Genome-wide CRISPR/Cas9 screening in mouse primordial germ cell specification

Authors

• XK Wang
• SR Malki
• B Bennett
• G Hu

Abstract

The germ cell lineage ensures the transmission of heritable genetic and epigenetic information to the next generation. In mammals, the pluripotent epiblast cells in the post-implantation embryos give rise to both the somatic and germ cell lineages during subsequent development. Under inductive cell interactions, a small number of epiblast cells adopt the fate of the primordial germ cells (PGCs) and become the multipotent precursor of germ cells. However, the underlying mechanism is not fully understood. Fortunately, the epiblast to PGC specification can be recapitulated in vitro using embryonic stem cells (ESCs), albeit with low efficiency. The specification of PGCs is accompanied by the induction of key germ-cell genes, repression of the nascent somatic program, and widespread epigenetic remodeling. Here, we carried out a CRISPR/Cas9-mediated genome-wide genetic screen in PGC-like cells (PGCLCs) in vitro differentiation system and identified several key factors that are epigenetic barriers to PGC specification. Phenotypically, when the knockout of these genes or the use of chemical inhibitors against them, it promoted PGCLCs differentiation significantly up to ~20% in vitro. Consistently, the transcriptional analysis showed the germ cell maker genes are highly activated in the knockout PGCLCs. We are currently using genomic approaches including RNA-seq and CUT&TAG to dissect the underlying mechanism. Further, we are generating knockout mouse models or embryo aggregation to test whether the deletion can promote PGC specification in vivo. Together, our study uncovered epigenetic barriers in PGC cell fate specification and illustrated the power of genetic screens in the study of cell fate transitions.

Scientific Focus Area: Stem Cell Biology

This page was last updated on Monday, September 25, 2023