Fig. 1: Mouse pre-implantation development from the fertilized one-cell stage to the blastocyst (64-cell) stage. Individual cells of cleavage-stage embryos (one-cell to eight-cell stages) remain totipotent, meaning that each one has the capacity to form an entire organism. By the 64-cell stage, three distinct cell types are distinguishable. The inner cell mass is visible at the top of the blastocyst.
© 2010 P. Robson
Researchers in Singapore have determined patterns of gene expression in single cells surgically isolated from very early mouse embryos to understand how different cell types arise before the implantation of the embryo into the mother’s uterus.
Soon after fertilization, a series of cleavages leads to the formation of a cell bundle called the morula, which later develops into the blastocyst containing a cavity that separates the outer cell layer from the inner cell mass (ICM) (Fig. 1).
At around the 64-cell stage, just before implantation, the blastocyst contains three different cell types—cells of the trophectoderm, the primitive endoderm, and the epiblast. Embryonic stem cells are derived from the epiblast.
“Until now, the regulatory genetic networks that give rise to these different cell types in the pre-implantation embryo have been poorly understood,” says team leader Paul Robson of the Genome Institute of Singapore, A*STAR. “We set out to define the earliest events leading to cell differentiation during the morula–blastocyst transition.”
Robson and his co-workers analyzed the expression of 48 different genes in more than 500 individual cells sampled from fertilization to the 64-cell stage. The genes studied mainly encode proteins called transcription factors that regulate the expression of other genes. They found that up to the early 16-cell stage, there were no recognizable differences between individual cells within an embryo.
“These early morula-stage cells all expressed a similar range of transcription factors, indicating that different cell types had not yet formed,” explains Robson. “But by the 64-cell stage, the three main cell types that make up the blastocyst were expressing characteristic suites of transcription factors and could be readily distinguished.
The team also showed that the earliest, detectable, differential expression occurred in the late morula, with the transcription factor Id2 marking outer cells and Sox2, a key reprogramming factor, marking the inner cells.
Later, at around the 32-cell stage, the researchers identified an inverse correlation in the expression of the signaling molecule FGF4 and its receptor between individual cells of the ICM. Inhibition of this signaling axis led to a block in primitive endoderm formation with all ICM cells becoming epiblastic. This suggests that signaling events determining cell position within the blastocyst occur before maturation of the transcriptional program.
“Our results, based on the powerful approach of single-cell gene expression profiling, have provided unprecedented insight into the regulation of cell differentiation during the morula–blastocyst transition in the mouse,” says Robson.
The A*STAR-affiliated researchers mentioned in this highlight are from the Genome Institute of Singapore.
