Epigenetic regulation of cell fate decisions in mouse gastrulation embryo at single-cell resolution

In mammals, implantation marks the initiation of cellular differentiation and exit from pluripotency. Alongside the apparent morphological changes of the gastrulating embryo, cells undergo lineage-commitment and early fate-choices shaped by complex layers of regulation. Specifically, various epigenetic regulators, such as the TET and DNMT proteins are critical for the progression of the embryo past the gastrulation phase, demonstrating the essential role of DNA methylation in early development. However, a comprehensive understanding of how DNA methylation is involved in defining cell states in vivo, through regulation of target genes, remains a formidable task. To address this, we characterized the developing embryo at unparalleled temporal resolution, by adopting a strategy of single-cell RNA sequencing (scRNA-seq) in single embryos. In this manner, we profiled the transcriptomes of >20,000 cells from >100 individual embryos from embryonic days 6.5-8.0. The resulting map provides a virtual “transcriptome-continuum” for gastrulating embryos, allowing us to compute a “molecular-age stamp”, correlated developmental milestones. To study the functional roles of DNA methylation during gastrulation, we have generated isogenic mouse embryonic stem cell lines, harboring knockouts of key regulators of DNA methylation. Analyzing chimeric embryos derived from isogenic clones, serves to separate cell-autonomous and non-autonomous effects of DNA methylation on cell states, within the context of the normally developing embryo. Our combined approaches will allow elucidating the functional roles of DNA methylation during early cell-fate changes, critically evaluating and substantiating casual relationships between DNA methylation changes and gene expression.