Enhancers reside in a unique epigenetic environment during early development

During development and differentiation, enhancers, and not promoters are most dynamic in their DNA methylation status. However, the causal relationship between enhancer activity and methylation is not clear. Here, we describe that during early zebrafish development, enhancer activity has little influence on DNA methylation, and that hypo-methylation is a unique feature of primed enhancers, whereas active enhancers are hyper-methylated. Hypo-methylated enhancers (hypo-enhancers) are enriched close to important transcription factors (TFs) that act later in development, are specifically de-methylated before the maternal-zygotic transition (MZT), and reside in a unique epigenetic environment. Finally, we demonstrate that hypo-enhancers are functionally active and that they are physically associated with the transcriptional start site (TSS) of target-genes, irrespective of target-gene activity. In conclusion, we demonstrate that early development in zebrafish represents a time-window characterized by non-canonical DNA methylation-enhancer relationships, including global DNA hypo-methylation of inactive enhancers and DNA hyper-methylation of active enhancers. Overall design: DNA methylation profiles of 4 time points during zebrafish early development.