Integrated single-cell sequencing reveals principles of epigenetic regulation of human gastrulation and germ cell development in a 3D organoid model

The emergence of different cell types and the role of the epigenome in regulating transcription is a key yet understudied event during human gastrulation. Investigating these questions remain infeasible due to the lack of availability of embryos at these stages of development. Further, human gastrulation is marked by dynamic changes in cell states that are difficult to isolate at high purity, thereby making it challenging to map how epigenetic reprogramming impacts gene expression and cellular phenotypes. To overcome these limitations, we describe scMAT-seq, a high-throughput one-pot single-cell multiomics technology to simultaneously quantify DNA methylation, DNA accessibility and the transcriptome from the same cell. Applying scMAT-seq to 3D human gastruloids, we characterized the epigenetic landscape of major cell types corresponding to the germ layers and primordial germ cell-like cells (hPGCLC). As the identity of the progenitors that give rise to human PGCLCs remain unclear, we used this system to discover that the progenitors emerge from epiblast cells and show transient characteristics of both amniotic- and mesoderm-like cells, before getting specified towards hPGCLCs. Finally, as cells differentiate along different lineages during gastrulation, we surprisingly find that while changes in DNA accessibility are tightly correlated to both upregulated and downregulated genes, reorganization of gene body DNA methylation is strongly related to only genes that get downregulated, with genes that turn on displaying a lineage trajectory-dependent correlation with DNA methylation. Collectively, these results demonstrate that scMAT-seq is a high-throughput and sensitive approach to elucidate epigenetic regulation of gene expression in complex systems such as human gastrulation that are marked by rapidly transitioning cell states.