Inheritance and reprogramming of non-CG methylation at satellite DNA repeats

In vertebrates, DNA methylation predominantly occurs at CG dinucleotides however, widespread non-CG methylation (mCH) has been reported in mammalian embryonic stem cells and in the brain. In mammals, mCH is found at CAC trinucleotides in the nervous system, where it is associated with transcriptional repression, and at CAG trinucleotides in embryonic stem cells, where it positively correlates with transcription. Moreover, CAC methylation appears to be a conserved feature of adult vertebrate brains. Unlike any of those methylation signatures, here we describe a novel form of mCH that occurs in the TGCT context within zebrafish mosaic satellite repeats. TGCT methylation is inherited from both male and female gametes, remodelled during mid-blastula transition, and re-established during gastrulation in all embryonic layers. Moreover, we identify DNA methyltransferase 3ba (Dnmt3ba) as the primary enzyme responsible for the deposition of this mCH mark. Finally, we observe that TGCT-methylated repeats are specifically associated with H3K9me3-marked heterochromatin suggestive of a functional interplay between these two gene-regulatory marks. Altogether, this work provides insight into a novel form of vertebrate mCH and highlights the substrate diversity of vertebrate DNA methyltransferases. Overall design: Enzymatic methylation sequencing (EM-seq): 24hpf (n=10 embryos / sample, n=2 samples); Reduced Representation Bisulfite sequencing (RRBS): 48hpf (n=10 embryos / sample, n=2 samples / condition); Whole Genome Bisulfite Sequencing (WGBS): 72h larvae (n=10 embryos / sample, n=2 samples / condition) and adult tissues (n=2 samples).