Mammalian embryo comparison identifies novel pluripotency genes associated with the naive or primed state

During early mammalian development transient pools of pluripotent cells emerge that can be immortalised upon stem cell derivation. The pluripotent state, "naive" or "primed", depends on the embryonic stage and derivation conditions used. Here we analyse the temporal gene expression patterns of mouse, cattle and porcine embryos at stages that harbour different types of pluripotent cells. We document conserved and divergent traits in gene expression, and identify predictor genes shared across the species that are associated with pluripotent states in vivo and in vitro Amongst these are the pluripotency-linked genes Klf4 and Lin28b The novel genes discovered include naive- (Spic, Scpep1 and Gjb5) and primed-associated (Sema6a and Jakmip2) genes as well as naive-to primed transition genes (Dusp6 and Trip6). Both Gjb5 and Dusp6 play a role in pluripotency since their knockdown results in differentiation and downregulation of key pluripotency genes. Together, our interspecies comparison revealed new insights of pluripotency, pluripotent stem cell identity and a new molecular criterion for distinguishing between pluripotent states in various species including human. RNA-Seq profiles of three embryonic stages, 3 replicas per stage. This profiling is performed for mouse (mus musculus), pig (sus scrofa) and cow (bos taurus). Furthermore 4 RNA-Seq samples on mouse liver RNA to investigate various methods of amplification for small quantities of RNA.在哺乳动物早期发育过程中，会出现短暂存在的多能细胞（pluripotent cells）群，这类细胞可通过干细胞建系实现永生化。多能状态可分为naive态（naive）与primed态（primed），其具体类型取决于胚胎发育阶段及所采用的建系条件。本研究针对携带有不同类型多能细胞的胚胎阶段，分析了小鼠、牛及猪胚胎的时序基因表达模式。本研究记录了基因表达中的保守特征与差异特征，并鉴定出跨物种共有的、与体内外多能状态相关的预测基因，其中包括与多能性相关的Klf4与Lin28b基因。本次发现的新基因包括与naive态相关的Spic、Scpep1及Gjb5，与primed态相关的Sema6a及Jakmip2，以及参与naive态向primed态转变的Dusp6与Trip6。Gjb5与Dusp6均参与多能性调控：对二者进行敲低会导致细胞分化，并下调关键多能性基因的表达。综上，本研究通过跨物种比较，为多能性调控机制、多能干细胞特性解析提供了新见解，并为包括人类在内的多个物种区分多能状态提供了全新的分子判定标准。本研究对3个胚胎发育阶段进行了RNA测序（RNA-Seq）分析，每个阶段设置3个生物学重复。测序样本涵盖小鼠（Mus musculus）、猪（Sus scrofa）及牛（Bos taurus）。此外，本研究还制备了4份小鼠肝脏RNA的RNA测序样本，用于探究针对微量RNA的多种扩增方法。