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Review
. 2018 Mar 1;17(2):80-88.
doi: 10.1093/bfgp/elx043.

Insights from resolving protein-DNA interactions at near base-pair resolution

Bryan J Venters
Review

Insights from resolving protein-DNA interactions at near base-pair resolution

Bryan J Venters. Brief Funct Genomics. .

Abstract

One of the central goals in molecular biology is to understand how cell-type-specific expression patterns arise through selective recruitment of RNA polymerase II (Pol II) to a subset of gene promoters. Pol II needs to be recruited to a precise genomic position at the proper time to produce messenger RNA from a DNA template. Ostensibly, transcription is a relatively simple cellular process; yet, experimentally measuring and then understanding the combinatorial possibilities of transcriptional regulators remain a daunting task. Since its introduction in 1985, chromatin immunoprecipitation (ChIP) has remained a key tool for investigating protein-DNA contacts in vivo. Over 30 years of intensive research using ChIP have provided numerous insights into mechanisms of gene regulation. As functional genomic technologies improve, they present new opportunities to address key biological questions. ChIP-exo is a refined version of ChIP-seq that significantly reduces background signal, while providing near base-pair mapping resolution for protein-DNA interactions. This review discusses the evolution of the ChIP assay over the years; the methodological differences between ChIP-seq, ChIP-exo and ChIP-nexus; and highlight new insights into epigenetic and transcriptional mechanisms that were uniquely enabled with the near base-pair resolution of ChIP-exo.

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Figures

Figure 1.
Figure 1.
Historical perspective on the development of the ChIP assay and detection methods. (A) Key technological advancements that ultimately led to ChIP-exo/nexus occurred in the following years 1985 [24], 2000 [25], 2007 [26, 27], 2011 [28] and 2015 [29]. (B) Current tallies of publications are shown using ChIP-exo, ChIP-nexus or developing bioinformatic tools specifically for ChIP-exo analysis. (C) Bar graph representation of publications with ‘ChIP-exo’ keyword by year as returned by PubMed.
Figure 2.
Figure 2.
Comparative workflow of ChIP-seq, ChIP-exo and ChIP-nexus technologies.
Figure 3.
Figure 3.
Models for how unique biological insights are revealed with the near base-pair resolution of ChIP-exo. (A) ChIP-exo signals delineate left and right protein–DNA cross-link borders, which enables precise TF motif identification. (B) ChIP-exo enables determination of structurally distinct modes of TF binding, such as the p53 tetrameric complex. (C) ChIP-exo distinguishes clustered TFs, such as the assembly of the enhanceosome. (D) Subnucleosomal structure of histone subunit–DNA contacts can be resolved by ChIP-exo. For example, the H2A.Z histone variant is asymmetrically incorporated into the +1 nucleosome. In addition, the ISW2 chromatin remodeler engages the +1 nucleosome on the promoter proximal side. (E) Ordered recruitment of Pol II elongation machinery at subnucleosomal resolution. (F) ChIP-exo resolves Pol II engaged at the PIC site, promotor proximal pause site and divergently oriented PIC.
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