Breaking the first lineage barrier: Direct induction of trophoblast stem cells from murine fibroblasts

Trophoblast stem cells represent the stem cell population of the extra-embryonic lineage and arise as a result of the first cell fate decision. From blastocyst stage onwards, a distinct epigenetic lineage barrier strictly separates mouse embryonic and extra-embryonic lineages. Recently, it has been shown that this epigenetic barrier cannot be fully overcome as the expression of TS-determining factors in embryonic stem cells lead to incomplete transdifferentiation. Here, we demonstrate that transient expression of Tfap2c, Gata3, Eomes and Ets2 in fibroblasts suffices to generate cells which are almost identical to trophoblast stem cells based on morphology, expression and methylation pattern. Further, these induced trophoblast stem cells display transgene independent self-renewal, differentiate along the extra-embryonic lineage and chimerize the placenta upon blastocyst injection. Our findings provide insights into the transcription factor networks governing trophoblast stem cell identity and offer a new tool for studying the hierarchy of those factors. 11 samples were analyzed: MEF4F, mouse embyronic fibroblasts (MEFs) transduced with Tfap2c, Gata3, Eomes and Ets2, 3 replicates. TS, mouse TS-EGFP trophoblast stem cells, 2 replicates. iTSC4F5, mouse induced trophoblast stem (TS) cells clone A from 4 factor transduced MEFs, 3 replicates. iTSC4F8, mouse induced trophoblast stem (TS) cells clone D from 4 factor transduced MEFs, 3 replicates.