Snail and the microRNA-200 Family Act in Opposition to Regulate Epithelial-to-Mesenchymal Transition and Germ Layer Fate Restriction in Differentiating ESCs

The reprogramming of somatic cells to inducible pluripotent stem cells requires a mesenchymal-to-epithelial transition. While differentiating ESCs can undergo the reverse process or epithelial-to-mesenchymal transition (EMT), little is known about the role of EMT in ESC differentiation and fate commitment. Here, we show that Snail homolog 1 (Snail) is expressed during ESC differentiation and is capable of inducing EMT on day 2 of ESC differentiation. Induction of EMT by Snail promotes mesoderm commitment while repressing markers of the primitive ectoderm and epiblast. Snail's impact on differentiation can be partly explained through its regulation of a number of ESC-associated microRNAs, including the microRNA-200 (miR-200) family. The miR-200 family is normally expressed in ESCs but is downregulated in a Wnt-dependent manner during EMT. Maintenance of miR-200 expression stalls differentiating ESCs at the epiblast-like stem cell (EpiSC) stage. Consistent with a role for activin in maintaining the EpiSC state, we find that inhibition of activin signaling decreases miR-200 expression and allows EMT to proceed with a bias toward neuroectoderm commitment. Furthermore, miR-200 requires activin to efficiently maintain cells at the epiblast stage. Together, these findings demonstrate that Snail and miR-200 act in opposition to regulate EMT and exit from the EpiSC stage toward induction of germ layer fates. By modulating expression levels of Snail, activin, and miR-200, we are able to control the order in which cells undergo EMT and transition out of the EpiSC state. STEM CELLS 2011;29:764-776