Genome-wide CRISPR-KO Screen Uncovers mTORC1-Mediated Gsk3 Regulation in Naive Pluripotency Maintenance and Dissolution

Abstract

Summary: The genetic basis of naive pluripotency maintenance and loss is a central question in embryonic stem cell biology. Here, we deploy CRISPR-knockout-based screens in mouse embryonic stem cells to interrogate this question through a genome-wide, non-biased approach using the Rex1GFP reporter as a phenotypic readout. This highly sensitive and efficient method identified genes in diverse biological processes and pathways. We uncovered a key role for negative regulators of mTORC1 in maintenance and exit from naive pluripotency and provided an integrated account of how mTORC1 activity influences naive pluripotency through Gsk3. Our study therefore reinforces Gsk3 as the central node and provides a comprehensive, data-rich resource that will improve our understanding of mechanisms regulating pluripotency and stimulate avenues for further mechanistic studies. : Li et al. conducted genome-wide CRISPR screens in mouse ESCs to identify genes affecting maintenance of and exit from naive pluripotency using a Rex1GFP reporter. They show that loss of two mTORC1-negative regulators, Tsc1/2 and Gator1, can cause opposing phenotypes through differential regulation of Gsk3 activity. Keywords: CRISPR, screening, naive pluripotency, exit from pluripotency, Akt, mTORC1, mTORC2, GATOR1, Nprl2, Tsc2