To better understand the regulation of methylation patterns during its most dynamic phase, we generated genome-scale maps of DNA methylation in both gametes and through the complete pre-implantation timeline. We find that methylation contributed by sperm to the zygote is most dramatically altered in retro-elements of specific families, while other elements remain more protected and retain higher methylation levels throughout development (Supplementary Fig. 11
). The methylation status of the oocyte is a strong predictor of levels in the zygote, and regions that are already hypomethylated in the oocyte could explain much of the disparities between the early embryo and sperm. Possibly, the mechanism and targets of DNA demethylation during female gametogenesis could be similar to those at work during fertilization34
. Regardless, the embryonic pattern more closely resembles that of the oocyte until the later stages of pre-implantation, where DNA methylation is further decreased.
In addition to classical ICRs, which exhibit parent-of-origin specific methylation maintained through adulthood, a substantial number of CpG island promoters are specifically hypermethylated in the oocyte, in agreement with a recent study25
. Surprisingly, these regions retain intermediate values indicating differential allelic methylation before gradually decreasing through ICM specification and gastrulation, where somatic methylation patterns are re-established (Supplementary Fig. 11
It remains to be investigated whether the diverse targets that exhibit low methylation levels during embryogenesis are the consequence of a single regulatory principle. LINE and LTR activity in the early embryo is associated with some of the earliest transcriptional events during zygotic genome activation. Targeted depletion by antisense oligonucleotides of the L1Md_T class as well as certain LTRs have demonstrated a general requirement for retrotransposon transcription for progression through cleavage48,49
. These observations may also support data suggesting the elongation factor/histone acetyltransferase ELP3 is a component of the DNA demethylation machinery and could explain a tight relationship between demethylation and transcription-associated complexes50
It is likely that current interest in hmC will spur technical improvements that will permit quantitative dissection of mC and hmC patterns, which will help answer remaining questions regarding Tet3’s universal necessity for conversion to unmethylated cytosines, as well as the effect hmC may have on Dnmt-mediated inheritance32
. Tet3’s global conversion to hmC of the paternal genome does not appear to lead to equivalently dramatic demethylation based on the retention of bisulfite-detected signal. The feature-specific dynamics of DNA methylation at fertilization suggest that Tet3 and hmC may be required for targeted demethylation, as well as for driving a gradual hypomethylation over cleavage9,10
. Further experiments will be required to characterize this division-dependent demethylation in more detail, and expand it to regions with lower GC content that are under-represented in RRBS. Importantly, other mechanisms must retain heritable methylation information because many targets display relative epigenetic stability from zygote onward and many of these features exhibit embryogenesis-specific methylation patterns.
In summary, our genome-scale single base resolution data provide improved understanding of the relationship and general regions exhibiting DNA demethylation at fertilization. This expands earlier models derived from immunohistochemistry-based observations and begins to address remaining open questions, setting the stage for future epigenetic studies in early mouse development.