Comparative analysis reveals epigenomic evolution related to species traits and genomic imprinting in mammals

Abstract

DNA methylation is an epigenetic modification that plays a crucial role in various regulatory processes, including gene expression regulation, transposable element repression, and genomic imprinting. However, most studies on DNA methylation have been conducted in humans and other model species, whereas the dynamics of DNA methylation across mammals remain poorly explored, limiting our understanding of epigenomic evolution in mammals and the evolutionary impacts of conserved and lineage-specific DNA methylation. Here, we generated and gathered comparative epigenomic data from 13 mammalian species, including two marsupial species, to demonstrate that DNA methylation plays critical roles in several aspects of gene evolution and species trait evolution. We found that the species-specific DNA methylation of promoters and noncoding elements correlates with species-specific traits such as body patterning, indicating that DNA methylation might help establish or maintain interspecies differences in gene regulation that shape phenotypes. For a broader view, we investigated the evolutionary histories of 88 known imprinting control regions across mammals to identify their evolutionary origins. By analyzing the features of known and newly identified potential imprints in all studied mammals, we found that genomic imprinting may function in embryonic development through the binding of specific transcription factors. Our findings show that DNA methylation and the complex interaction between the genome and epigenome have a significant impact on mammalian evolution, suggesting that evolutionary epigenomics should be incorporated to develop a unified evolutionary theory.

Graphical abstract

Figure 1

Interspecies conservation of DNA methylation among various genomic features (A) Each violin plot depicts the distribution of the DNA methylation levels in the genomic regions of gene bodies, exons, introns, intergenic regions, and TEs in the species that we analyzed, as shown in the phylogenetic tree. (B) This figure shows the methylation levels of regions 10 kb upstream and 10 kb downstream of the TSS of genes in each species calculated using 100 bp bins, indicating that the regions around the TSS have the lowest methylation levels. (C) This figure shows the average methylation levels of each genomic feature in each species.

Figure 2

Epigenomic evolution of promoter methylation in synchrony with the molecular evolution of genes (A) The boxplot shows the distribution of promoter methylation divergence values for ∼660,000 orthologous gene pairs with dS < 3, divided into six bins by dS values. The regression line with the 95% confidence interval demonstrates a significant correlation (p < 2.2 × 10⁻¹⁶) between promoter methylation divergence and dS of all gene pairs, indicating that gene pairs with larger dS values and higher evolutionary distances tended to differ in promoter methylation. (B) The boxplot shows the distribution of mean promoter methylation levels for more than 10,000 genes, divided into five bins by dN/dS values. The regression line with the 95% confidence interval demonstrates a significant correlation (p < 2.2 × 10⁻¹⁶) between mean promoter methylation and dN/dS of all these genes, indicating that genes with lower evolutionary rates tend to have lower mean promoter methylation levels.

Figure 3

Promoter methylation patterns of orthologous genes are associated with important developmental processes and species traits (A) This heatmap shows the methylation levels of four groups of gene promoters: consistently hypomethylated, lineage-specifically hypermethylated, lineage-specifically hypomethylated, and consistently hypermethylated. (B) The functional analysis of consistently hypomethylated gene promoters reveals the enrichment of developmental processes, including cell development, tissue and system development, embryo development, and organism morphogenesis. (C) Promoters and noncoding sequences with lineage-specific altered methylation patterns exhibit enriched GO annotations (selected from all significant results) associated with species traits. The biological processes in black letters are enriched in the promoters, whereas the biological processes in blue letters are enriched in both the promoters and noncoding sequences.

Figure 4

Evolutionary origins of documented human and mouse ICRs Arrows indicate that ICRs may arise during the evolution of the branch. Somatic ICRs are marked by an asterisk (“∗”), and the rest are germline ICRs. Two somatic ICRs (MEG3 and MEG8) are located within the imprinting cluster of the germline ICR DLK1-DIO3, and two others (NESPAS-GNASXL, GNAS1A) are located within the cluster of the germline ICR GNAS. The divergence time of each node was extracted from TimeTree.

Figure 5

Genomic distribution, functional enrichment, and motif enrichment of AMRs (A) The pie chart indicates the proportions of AMRs that overlapped with genes (including protein-coding genes, lncRNA genes, and both) and intergenic regions. (B) The distances of the AMRs to the nearest transcription start sites of genes (black line) were significantly closer compared with those of random regions of similar length (gray line). (C) Gene functions enriched in the commonly shared AMRs were mostly associated with embryonic development. (D) Motifs enriched in the commonly shared AMRs were associated with development and gene expression regulation.