Wnt3a Activates the WNT-YAP/TAZ Pathway to Sustain Expression in Bovine Trophoblast Stem Cells.

Trophoblast stem cells (TSCs), the precursors of placental cells, are effective for studying placental formation . Using a dual inhibition (2i) medium and mixed L-Wnt3a/mouse embryonic fibroblast feeder cells, we previously established the bovine trophoblast cell line BTS-1. In this study, we used bovine fetal fibroblasts and added Wnt3a to the 2i medium to establish another bovine TSC line (BTSW). BTSW cells expressed pluripotency markers, including , , , , , , , and , and TSC markers , , and . Methylation sequencing of the promoter regions of , , and revealed no significant differences between BTS-1 and BTSW cells. Removal of Wnt3a from the culture medium resulted in downregulation ( < 0.05) of , , , and TSC marker genes, and upregulation of TSC differentiation markers, including , , and . Western blotting indicated activation of the WNT-YAP/TAZ signaling pathway in BTS-1 and BTSW cells, consequently activating transcription. However, this pathway was not activated in BCFF cells, an established bovine embryonic stem-like cell line that expresses , , and , but not . Thus, Wnt3a may play a critical role in bovine TSC maintenance by activating and regulating expression through the WNT-YAP/TAZ signaling pathway.