High-resolution digital profiling of the epigenome

Key Points

  • The rapid decrease in sequencing costs has led to the recent development of a plethora of epigenomic techniques that use short-read sequencing as a readout.

  • Enzymatic footprinting (using micrococcal nuclease (MNase), deoxyribonuclease (DNase), Tn5 transposase or methyltransferases) provides a means to assess the occupancy of both nucleosomes and non-histone proteins in a single experiment.

  • Enzymatic digestion of chromatin before or after chromatin immunoprecipitation (ChIP) greatly increases its resolution.

  • Mapping the last nucleotide added to a nascent RNA chain provides base-pair resolution maps of RNA polymerase occupancy.

Abstract

The widespread adoption of short-read DNA sequencing as a digital epigenomic readout platform has motivated the development of genome-wide tools that achieve base-pair resolution. New methods for footprinting and affinity purification of nucleosomes, RNA polymerases, chromatin remodellers and transcription factors have increased the resolution of epigenomic profiling by two orders of magnitude, leading to new insights into how the chromatin landscape affects gene regulation. These digital epigenomic tools have also been applied to directly profile both turnover kinetics and transcription in situ. In this Review, we describe how these new genome-wide tools allow interrogation of diverse aspects of the epigenome.

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Figure 1: Methods for chromatin footprinting.
Figure 2: High-resolution ChIP methodologies.
Figure 3: Methods for analysing chromatin dynamics.
Figure 4: Profiling transcription.

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Acknowledgements

The authors thank S. Kasinathan and S. Ramachandran for critical reading of the manuscript and C. Weber for discussions. Work in the authors' laboratory is supported by the US National Institutes of Health grants 5U01 HG004274, U54 CA143862, and R01 ES020116 and by the Howard Hughes Medical Institute.

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Correspondence to Steven Henikoff.

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Glossary

Digital epigenomic analysis

The use of methods with sequencing-based readouts to interrogate the epigenome.

Sequencing library

A collection of DNA fragments prepared for high-throughput sequencing by the addition of specific adapter sequences.

Fragment midpoint-versus-length plots

(V-plots). Representations of paired-end sequencing data in which a point corresponding to the midpoint of a paired-end read is plotted in two-dimensional space. The x coordinate of the point represents the distance of the read midpoint from a defined genomic feature, and its y coordinate represents the length of the fragment from which it was derived.

Affinity reagents

Antibodies or other molecules used to recover specific proteins from a complex mixture.

Tagmentation

Simultaneous fragmentation and incorporation of sequencing adapters into chromatin using Tn5 transposase.

Native ChIP

Chromatin immunoprecipitation (ChIP) using chromatin that has not been crosslinked with formaldehyde or any other crosslinking agent.

Nuclear run-on

A technique in which transcription is reinitated in isolated nuclei to determine the rates at which genes are transcribed.

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Zentner, G., Henikoff, S. High-resolution digital profiling of the epigenome. Nat Rev Genet 15, 814–827 (2014). https://doi.org/10.1038/nrg3798

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