Fan Y, Melhem M F, Chaillet J R
Department of Biological Sciences, University of Pittsburgh, Pittsburgh, Pennsylvania, 15260, USA.
Dev Biol. 1999 Jun 15;210(2):481-96. doi: 10.1006/dbio.1999.9279.
Similarities in the differentiation of mouse embryos and ES cell embryoid bodies suggest that aspects of early mammalian embryogenesis can be studied in ES cell embryoid bodies. In an effort to understand the regulation of cellular differentiation during early mouse embryogenesis, we altered the expression of the Pem homeobox-containing gene in ES cells. Pem is normally expressed in the preimplantation embryo and expressed in a lineage-restricted fashion following implantation, suggesting a role for Pem in regulating cellular differentiation in the early embryo. Here, we show that the forced expression of Pem from the mouse Pgk-1 promoter in ES cells blocks the in vitro and in vivo differentiation of the cells. In particular, embryoid bodies produced from these Pgk-Pem ES cells do not differentiate into primitive endoderm or embryonic ectoderm, which are prominent features of early embryoid bodies from normal ES cells. This Pgk-Pem phenotype is also different from the null phenotype, as embryoid bodies derived from ES cells in which endogenous Pem gene expression has been blocked show a pattern of differentiation similar to that of normal ES cells. When the Pgk-Pem ES cells were introduced into subcutaneous sites of nude mice, only undifferentiated EC-like cells were found in the teratomas derived from the injected cells. The Pem-dependent block of ES cell differentiation appears to be cell autonomous; Pgk-Pem ES cells did not differentiate when mixed with normal, differentiating ES cells. A block to ES cell differentiation, resulting from the forced expression of Pem, can also be produced by the forced expression of the nonhomeodomain region of Pem. These studies are consistent with a role for Pem in regulating the transition between undifferentiated and differentiated cells of the early mouse embryo.
小鼠胚胎与胚胎干细胞来源的拟胚体在分化过程中的相似性表明，早期哺乳动物胚胎发育的某些方面可以在胚胎干细胞来源的拟胚体中进行研究。为了理解小鼠早期胚胎发育过程中细胞分化的调控机制，我们改变了胚胎干细胞中含Pem同源框基因的表达。Pem通常在植入前胚胎中表达，植入后以谱系限制的方式表达，这表明Pem在调节早期胚胎细胞分化中发挥作用。在此，我们表明，在胚胎干细胞中由小鼠Pgk-1启动子强制表达Pem会阻断细胞的体外和体内分化。特别是，这些Pgk-Pem胚胎干细胞产生的拟胚体不会分化为原始内胚层或胚胎外胚层，而这是正常胚胎干细胞来源的早期拟胚体的显著特征。这种Pgk-Pem表型也不同于基因敲除表型，因为源自内源性Pem基因表达被阻断的胚胎干细胞的拟胚体显示出与正常胚胎干细胞相似的分化模式。当将Pgk-Pem胚胎干细胞引入裸鼠皮下部位时，在源自注射细胞的畸胎瘤中仅发现未分化的类EC细胞。Pem依赖的胚胎干细胞分化阻断似乎是细胞自主性的；Pgk-Pem胚胎干细胞与正常的、正在分化的胚胎干细胞混合时不会分化。由Pem的强制表达导致的胚胎干细胞分化阻断，也可由Pem的非同源结构域区域的强制表达产生。这些研究结果与Pem在调节小鼠早期胚胎未分化细胞和分化细胞之间的转变中所起的作用一致。
