mA modulates haematopoietic stem and progenitor cell specification.

N-methyladenosine (mA) has been identified as the most abundant modification on eukaryote messenger RNA (mRNA). Although the rapid development of high-throughput sequencing technologies has enabled insight into the biological functions of mA modification, the function of mA during vertebrate embryogenesis remains poorly understood. Here we show that mA determines cell fate during the endothelial-to-haematopoietic transition (EHT) to specify the earliest haematopoietic stem/progenitor cells (HSPCs) during zebrafish embryogenesis. mA-specific methylated RNA immunoprecipitation combined with high-throughput sequencing (MeRIP-seq) and mA individual-nucleotide-resolution cross-linking and immunoprecipitation with sequencing (miCLIP-seq) analyses reveal conserved features on zebrafish mA methylome and preferential distribution of mA peaks near the stop codon with a consensus RRACH motif. In mettl3-deficient embryos, levels of mA are significantly decreased and emergence of HSPCs is blocked. Mechanistically, we identify that the delayed YTHDF2-mediated mRNA decay of the arterial endothelial genes notch1a and rhoca contributes to this deleterious effect. The continuous activation of Notch signalling in arterial endothelial cells of mettl3-deficient embryos blocks EHT, thereby repressing the generation of the earliest HSPCs. Furthermore, knockdown of Mettl3 in mice confers a similar phenotype. Collectively, our findings demonstrate the critical function of mA modification in the fate determination of HSPCs during vertebrate embryogenesis.