UCSF

Mouse embryonic stem cells (mESCs), an in vitro model for naive pluripotent differentiation into primed pluripotent epiblast-like cells (EpiLCs), have been used to reveal transcriptional and epigenetic control of early embryonic development. However, the control and significance of morphological changes during lineage specification remain less defined. We show with quantitative time-lapse imaging marked changes in morphology and actin architectures during differentiation that depend on activity of the Arp2/3 complex, an actin filament nucleator. Inhibiting Arp2/3 complex activity pharmacologically or genetically does not block exit from naive pluripotency but attenuates increases in EpiLC markers compared with controls. We found that loss of Arp2/3 complex activity delays entry into the intermediate formative pluripotent state, resulting in globally defective lineage specification as indicated by RNA-sequencing with marked effects on TBX3-depedendent transcriptional programs. Further, we identified two previously unreported indicators of pluripotent status; MRTF and FHL2, which have inverse actin-dependent nuclear translocation as competing SRF co-factors that is dependent on Arp2/3 complex but not formin activity. These data reveal a previously unrecognized role for Arp2/3 complex-dependent actin remodeling in mESC differentiation, timing of formative pluripotency, and TBX3-dependent lineage specification, as well as newly identified MRTF and FHL2 nuclear shuttling. Moreover, our current findings on a role for Arp2/3 complex activity in mESC differentiation compared with the established role for formin activity in epithelial to mesenchymal transition indicate that distinct actin nucleators regulate distinct modes of epithelial plasticity.