Regulatory and evolutionary impact of DNA methylation

Regulation of transcription by DNA methylation in the 5'-CpG-3' context is a widespread phenotype allowing differential expression of genetically identical cells to persist throughout development. Consequently, differences in DNA methylation between cells, tissues, populations and species could reinforce differences in gene expression. Despite a surge in studies on DNA methylation we do not know the importance of this mechanism in population differentiation and speciation. Here we investigate the regulatory and evolutionary impact of DNA methylation in five tissues of two Ficedula flycatcher species and their naturally occurring F1 hybrids. We show that the density of CpG in the promoters of genes determines the strength of the association between gene expression and DNA methylation. The impact of DNA methylation on gene expression varies among tissues with brain showing unique patterns. Differentially expressed genes between parental species are predicted by genetic- and methylation differentiation in CpG-rich promoters. Both factors fail to predict hybrid misexpression. This suggests that promoter mismethylation is not a main determinant of hybrid misexpression in these Ficedula flycatchers. Using allele-specific methylation estimates in hybrids we also determine the genome-wide contribution of cis- and trans effects in DNA methylation differentiation. These distinct mechanisms are roughly balanced in all tissues except brain, where trans differences dominate. Overall, this study provides insight on the regulatory and evolutionary impact of DNA methylation in songbirds.