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A multi-tissue epigenomic atlas links the sheep non-coding genome to domestication and complex traits

Zhu MengiD, Shiwen ZhangiD, Ziyang Zhuang, Jiayi HeiD, Fei Liu, Lingyun Wu, Xiangrong SuniD, Jingsheng LuiD, Xinyi LiiD, Hao YangiD, Mian GongiD, Xiaolong DuiD, Qiao Mei, Hao LiiD, Guoqing ZhangiD, Xiaoyun HeiD, Jianning HeiD, Xiaofei GuoiD, Sijia Nie, Xiaoxu ZhangiD, Mingzhu Shan, Jianqi Yang, Xinyue LiiD, 杨怀强, Yangshuo HuiD, , Zhangyuan PaniD

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Abstract

Non-coding regulatory variation drives complex traits, domestication, and evolutionary adaptation, yet the sheep genome lacks high-resolution functional annotation. Here we present SheepEpimap, a multi-tissue regulatory atlas harmonizing 516 CUT&Tag histone modifications, ATAC-seq, and RNA-seq datasets across 43 adult tissues in sheep. We annotated 2.93 million cis-regulatory elements, yielding 557,441 enhancer-gene pairs and 145,407 variants with allele-specific effects. By training a sequence-to-function deep-learning model, we decoded the base-pair syntax of chromatin accessibility, annotated transcription factor motif instances genome-wide, and constructed 12,210 tissue-specific gene regulatory networks (GRNs). Integrating this resource with multi-tissue expression quantitative trait loci, selection sweeps, and genome-wide association studies prioritized non-coding variants driving domestication and complex traits. Finally, cross-species analysis revealed that sequence-conserved, tissue-matched enhancers were significantly enriched in the heritability of complex traits and diseases in humans. In summary, SheepEpimap (https://genome.ucsc.edu/s/mengzhu/SheepEpimap) provides an open-access foundational ecosystem for sheep functional genomics, precision breeding, and comparative biology.

In-depth summary

Claude Opus 5.5 · xhigh effort · 8 Nov 2026 · from the full text of version 1 (26 Aug 2026) · CC BY 4.0

SheepEpimap combines 516 histone CUT&Tag, ATAC-seq and RNA-seq datasets from 43 sheep tissues to annotate 2.93 million cis-regulatory elements and 145,407 allele-specific variants for trait interpretation.

Sheep complex traits are thought to be shaped mainly by non-coding variants, yet earlier sheep epigenomic maps were limited by narrow tissue coverage, few assay types or missing network inference, and how SheepGTEx molecular QTL act mechanistically was never systematically dissected. SheepEpimap unifies chromatin states, enhancer-gene links, sequence-to-function models and allele-specific imbalance, paralleling FAANG and FarmGTEx resources for pig, chicken and cattle.

Key results

  1. ShownAn 11-state ChromHMM model built from four histone marks plus ATAC-seq divides the sheep genome into promoter, enhancer, repressed and quiescent classes. Correlating H3K27ac signal with expression links enhancers to putative target genes.

    Atlas of 344 CUT&Tag, 86 ATAC-seq and 86 RNA-seq profiles; 2,931,347 non-redundant elements (675,096 promoters, 2,004,542 enhancers, 251,709 repressed) after excluding quiescent states; 141,809 super-enhancers via ROSE; 226,734 enhancers linked to 24,145 genes (557,441 pairs; Pearson r > 0.3, q ≤ 0.05). · Overview of functional genomic data in the…; Enhancer–gene pair prediction and functional…

  2. ShownTissue-specific ChromBPNet models decoded base-resolution sequence determinants of accessibility. The resulting de novo motif atlas, which includes tissue-restricted motifs such as liver HNF4A and intestinal CDX4, was used to assemble tissue-resolved gene regulatory networks (GRNs).

