HAND1 controls the lineage divergence between amnion and trophoblast from human pluripotent stem cells

Trophoblasts and amnion, as extra-embryonic lineages, can be differentiated from human pluripotent stem cells (PSCs) under defined culture conditions, but the regulatory mechanisms that coordinate the cell fate decisions between these lineages during PSC differentiation remain poorly defined. In this study, leveraging CRISPR/Cas9-mediated targeted screening and lineage reporter pluripotent stem cells (PSCs), we identified the transcription factor HAND1 as a critical determinant governing the divergence of trophoblast and amnion lineages in PSCs. Genetic ablation of HAND1 effectively suppressed the amnion differentiation capacity of EPSCs while enhancing their trophoblast differentiation potential. Conversely, ectopic overexpression of HAND1 have adverse effects. Notably, forced expression of HAND1 activated amnionic transcription program in human trophoblast stem cells (TSCs), demonstrating its lineage-reprogramming capability. Mechanistic analyses revealed that HAND1 cooperates with Wnt signaling pathway co-factors TCF/b-catenin to form a transcriptional complex that antagonistically regulates trophoblast- and amnion-associated gene networks. Collectively, our findings establish HAND1 as a pivotal regulator that orchestrates the fate choice between amnionic and trophoblast lineages during human PSC differentiation, providing insights into the molecular logic of extra-embryonic lineage specification.