Single-cell omics uncover gene dynamics shaping embryonic and extra embryonic lineages in pig blastocysts

Late blastocyst development before implantation is a unique feature of ungulates, during which the epiblast proliferates and maintains pluripotency while extra-embryonic tissues expand dramatically, elongating to several tens of centimeters. The mechanisms coordinating these processes are not well understood. We performed single-cell omics profiling of porcine blastocysts from the hatched stage (E7) through early (E9) and late ovoid stages (E11). From 15,370 cells, we identified distinct embryonic and extra-embryonic populations with characteristic chromatin accessibility profiles. We reconstructed gene regulatory networks using enhancer-based eRegulons and validated them through motif occupancy analysis. Extra-embryonic tissues showed strong shifts in gene regulatory module activity at the onset of elongation, reflecting major transitions in morphogenesis and differentiation and the activation of pathways linked to cell morphology, proliferation, metabolism, trafficking, and biomolecule transport. In contrast, epiblast cells retained a stable transcriptional and regulatory identity from day 7 to day 11, immediately preceding the onset of gastrulation.