Efficient chromatin accessibility mapping in situ by nucleosome-tethered tagmentation
Abstract
Chromatin accessibility mapping is a powerful approach to identify potential regulatory elements. A popular example is ATAC-seq, whereby Tn5 transposase inserts sequencing adapters into accessible DNA ('tagmentation'). CUT&Tag is a tagmentation-based epigenomic profiling method in which antibody tethering of Tn5 to a chromatin epitope of interest profiles specific chromatin features in small samples and single cells. Here we show that by simply modifying the tagmentation conditions for histone H3K4me2 or H3K4me3 CUT&Tag, antibody-tethered tagmentation of accessible DNA sites is redirected to produce chromatin accessibility maps that are indistinguishable from the best ATAC-seq maps. Thus, chromatin accessibility maps can be produced in parallel with CUT&Tag maps of other epitopes with all steps from nuclei to amplified sequencing-ready libraries performed in single PCR tubes in the laboratory or on a home workbench. As H3K4 methylation is produced by transcription at promoters and enhancers, our method identifies transcription-coupled accessible regulatory sites.
Data availability
Sequencing data have been deposited in GEO under accession code GSE158327
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Efficient transcription-coupled chromatin accessibility mapping in situNCBI Gene Expression Omnibus, GSE158327.
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CUT&Tag for efficient epigenomic profiling of small samples and single cellsNCBI Gene Expression Omnibus, GSE124557.
Article and author information
Author details
Funding
National Institutes of Health (R01 HG010492)
- Steven Henikoff
National Institutes of Health (R01 GM108699)
- Kami Ahmad
Chan Zuckerberg Initiative (Fred Hutch HCA Seed Network)
- Steven Henikoff
- Kami Ahmad
Howard Hughes Medical Institute (Henikoff)
- Steven Henikoff
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2020, Henikoff et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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Further reading
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- Cell Biology
- Chromosomes and Gene Expression
During oncogene-induced senescence there are striking changes in the organisation of heterochromatin in the nucleus. This is accompanied by activation of a pro-inflammatory gene expression programme – the senescence-associated secretory phenotype (SASP) – driven by transcription factors such as NF-κB. The relationship between heterochromatin re-organisation and the SASP has been unclear. Here, we show that TPR, a protein of the nuclear pore complex basket required for heterochromatin re-organisation during senescence, is also required for the very early activation of NF-κB signalling during the stress-response phase of oncogene-induced senescence. This is prior to activation of the SASP and occurs without affecting NF-κB nuclear import. We show that TPR is required for the activation of innate immune signalling at these early stages of senescence and we link this to the formation of heterochromatin-enriched cytoplasmic chromatin fragments thought to bleb off from the nuclear periphery. We show that HMGA1 is also required for cytoplasmic chromatin fragment formation. Together these data suggest that re-organisation of heterochromatin is involved in altered structural integrity of the nuclear periphery during senescence, and that this can lead to activation of cytoplasmic nucleic acid sensing, NF-κB signalling, and activation of the SASP.
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- Chromosomes and Gene Expression
- Evolutionary Biology
Gene regulation is essential for life and controlled by regulatory DNA. Mutations can modify the activity of regulatory DNA, and also create new regulatory DNA, a process called regulatory emergence. Non-regulatory and regulatory DNA contain motifs to which transcription factors may bind. In prokaryotes, gene expression requires a stretch of DNA called a promoter, which contains two motifs called –10 and –35 boxes. However, these motifs may occur in both promoters and non-promoter DNA in multiple copies. They have been implicated in some studies to improve promoter activity, and in others to repress it. Here, we ask whether the presence of such motifs in different genetic sequences influences promoter evolution and emergence. To understand whether and how promoter motifs influence promoter emergence and evolution, we start from 50 ‘promoter islands’, DNA sequences enriched with –10 and –35 boxes. We mutagenize these starting ‘parent’ sequences, and measure gene expression driven by 240,000 of the resulting mutants. We find that the probability that mutations create an active promoter varies more than 200-fold, and is not correlated with the number of promoter motifs. For parent sequences without promoter activity, mutations created over 1500 new –10 and –35 boxes at unique positions in the library, but only ~0.3% of these resulted in de-novo promoter activity. Only ~13% of all –10 and –35 boxes contribute to de-novo promoter activity. For parent sequences with promoter activity, mutations created new –10 and –35 boxes in 11 specific positions that partially overlap with preexisting ones to modulate expression. We also find that –10 and –35 boxes do not repress promoter activity. Overall, our work demonstrates how promoter motifs influence promoter emergence and evolution. It has implications for predicting and understanding regulatory evolution, de novo genes, and phenotypic evolution.