    Models passing held-out Pearson r > 0.5 were retained for 38 of 43 tissues; TF-MoDISco found 118 de novo motifs (90 matched known, 28 unannotated); Fi-NeMo mapped a mean of 91,516 instances per motif; TOBIAS footprints agreed for CTCF#2; 12,210 GRNs (mean 407 per tissue) spanned 75 TFs, 1,946 enhancers, 619 promoters and 560 genes. · ChromBPNet reveals sequence syntax underlying…; Construction of a sheep gene regulatory network; ChromBPNet modeling of regulatory sequence…

  3. ShownAllele-specific analysis across six assays identified cis-acting imbalanced SNPs. Tissue-restricted enhancers carried larger allelic effects than broadly shared, more constrained enhancers, and allele-specific elements were enriched for molQTL and GWAS loci.

    Binomial tests at 13,366,709 heterozygous SNPs yielded 145,407 imbalanced SNPs (BH-adjusted P < 0.1): 56,167 RNA-seq, 48,536 ATAC-seq, 28,968 H3K4me3, 7,183 H3K27ac, 4,295 H3K4me1, 258 H3K27me3; allelic effects correlated with an independent liver dataset; ES-EnhA (21-43 tissues) showed the smallest ATAC, H3K4me1 and H3K27ac effects. · Allele-specific regulatory variants; Identification of allele-specific events

  4. ShownRegulatory annotations overlap most known monogenic and QTL records and show tissue-matched enrichment of sheep GWAS and selection signals. This nominates candidate variants such as rs426482293 in a colon enhancer at LCORL.

    25 of 38 OMIA loci (65.8%) and 3,759 of 6,937 AnimalQTLdb QTL (54.2%) overlapped regulatory elements; eQTL enriched in promoter and enQTL in enhancer states; litter-size GWAS signals enriched in uterus, ovary and oviduct enhancers; sweeps contained 6,986 candidate variants (98% noncoding, 87% overlapping or flanking tissue-specific regions). · Integrating SheepEpimap with molQTL and GWAS…; SheepEpimap prioritizes candidate regulatory…; Regulatory annotation and enrichment analysis of…; Identification and regulatory enrichment…

  5. SuggestedSheep tissue-specific enhancers lifted over to the human genome capture tissue-matched human trait heritability, and human elements conserved in both sequence and chromatin state in matched sheep tissues show nominal heritability enrichment in most trait-tissue pairs tested.

    Lifted sheep uterus, heart and brain enhancers were enriched for human multiple-birth, pulse-rate and neuroticism heritability; across 12 matched tissue pairs, 3,290,130 human CREs split into 529,432 sfCREs, 949,258 sdCREs, 1,335,701 soCREs and 475,739 ssCREs; sfCREs showed nominal (uncorrected) LDSC enrichment for 24 of 37 trait-tissue pairs (64.9%). · Translational analysis of SheepEpimap in human…; Cross-species CRE classification and…

Methods, data and reproducibility

Bulk tissues from two rams and two ewes (Small-Tailed Han) were profiled by CUT&Tag for four histone marks, ATAC-seq and RNA-seq; ChromHMM states, correlation-based enhancer-gene links, ChromBPNet models, motif-based GRNs and WASP-corrected allelic-imbalance tests were integrated with SheepGTEx molQTL, sheep GWAS, selection scans and human LDSC.

