Tracing the Transitions from Pluripotency to Germ Cell Fate with CRISPR Screening

Abstract

Early mammalian development entails transit through naïve pluripotency towards post-implantation epiblast, which subsequently gives rise to primordial germ cells (PGC), the founding germline population. To investigate these cell fate transitions, we developed a compound-reporter to track cellular identity in a model of PGC specification (PGC-like cells;PGCLC), and coupled it with genome-wide CRISPR-screening. We identify key genes both for exit from pluripotency and for acquisition of PGC fate, and characterise a central role for the transcription-regulators Nr5a2 and Zfp296 in germline ontogeny. Abrogation of these genes results in widespread activation (Nr5a2-/-) or inhibition (Zfp296-/-) of WNT-pathway factors in PGCLC. This leads to aberrant upregulation of the somatic programme or failure to activate germline-genes, respectively, and consequently loss of germ cell identity. Our study places Zfp296 and Nr5a2 as key components of an expanded PGC gene regulatory network, and outlines a transferable strategy for identifying critical regulators of complex cell fate decisions.