Loss of FANCM impairs primordial germ cell differentiation in vitro

Abstract

Pathogenic variants in FANCM have been implicated in premature ovarian insufficiency, suggesting a critical role for FANCM in early germline development. To investigate this genes function in a controlled system, we evaluated whether a mouse in vitro primordial germ cell (PGC) model could be used to interrogate the consequences of FANCM loss on germ cell specification. Using CRISPR-Cas9 gene editing, we introduced a premature stop codon into exon 1 of Fancm in mouse embryonic stem cells. These edited cells were differentiated into epiblast-like cells and subsequently into primordial germ cell-like cells (PGCLCs), which were assessed for their developmental competence. After successful generation of in vitro-derived PGCLCs, we found that loss of FANCM markedly reduced PGCLC formation, consistent with previously reported in vivo phenotypes. Our findings demonstrate that this in vitro system provides a tractable platform for dissecting gene function in otherwise inaccessible stages of germline development. Moreover, our data suggest that FANCM is required at the earliest stages of PGC specification, potentially as early as the developmental window equivalent to embryonic day 8.5, a period not previously examined in vivo.