CRISPR Screening Reveals a Novel Role for FOXH1 in Regulating Pluripotency of Porcine Embryonic Stem Cells

ABSTRACT: pESCs hold significant potential for applications in regenerative medicine and stem cell-based breeding programs. However, the regulatory mechanisms governing pluripotency in pESCs, including aspects of cellular adaptability and the maintenance of a pluripotent identity, are not yet fully understood.To distinguish these two features of pESCs, we conducted two types of CRISPR-Cas9 knockout screenings targeting all known transcription factors and epigenetic regulators in the porcine genome. Through the CRISPR-Cas9 screenings, we identified 900 essential genes, including the species-conserved gene PRMT1 and the pig-specific gene NASP, which were experimentally validated for their roles in preserving the survival of pESCs. In addition, we found that FOXH1 is indispensable for maintaining pESCs pluripotency. pESCs deficient in FOXH1 displayed a flatter and more dispersed clonal morphology, accompanied by downregulation of pluripotency genes and upregulation of lineage-specific genes. In parallel, FOXH1 knockdown significantly impaired blastocyst formation during early pig embryogenesis, demonstrating an unrecognized role of FOXH1 that is not conserved in hESCs. At the molecular level, FOXH1 exhibits a dual role by regulating chromatin accessibility at pluripotency gene loci and H3K4me3 modifications at the promoters of developmental genes. The resources developed through this study not only lay the groundwork for more extensive CRISPR screenings but also provide crucial theoretical support for the establishment of pPSC cell lines with higher differentiation potential.

ORGANISM(S): Sus scrofa

PROVIDER: GSE283822 | GEO | 2025/07/01

REPOSITORIES: GEO