MOTIVATION: Cleavage Under Targets and Release Using Nuclease (CUT&RUN) has rapidly gained prominence as an effective approach for mapping protein-DNA interactions, especially histone modifications, offering substantial improvements over conventional chromatin immunoprecipitation sequencing (ChIP-seq). However, the effectiveness of this technique is contingent upon accurate peak identification, necessitating the use of optimal peak calling methods tailored to the unique characteristics of CUT&RUN data. RESULTS: Here, we benchmark four prominent peak calling tools, MACS2, SEACR, GoPeaks, and LanceOtron, evaluating their performance in identifying peaks from CUT&RUN datasets. Our analysis utilizes in-house data of three histone marks (H3K4me3, H3K27ac, and H3K27me3) from mouse brain tissue, as well as samples from the 4D Nucleome database. We systematically assess these tools based on parameters such as the number of peaks called, peak length distribution, signal enrichment, and reproducibility across biological replicates. Our findings reveal substantial variability in peak calling efficacy, with each method demonstrating distinct strengths in sensitivity, precision, and applicability depending on the histone mark in question. These insights provide a comprehensive evaluation that will assist in selecting the most suitable peak caller for high-confidence identification of regions of interest in CUT&RUN experiments, ultimately enhancing the study of chromatin dynamics and transcriptional regulation. AVAILABILITY AND IMPLEMENTATION: The CUT&RUN data generated in this study have been deposited in the Gene Expression Omnibus (GEO) under the accession number GSE282809. All the 4D Nucleome datasets can be obtained from the 4D Nucleome Data Portal (https://data.4dnucleome.org/). All scripts used for data processing, figure generation, and analysis are available in the following GitHub repository: https://github.com/OroujiLab/CUTandRun_Peak_Calling/, and have also been archived on Zenodo.

The human genome is marked by several singular and combinatorial histone modifications that shape the different states of chromatin and its three-dimensional organization. Genome-wide mapping of these marks as well as histone variants and open chromatin regions is commonly carried out via profiling DNA-protein binding or via chromatin accessibility methods. After the generation of epigenomic datasets in a cell type, statistical models can be used to annotate the noncoding regions of DNA and infer the combinatorial histone marks or chromatin states (CS). These methods involve partitioning the genome and labeling individual segments based on their CS patterns. Chromatin labels enable the systematic discovery of genomic function and activity and can label the gene body, promoters or enhancers without using other genomic maps. CSs are dynamic and change under different cell conditions, such as in normal, preneoplastic or tumor cells. This review aims to explore the available computational tools that have been developed to capture CS alterations under two or more cellular conditions.

MOTIVATION: DNA methylation is a key epigenetic factor regulating gene expression. While promoter methylation has been well studied, recent publications have revealed that functionally important methylation also occurs in intergenic and distal regions, and varies across genes and tissue types. Given the growing importance of inter-platform integrative genomic analyses, there is an urgent need to develop methods to discover and characterize gene-level relationships between methylation and expression. RESULTS: We introduce a novel sequential penalized regression approach to identify methylation-expression quantitative trait loci (methyl-eQTLs), a term that we have coined to represent, for each gene and tissue type, a sparse set of CpG loci best explaining gene expression and accompanying weights indicating direction and strength of association. Using TCGA and MD Anderson colorectal cohorts to build and validate our models, we demonstrate our strategy better explains expression variability than current commonly used gene-level methylation summaries. The methyl-eQTLs identified by our approach can be used to construct gene-level methylation summaries that are maximally correlated with gene expression for use in integrative models, and produce a tissue-specific summary of which genes appear to be strongly regulated by methylation. Our results introduce an important resource to the biomedical community for integrative genomics analyses involving DNA methylation. AVAILABILITY AND IMPLEMENTATION: We produce an R Shiny app (https://rstudio-prd-c1.pmacs.upenn.edu/methyl-eQTL/) that interactively presents methyl-eQTL results for colorectal, breast and pancreatic cancer. The source R code for this work is provided in the Supplementary Material. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.