Functionally Annotating Regulatory Elements in the Equine Genome Using Histone Mark ChIP-Seq

Abstract

One of the primary aims of the Functional Annotation of ANimal Genomes (FAANG) initiative is to characterize tissue-specific regulation within animal genomes. To this end, we used chromatin immunoprecipitation followed by sequencing (ChIP-Seq) to map four histone modifications (H3K4me1, H3K4me3, H3K27ac, and H3K27me3) in eight prioritized tissues collected as part of the FAANG equine biobank from two thoroughbred mares. Data were generated according to optimized experimental parameters developed during quality control testing. To ensure that we obtained sufficient ChIP and successful peak-calling, data and peak-calls were assessed using six quality metrics, replicate comparisons, and site-specific evaluations. Tissue specificity was explored by identifying binding motifs within unique active regions, and motifs were further characterized by gene ontology (GO) and protein-protein interaction analyses. The histone marks identified in this study represent some of the first resources for tissue-specific regulation within the equine genome. As such, these publicly available annotation data can be used to advance equine studies investigating health, performance, reproduction, and other traits of economic interest in the horse.

Genome-Wide Histone Modifications and CTCF Enrichment Predict Gene Expression in Sheep Macrophages

“…Heterochromatin and silencer sequences were shown to be depleted for GC content ( Glass and Natoli, 2016 ) like our H3K27me3-enriched regions (see Table 2 for GC content summary). We found a clear bimodal distribution of active promoters and enhancers around the TSS ( Figure 3 ) as has been reported by others for a variety of tissue types ( Kingsley et al, 2020 ). At the TSS there was a slight depression in signal reflecting this nucleosome depletion that would allow for the positioning of the initiation complex and RNA polymerase along the chromatin.…”

“…Several groups estimated that over 90% of causal mutations that explain phenotypic variation laid outside of genes within regulatory elements ( Hindorff et al, 2009 ; Maurano et al, 2012 ; Albert and Kruglyak, 2015 ). Currently, little is known regarding in vivo tissue annotation of regulatory elements in livestock species ( Villar et al, 2015 ; Zhao et al, 2015 ; Wang et al, 2017 , 2018 ; Naval-Sanchez et al, 2018 ; Nguyen et al, 2018 ; Fang et al, 2019 ; Hall et al, 2020 ; Kingsley et al, 2020 ). Therefore, the FAANG consortium recognized this need and formed a global network of researchers for epigenetic discovery in food animal species ( Andersson et al, 2015 ; Tuggle et al, 2016 ; Giuffra and Tuggle, 2019 ).…”

Whole genome sequencing identified a 16 kilobase deletion on ECA13 associated with distichiasis in Friesian horses

“…As this locus on ECA13 contains no annotated protein-coding genes and no functional data currently exists for eyelid tissue or Meibomian glands, the equine data collected for the Functional Annotation of Animal Genomes (FAANG) project, including RNA-seq [ 26 , 27 ] and ChIP-seq data [ 28 ] from two clinically healthy Thoroughbred mares, were evaluated to develop additional hypotheses on the functional mechanisms of this variant. To investigate if there was tissue-specific variation in gene expression in clinically normal horses, the three genes near the deletion ( FAM20C , PRKAR1B and PDGFA ) were assessed in the RNA-seq data.…”

“…As the deletion identified is located in an intergenic region, computational analyses were performed to assess the potential regulatory role of this region as the cause of distichiasis. The annotated locations of FAM20C , PDGFA and PRKAR1B from the FAANG RNA-seq data were visualized using IGV [ 28 , 29 ]. Publicly available equine FAANG ChIP-Seq data (H3K4me1, H3K4me, H3K27ac, and H3K27me3) from eight tissues (adipose, brain, heart, lamina, liver, lung, ovary, skeletal muscle) were assessed to investigate potential regulatory regions [ 30 ].…”

Pig genome functional annotation enhances biological interpretations of complex traits and comparative epigenomics

“…Following ENCODE and Roadmap Epigenomics projects 8 , the Functional Annotation of Animal Genomes (FAANG) initiative 24 , although still in its infancy, has made great progress towards annotating functional elements in many tissues across multiple domestic species including pigs [25][26][27][28][29][30] . Here, we present 95 new genome-wide sequencing datasets from six gut-associated porcine tissues and integrate them with 128 previously published FAANG datasets from eight biologically distinct tissues.…”