The onset of lineage specification and loss of pluripotency during embryonic development occurs during gastrulation, the process by which the three germ layers are established from the pluripotent epiblast. The hallmark of gastrulation is the formation of the primitive streak (PS) a transient structure that defines the posterior of the epiblast and the basic embryonic body plan. Prior to PS formation, the epiblast is essentially symmetrical; however, localized gene ex
The signaling pathways activated by the ligands expressed in the EXE signaling center function to promote the formation of the PS. Genetic ablation of Wnt3, Betacatenin, or the Wnt receptors Lrp5/6 completely blocked PS formation. Conversely, over-activation of Wnt signaling by forced ex
At approximately e6.5, this localization of Wnt, Nodal, and BMP signaling activity results in the induction of PS formation at the proximal posterior region of the epiblast, which defines the embryonic posterior. At this site, both morphological changes and lineage specification are induced in epiblast cells. Epiblast cells in the PS begin to express markers of mesoderm lineage specification and undergo an epithelial-to-mesenchymal transition. The result of this is the formation of a new layer of cells: the mesoderm. As gastrulation proceeds, epiblast cells rapidly proliferate and migrate towards the PS. Subsequently, the mesoderm layer expands laterally around the epiblast. Cells from the PS also contribute to the outer layer of definitive endoderm by intercalating into existing endoderm layer. Finally, anterior epiblast cells that do not enter the PS undergo neurectoderm lineage specification. As such, by e7.5, the three germ layers are specified and the basic embryonic body plan is established.
During PS formation, the pluripotency of the epiblast is progressively lost as cells undergo lineage specification. Cell labeling and transplantation experiments showed that up until the early stages of PS formation (~e6.5), the epiblast was uniformly pluripotent and cells from any part of the epiblast could contribute to all three germ layers. However, as PS formation progressed, the developmental potential of epiblast cells was restricted. In embryos with a fully formed PS (~e7.5), cells from the posterior epiblast predominantly contributed to mesodermal cell types and cells from the anterior contributed to neural and ectodermal cell types. In teratoma formation assays, embryos could give contribute to all three germ layers up until e7.5, suggesting that the pluripotency of embryos was lost after PS formation. In a more recent series of experiments, the Chambers and Wilson groups examined epiblast pluripotency by deriving EpiSC from embryos at various stages of PS formation. They found that EpiSC could only be derived from embryos up until ~e8.0, validating the previous findings that epiblast pluripotency was lost following PS formation.