A developmental coordinate of the spectrum of pluripotency among mice, monkeys, and humans

The epiblast (EPI) is the origin of all somatic and germ cells in mammals, and of the spectrum of pluripotent stem cells (PSCs) in vitro. To explore the ontogeny of human/primate pluripotency, we performed comprehensive single-cell RNA sequencing for pre- and post-implantation EPI development in cynomolgus monkeys. Here we show that after specification in the blastocysts [embryonic day (E)7], cyEPI undergoes major transcriptome changes upon implantation. Thereafter, cyEPI, while generating gastrulating cells (~E13), maintains its transcriptome relatively stably over a week, retaining a unique set of pluripotency genes while acquiring properties for "neuron differentiation." h/cyPSCs show the highest similarity to post-implantation late cyEPI (~E17), which, despite co-existing with gastrulating cells, bears characteristics of pre-gastrulating mouse EPI (E5.5) and epiblast-like cells (EpiLCs) in vitro. These findings not only reveal divergence/coherence of EPI development, but also identify a developmental coordinate of the spectrum of pluripotency among key species, providing a basis for better regulation of human pluripotency in vitro. Single cell transcriptome analysis of cynomolgus monkey embryo E6-17 and mouse embryo E4.5 - E6.5 as well as of cynomogus monkey embryonic stem cells (ESCs) and mouse ESC, epiblast like cells (EpiLCs), using SC3-seq technology.上胚层（epiblast，EPI）是哺乳动物所有体细胞、生殖细胞以及体外各类多能干细胞（pluripotent stem cells，PSCs）的起源。为探究人类/灵长类多能性的个体发育过程，我们对食蟹猴植入前及植入后的上胚层发育阶段开展了全面的单细胞RNA测序。本研究显示，在囊胚中完成特化后（胚胎日E7），食蟹猴上胚层（cyEPI）在植入后会发生显著的转录组变化。此后，cyEPI在产生原肠胚形成细胞（约E13）的同时，在长达一周的时间内维持相对稳定的转录组特征，既保留了独特的多能性基因谱，又逐步获得了“神经元分化”相关特性。人类/食蟹猴多能干细胞（h/cyPSCs）与植入后晚期的cyEPI（约E17）相似度最高；尽管该阶段的cyEPI与原肠胚形成细胞共存，但其特征更接近于原肠胚形成前的小鼠上胚层（E5.5）以及体外培养的上胚层样细胞（epiblast-like cells，EpiLCs）。本研究结果不仅揭示了上胚层发育的物种差异与保守性，还明确了关键物种间多能干细胞谱系的发育坐标，为体外调控人类多能性提供了理论基础。本研究采用SC3-seq技术，对食蟹猴胚胎E6-E17阶段、小鼠胚胎E4.5-E6.5阶段的细胞，以及食蟹猴胚胎干细胞（embryonic stem cells，ESCs）、小鼠胚胎干细胞、上胚层样细胞（EpiLCs）开展了单细胞转录组分析。