization. Genome Biol 6:R96PubMedCrossRefGoogle Scholar

Copyright information

© Humana Press, a part of Springer Science+Business Media, LLC 2008

Authors and Affiliations

  • Byung-Kwan Cho
    • 1
  • Eric M. Knight
    • 1
  • Bernhard Ø. Palsson
    • 1
  1. 1.Department of BioengineeringUniversity of California-San DiegoLa JollaUSA

Personalised recommendations