Specification of human germ cell fate with enhanced progression capability

Human primordial germ cells (hPGCs) are the embryonic precursors of sperm and eggs. While the exact temporal window of hPGC specification and the cellular identity of their precursors remains unclear, few studies on ex vivo and in vitro cultured human embryos reported plausible hPGCs on embryonic day (E) 12-13, and in a E16-17 gastrulating embryo. In vitro, hPGC-like cells (hPGCLCs) can be specified from intermediary pluripotent stage or peri-gastrulation precursors. Nevertheless, it remains unclear the extent of the spectrum of precursors for hPGCLC specification and how these different cellular identities impact hPGCLC development. Here we show that capacitating and resetting precursors respond to BMP and induce resetting hPGCLCs (rhPGCLCs). Strikingly, rhPGCLC in the human hindgut organoid co-cultures show progression at a pace reminiscent of in vivo timing, demonstrating their higher potential for progression compared to those derived from peri-gastrulation precursors. Notably, EOMES and TBXT act synergistically during rhPGCLC specification, while only EOMES is necessary for peri-gastrulation hPGCLC specification. Our results emphasize the notion that the function of a cell identity can be largely influenced by features of their precursors rather than their convergent identity. Importantly, our study provides the foundation for refined and efficient in vitro models of human gametogenesis, which may contribute to design new therapeutic approaches for human infertility. Overall design: RNA-seq of hESC-derived cell types (2-4 biological replicates for each cell type)

人类原始生殖细胞（human primordial germ cells, hPGCs）是精子与卵子的胚胎前体。尽管hPGCs的精确特化时间窗口及其前体细胞的细胞身份仍不明确，目前仅有少数针对离体（ex vivo）与体外（in vitro）培养人类胚胎的研究，分别在胚胎第12-13天（E12-13）以及16-17天的原肠胚形成胚胎中报道了疑似的hPGCs。在体外环境中，人类原始生殖细胞样细胞（human primordial germ cell-like cells, hPGCLCs）可由中间多能阶段或原肠运动前期前体细胞特化而来。然而，hPGCLC特化的前体细胞谱系范围究竟有多大，以及这些不同的细胞身份如何影响hPGCLC的发育，目前仍不清楚。本研究发现，活化型与重置型前体细胞可响应骨形态发生蛋白（bone morphogenetic protein, BMP），并诱导产生重置型人类原始生殖细胞样细胞（resetting hPGCLCs, rhPGCLCs）。值得注意的是，在人类后肠类器官共培养体系中的rhPGCLCs，其发育进程与体内时序高度一致，这表明相较于源自原肠运动前期前体细胞的hPGCLCs，此类细胞具备更强的发育潜能。尤为关键的是，EOMES与TBXT在rhPGCLC特化过程中具有协同作用，而仅EOMES对于原肠运动前期hPGCLC特化是必需的。本研究结果印证了如下观点：细胞身份的功能在很大程度上受其前体细胞的特征影响，而非其趋同后的细胞身份。重要的是，本研究为构建更精准高效的人类配子发生体外模型奠定了基础，有望为人类不孕症的新型治疗方案研发提供思路。实验设计：对人类胚胎干细胞（human embryonic stem cells, hESC）来源的各类细胞进行RNA测序（RNA-seq），每种细胞类型设置2-4次生物学重复。