Cited by 567Genomics and Chromatin DynamicsEpigenetics and DNA MethylationChromatin Remodeling and Cancer

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Peak calling by Sparse Enrichment Analysis for CUT&RUN chromatin profiling

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Meers, Michael P., Tenenbaum, Dan, Henikoff, Steven·2019·Epigenetics & Chromatin

DOI 10.1186/s13072-019-0287-4PMC PMC6624997

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Abstract

BACKGROUND: CUT&RUN is an efficient epigenome profiling method that identifies sites of DNA binding protein enrichment genome-wide with high signal to noise and low sequencing requirements. Currently, the analysis of CUT&RUN data is complicated by its exceptionally low background, which renders programs designed for analysis of ChIP-seq data vulnerable to oversensitivity in identifying sites of protein binding. RESULTS: Here we introduce Sparse Enrichment Analysis for CUT&RUN (SEACR), an analysis strategy that uses the global distribution of background signal to calibrate a simple threshold for peak calling. SEACR discriminates between true and false-positive peaks with near-perfect specificity from "gold standard" CUT&RUN datasets and efficiently identifies enriched regions for several different protein targets. We also introduce a web server ( http://seacr.fredhutch.org ) for plug-and-play analysis with SEACR that facilitates maximum accessibility across users of all skill levels. CONCLUSIONS: SEACR is a highly selective peak caller that definitively validates the accuracy of CUT&RUN for datasets with known true negatives. Its ease of use and performance in comparison with existing peak calling strategies make it an ideal choice for analyzing CUT&RUN data.

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Fig. 1 SEACR enforces peak calling specificity across a range of read depths. a Schematic of SEACR methodology. Contiguous signal blocks (top) are identified and plotted by the percentage of blocks exceeding a total signal threshold (right), and an optimal threshold is empirically identified and used for filtering (bottom). b Plot of area under the precision–recall curve (AUPR) for peaks called from H3K4me3 CUT&RUN data by SEACR, MACS2, MACS2 with local lambda inactivated (MACS2 llocal), and HOMER that were compared to a stringent test set of H3K4me3 peaks called from ENCODE ChIP-seq data. Read subsampling levels are indicated on the X-axisCC BY · PubMed Central

Fig. 2 SEACR minimizes true negative detection from a gold standard dataset. a Schematic of SEACR threshold selection. A target/total signal block curve (right) is calculated, and the stringent and relaxed thresholds are set based on the knee and the peak of the curve. b Peaks were called from Sox2 or FoxA2 experiments carried out in either hESCs or Endoderm cells using SEACR in stringent or relaxed mode; MACS2; or HOMERCC BY · PubMed Central

Fig. 3 SEACR thresholding provides an improved specificity/sensitivity trade-off. a Precision–recall curves derived from a comparison of peaks called from subsampled H3K4me2 CUT&RUN to ENCODE peaks called from H3K4me2 ChIP-seq data. Colors denote peak calling method, size of points denotes read subsampling level, and dotted black line denotes 85% precision in peak calling. b F1 score for each point on the plots in ACC BY · PubMed Central

Fig. 4 SEACR identifies coherent broad domains. a CUT&RUN domains identified by SEACR are broader than those found by MACS2 or HOMER. b A representative region of a H3K27me3 profile is shown. SEACR domains are relatively coherent compared with HOMER and MACS2 domains, which are fragmented into mixtures of wide and narrow segmentsCC BY · PubMed Central

Fig. 5 SEACR exhibits competitive run time and memory usage. Comparison of run time (a), average disk read memory (b), and average disk write memory (c) recorded for SEACR, MACS2, and HOMER peak calling from H3K27me3 data at read depths indicated. Error bars indicate standard deviation from the mean of 10 trialsCC BY · PubMed Central

Fig. 6 A web server for SEACR analysis. a The SEACR web server home page (https://seacr.fredhutch.org) contains browse functions for bedgraph input files and toggles for normalization and stringent/relaxed mode options. b The SEACR web server output page features a progress window with regular updates, a link for downloadable results in BED format, and the empirical false discovery rate (FDR) calculated by quantifying the percentage of control signal blocks remaining out of the total above the thresholdCC BY · PubMed Central

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Authors: Meers, M. P., Tenenbaum, D., & Henikoff, S.
Affiliations: Fred Hutch Cancer Center (US)
Howard Hughes Medical Institute (US)
Corresponding author: Henikoff, Steven
ORCID: Meers, Michael P.
Henikoff, Steven
Venue: Epigenetics & Chromatin
Publisher: Springer Science and Business Media LLC
Published: 2019-07-12
Volume: 12
Issue: 1
Pages: 42-42
ISSN: 1756-8935 (ISSN-L)
DOI: 10.1186/s13072-019-0287-4
PMC: PMC6624997
OpenAlex: W2958184371
Cited by: 556
References: 21
Citations by year: 2026: 86 · 2025: 123 · 2024: 126 · 2023: 95 · 2022: 65 · 2021: 44 · 2020: 16
FWCI: 10.34
Type: journal article
Language: en
Open access: gold
doi.org
Licence: cc-by
creativecommons.org/licenses/by/4.0/ (tdm)
creativecommons.org/licenses/by/4.0/ (vor)
Locations: Epigenetics & Chromatin (publishedVersion, cc-by)
PubMed Central (publishedVersion, cc-by)
PubMed (publishedVersion)
DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
Funders: Howard Hughes Medical Institute (Henikoff)
National Human Genome Research Institute (1R01HG010492)
Topics: Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Genomics and Chromatin Dynamics
Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Epigenetics and DNA Methylation
Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Chromatin Remodeling and Cancer
Keywords: peak calling, CUT&RUN, chromatin profiling, protein-DNA binding sites, epigenetic profiling, web server
MeSH terms: Algorithms, Animals, Binding Sites, Chromatin, DNA-Binding Proteins, Humans, Protein Binding, Genome, Sequence Analysis, DNA, Epigenesis, Genetic, Chromatin Immunoprecipitation, Epigenomics

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Peak calling by Sparse Enrichment Analysis for CUT&RUN chromatin profiling

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