  • Newly generated CUT&Tag (H3K4me1, H3K4me3, H3K27ac, H3K27me3), ATAC-seq and RNA-seq across 43 adult tissues (most from two ewes; testis and epididymis from two rams) · sheep (Ovis aries), Small-Tailed Han breed · n = 516 datasets (344 CUT&Tag, 86 ATAC-seq, 86 RNA-seq); 43 tissues · SRA BioProject PRJNA1237432
  • SheepGTEx multi-tissue expression and enhancer molecular QTL (eQTL, enQTL) · sheep (Ovis aries) · n = 19,054,283 eQTL/enQTL
  • Sheep GWAS summary statistics for 34 complex traits (growth, body size, reproduction, wool), meta-analyzed across populations with METAL · sheep (Ovis aries) · n = 21,531 GWAS signals across 34 traits
  • OMIA monogenic variants and Animal QTLdb trait-associated QTL records · sheep (Ovis aries) · n = 38 OMIA variants; 6,937 QTL
  • Population genotypes from SheepGTEx and Li et al. for four selection contrasts (F_ST on 21,542,313 common SNPs; XP-CLR and pi ratio for wild vs domestic) · sheep (Ovis aries) and wild Ovis orientalis · n = CEA 1,104; EUR 710; ancient CEA 21, EUR 31; domestic 738; wild 72
  • ENCODE and Roadmap Epigenomics human epigenomes re-annotated with the same 11-state ChromHMM model for sheep-human CRE comparison · human · n = 111 epigenomes; 12 matched tissue pairs
  • Human GWAS summary statistics for LDSC partitioned heritability, using 1000 Genomes Phase 3 European reference and the baselineLD model · human · n = 37 traits
  • Public sheep Hi-C (Yan et al.) for TAD calling and EBI WGBS for muscle, ovary, skin and liver · sheep (Ovis aries)
Methods & tools
ChromHMM (v1.26) 11-state chromatin-state model, ChromImpute (v1.0.5) for track imputation, ChromBPNet (v1.0) sequence-to-function models, TF-MoDISco and Fi-NeMo motif discovery and instance calling, TOBIAS footprinting, ROSE super-enhancer calling, HOMER (v4.11) and MEME FIMO (v5.5.9) motif analysis, GATK (v4.6.2.0), Beagle (v5.4) and WASP (v0.16.3) for allele-specific analysis, MACS2 (v2.2.9.1), STAR (v2.7.11a), Bowtie2 (v2.5.3), METAL, VCFtools (v0.1.16) F_ST, XP-CLR and GAT for GWAS and selection analyses, LDSC (v1.0.0) partitioned heritability with liftOver, GREAT and clusterProfiler (v4.4.1) enrichment
Software & environment
Trim Galore v0.6.5, STAR v2.7.11a, Bowtie2 v2.5.3, SAMtools v1.18, MACS2 v2.2.9.1, ChromHMM v1.26, ChromImpute v1.0.5, ChromBPNet v1.0, GATK v4.6.2.0, WASP v0.16.3, LDSC v1.0.0; reference genome ARS-UI_Ramb_v2.0 (GCF_016772045.1)
Reproducibility gaps
Trained ChromBPNet model weights and the meta-analyzed sheep GWAS summary statistics are not stated as deposited; Hi-C, WGBS, SheepGTEx QTL and human GWAS inputs must be retrieved from cited sources.
Limitations (5)
  • Built from bulk adult tissue of few animals, the atlas cannot resolve cell-type heterogeneity, developmental changes or population-level regulatory variation. · stated by the authors
  • Enhancer-gene links, GRNs and ChromBPNet allelic effects rest on correlation, chromatin marks and sequence-model predictions, so they are prioritized candidates rather than causal evidence; CRISPR, STARR-seq or reporter assays are needed. · stated by the authors
  • Sheep GWAS datasets have limited power, so enrichment relied on suggestive rather than genome-wide significant loci; larger GWAS, fine-mapping and molecular QTL maps are needed to link regulatory variants to traits. · stated by the authors
  • Cross-species LDSC enrichment P values were not corrected for multiple testing, so some of the 24 of 37 sfCRE trait-tissue pairs called significant may be false positives. · noted in this summary
  • Allele-specific calls use a lenient BH-FDR < 0.10 in a few animals, and external replication was shown only for liver, so reproducibility in other tissues and assays is untested. · noted in this summary

Read this if you work on livestock functional genomics, need sheep enhancer, chromatin-state or ChromBPNet motif annotations, or want to interpret sheep GWAS, molQTL or selection signals in a tissue-resolved regulatory context.

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