Comprehensive genome-wide protein-DNA interactions detected at single-nucleotide resolution

Ho Sung Rhee, B. Franklin Pugh

Research output: Contribution to journalArticle

407 Citations (Scopus)

Abstract

Chromatin immunoprecipitation (ChIP-chip and ChIP-seq) assays identify where proteins bind throughout a genome. However, DNA contamination and DNA fragmentation heterogeneity produce false positives (erroneous calls) and imprecision in mapping. Consequently, stringent data filtering produces false negatives (missed calls). Here we describe ChIP-exo, where an exonuclease trims ChIP DNA to a precise distance from the crosslinking site. Bound locations are detectable as peak pairs by deep sequencing. Contaminating DNA is degraded or fails to form complementary peak pairs. With the single bp accuracy provided by ChIP-exo, we show an unprecedented view into genome-wide binding of the yeast transcription factors Reb1, Gal4, Phd1, Rap1, and human CTCF. Each of these factors was chosen to address potential limitations of ChIP-exo. We found that binding sites become unambiguous and reveal diverse tendencies governing in vivo DNA-binding specificity that include sequence variants, functionally distinct motifs, motif clustering, secondary interactions, and combinatorial modules within a compound motif.

Original languageEnglish (US)
Pages (from-to)1408-1419
Number of pages12
JournalCell
Volume147
Issue number6
DOIs
StatePublished - Dec 9 2011

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Nucleotides
Genes
Genome
DNA
DNA Contamination
High-Throughput Nucleotide Sequencing
Exonucleases
Proteins
Chromatin Immunoprecipitation
DNA Fragmentation
Cluster Analysis
Transcription Factors
Yeasts
Binding Sites
Crosslinking
Yeast
Chromatin
Assays
Contamination

All Science Journal Classification (ASJC) codes

  • Biochemistry, Genetics and Molecular Biology(all)

Cite this

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abstract = "Chromatin immunoprecipitation (ChIP-chip and ChIP-seq) assays identify where proteins bind throughout a genome. However, DNA contamination and DNA fragmentation heterogeneity produce false positives (erroneous calls) and imprecision in mapping. Consequently, stringent data filtering produces false negatives (missed calls). Here we describe ChIP-exo, where an exonuclease trims ChIP DNA to a precise distance from the crosslinking site. Bound locations are detectable as peak pairs by deep sequencing. Contaminating DNA is degraded or fails to form complementary peak pairs. With the single bp accuracy provided by ChIP-exo, we show an unprecedented view into genome-wide binding of the yeast transcription factors Reb1, Gal4, Phd1, Rap1, and human CTCF. Each of these factors was chosen to address potential limitations of ChIP-exo. We found that binding sites become unambiguous and reveal diverse tendencies governing in vivo DNA-binding specificity that include sequence variants, functionally distinct motifs, motif clustering, secondary interactions, and combinatorial modules within a compound motif.",
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Comprehensive genome-wide protein-DNA interactions detected at single-nucleotide resolution. / Rhee, Ho Sung; Pugh, B. Franklin.

In: Cell, Vol. 147, No. 6, 09.12.2011, p. 1408-1419.

Research output: Contribution to journalArticle

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