Human Primordial Germ Cells are Specified from Lineage Primed Progenitors

In vitro gametogenesis is the process of making germline cells from human pluripotent stem cells. The foundation of this model is the quality of the first progenitors called primordial germ cells (PGCs), which in vivo are specified during the peri-implantation window of human development. Here, we show using human embryo attachment culture that hPGC specification begins at day 12 post fertilization. Using single cell RNA-sequencing of hPGC-like cells (hPGCLCs) differentiated from pluripotent stem cells we discovered that hPGCLC specification involves resetting pluripotency towards a transitional state with shared characteristics between naïve and primed pluripotency followed by differentiation into lineage primed TFAP2A+ progenitors. Applying the germline trajectory to TFAP2C mutants reveals that TFAP2C functions in the TFAP2A+ progenitors upstream of PRDM1 to regulate the expression of SOX17. This serves to protect hPGCLCs from crossing the Weismann’s barrier to adopt somatic cell fates and therefore is an essential mechanism for successfully initiating in vitro gametogenesis.