EpCAM Supports Exit from Pluripotency of Embryonic Stem Cells via Eomes

Gene expression association of Epcam with Eomes, Foxa2, and Gata6 was prominent at day three of spontaneous differentiation, representing primitive streak formation in EBs. Interrogation of public single-cell RNA sequencing (scRNAseq) datasets supported a co-expression of Epcam and Eomes at early stages of murine embryogenesis in epiblast, primitive streak, nascent mesoderm, extraembryonic ectoderm and endoderm. Newly generated scRNAseq of wildtype mESCs in spontaneous differentiation confirmed the formation of essential stages including epiblast, primitive streak, endo- and mesoderm cells, and cardiomyocytes.

Expression and pseudotime analysis positioned Epcam expression slightly ahead of Eomes at the transition of early to late primitive streak, along with rising of Wnt signaling. Accordingly, conditional re-expression of Epcam or Eomes but not of Foxa2 or Gata6 complemented differentiation defects of Epcam-knockouts and confirmed an involvement of Wnt signaling in the EpCAM-dependent activation of Eomes. Hence, defective exit of pluripotency in Epcam-deficient ESCs is linked to Eomes regulation via Wnt signaling.