Study of 5mC-DNA, m6A-RNA and RNA-expression in HeLa and EB [5mC in HeLa]

Epigenetic marking of the genome via modifications of DNA and histones is central to the regulation of gene expression. DNA methylation is a well-characterized modification involved in regulating chromatin states. RNA also undergoes modifications. The most prevalent internal modification of mRNA is N6-methyladenosine (m6A) installed co-transcriptionally by the METTL3-METTL14 methyltransferases. This process is an essential post-transcriptional mechanism of gene regulation. Recent evidence links m6A function to histone modifications. Here we reveal a direct connection between m6A and DNA methylation. We find that the DNA methyltransferase DNMT1 interacts both in vitro and in vivo with METTL14 within the METTL3-METTL14 complex. Using genome- and transcriptome-wide mapping, we show that methylated cytosine (5mC) tends to occur in DNA near positions corresponding to m6A sites in mRNAs, especially in gene bodies, this being associated with increased gene expression. This tight co-occurrence of 5mC and m6A appears to be favoured by a high m6A density. METTL3-depleted cells display strong intragenic DNA hypomethylation and reduced genome-wide binding of DNMT1 to DNA as evidenced by ChIP-Seq, this leading to reduced expression of the corresponding genes. We provide further mechanistic insights by showing in vitro that m6A impedes binding of DNMT1 to RNA. Accordingly, transcriptome-wide mapping of DNMT1 in vivo confirms that METTL3-mediated m6A formation interferes with DNMT1 binding to mRNA targets, causing increased gene expression. Together, our results reveal that METTL3-METTL14 favors intragenic DNA methylation by causing DNMT1 to be repelled from mRNA by m6A and targeted to gene bodies, thereby promoting transcriptional activation. They highlight a previously unrecognized layer of gene expression regulation, involving a direct mechanistic link between DNA methylation and RNA modification. Total genomic DNA from HeLa cells wild type and knocked out for METTL3 were analyzed for 5mC changes using Infinium 850k