FGFR-mediated ERK1/2 signaling contributes to mesendoderm and definitive endoderm formation in vitro

The differentiation of human pluripotent stem cells into the SOX17(+) definitive endoderm (DE) germ layer is important for generating tissues for regenerative medicine. Multiple developmental and stem cell studies have demonstrated that Activin/Nodal signaling is the primary driver of definitive endoderm forma-tion. Here, we uncover that the FGF2-FGFR-ERK1/2 signaling contributes to mes-endoderm and SOX17(+) DE formation. Without ERK1/2 signaling, the Activin/ Nodal signaling is insufficient to drive mesendoderm and DE formation. Besides FGF2-FGFR-mediated signaling, IGF1R signaling possibly contributes to the ERK1/2 signaling for DE formation. We identified a temporal relationship be-tween Activin/Nodal-SMAD2 and FGF2-FGFR-ERK1/2 signaling in which Activin/Nodal-SMAD2 participates in the initiation of mesendoderm and DE spec-ification that is followed by increasing activity of FGF2-FGFR-ERK1/2 to facilitate and permit the successful generation of SOX17(+) DE. Overall, besides the role of Activin/Nodal signaling for DE formation, our findings shed light on the contribu-tion of ERK1/2 signaling for mesendoderm and DE formation.