Factors that sustain self-renewal of mouse embryonic stem cells (ESCs) are well described. In contrast, the machinery regulating exit from pluripotency is ill defined. In a large-scale small interfering RNA (siRNA) screen, we found that knockdown of the tumor suppressors Folliculin ( Flcn) and Tsc2 prevent ESC commitment. Tsc2 lies upstream of mammalian target of rapamycin (mTOR), whereas Flcn acts downstream and in parallel. Flcn with its interaction partners Fnip1 and Fnip2 drives differentiation by restricting nuclear localization and activity of the bHLH transcription factor Tfe3. Conversely, enforced nuclear Tfe3 enables ESCs to withstand differentiation conditions. Genome-wide location and functional analyses showed that Tfe3 directly integrates into the pluripotency circuitry through transcriptional regulation of Esrrb. These findings identify a cell-intrinsic rheostat for destabilizing ground-state pluripotency to allow lineage commitment. Congruently, stage-specific subcellular relocalization of Tfe3 suggests that Flcn-Fnip1/2 contributes to developmental progression of the pluripotent epiblast in vivo.
► A large-scale siRNA screen identifies drivers of exit from ground-state pluripotency ► Tsc1/2 and Flcn-Fnip1/2 promote ESC commitment via nuclear exclusion of Tfe3 ► Nuclear Tfe3 confers autonomous ESC self-renewal ► Tfe3 directly regulates transcription of the core pluripotency factor Esrrb
An siRNA screen in mouse embryonic stem cells uncovers a signaling axis at the onset of differentiation that stimulates nuclear export of Tfe3, a transcription factor that regulates the pluripotent transcription factor network. This study highlights subcellular compartmentalization of cell-state regulators as a means of enabling cell-fate transition.