Profiling Chromatin Accessibility on Replicated DNA with repli-ATAC-Seq.

Open or accessible chromatin typifies euchromatic regions and helps define cell type-specific transcription programs. DNA replication massively disorders chromatin composition and structure, and how accessible regions are affected by and recover from this disruption has been unclear. Here, we present repli-ATAC-seq, a protocol to profile accessible chromatin genome-wide on replicated DNA starting from 100,000 cells.

Recycling of modified H2A-H2B provides short-term memory of chromatin states.

Chromatin landscapes are disrupted during DNA replication and must be restored faithfully to maintain genome regulation and cell identity. The histone H3-H4 modification landscape is restored by parental histone recycling and modification of new histones. How DNA replication impacts on histone H2A-H2B is currently unknown.

Quantifying propagation of DNA methylation and hydroxymethylation with iDEMS.

DNA methylation is a critical epigenetic mark in mammalian cells. Many aspects of DNA methylation maintenance have been characterized; however, the exact kinetics of post-replicative methylation maintenance remain a subject of debate. Here we develop isolation of DNA by 5-ethynyl-deoxyuridine labelling for mass spectrometry (iDEMS), a highly sensitive, quantitative mass spectrometry-based method for measuring DNA modifications on metabolically labelled DNA.

Chromatin replication and epigenetic cell memory.

Propagation of the chromatin landscape across cell divisions is central to epigenetic cell memory. Mechanistic analysis of the interplay between DNA replication, the cell cycle, and the epigenome has provided insights into replication-coupled chromatin assembly and post-replicative chromatin maintenance. These breakthroughs are critical for defining how proliferation impacts the epigenome during cell identity changes in development and disease.

Transcription Restart Establishes Chromatin Accessibility after DNA Replication.

DNA replication is highly disruptive to chromatin, leading to eviction of nucleosomes, RNA polymerase, and regulatory factors. When and how transcription resumes on DNA following DNA replication is unknown. Here we develop a replication-coupled assay for transposase-accessible chromatin (repli-ATAC-seq) to investigate active chromatin restoration post-replication in mouse embryonic stem cells.

A DNMT3A PWWP mutation leads to methylation of bivalent chromatin and growth retardation in mice.

DNA methyltransferases (DNMTs) deposit DNA methylation, which regulates gene expression and is essential for mammalian development. Histone post-translational modifications modulate the recruitment and activity of DNMTs. The PWWP domains of DNMT3A and DNMT3B are posited to interact with histone 3 lysine 36 trimethylation (H3K36me3); however, the functionality of this interaction for DNMT3A remains untested in vivo.

Accurate Recycling of Parental Histones Reproduces the Histone Modification Landscape during DNA Replication.

Chromatin organization is disrupted genome-wide during DNA replication. On newly synthesized DNA, nucleosomes are assembled from new naive histones and old modified histones. It remains unknown whether the landscape of histone post-translational modifications (PTMs) is faithfully copied during DNA replication or the epigenome is perturbed.