Targeted gene correction ofFKRPby CRISPR/Cas9 restores functional glycosylation of α-dystroglycan in cortical neurons derived from human induced pluripotent stem cells

Summary Mutations in genes required for functional glycosylation of α-dystroglycan cause a group of congenital muscular dystrophies associated with brain malformations, referred to as dystroglycanopathies. The lack of isogenic, physiology-relevant human cellular models has limited our understanding of the cortical abnormalities in dystroglycanopathies. Here we generate induced pluripotent stem cells (iPSCs) from a severe dystroglycanopathy patient with homozygous mutations in the ribitol-5-phosphate transferase gene, FKRP . We carry out targeted gene correction in FKRP-iPSCs using CRISPR/Cas9-mediated genome editing. We characterise the directed differentiation of FKRP- and corrected-iPSCs to neural stem cells, cortical progenitors and cortical neurons. Importantly, we show that targeted gene correction of FKRP restores functional glycosylation of α-dystroglycan in iPSC-derived cortical neurons. We independently validate this result by showing targeted gene mutation of FKRP disrupts functional glycosylation of α-dystroglycan. This work demonstrates the feasibility of using CRISPR/Cas9-engineered human iPSCs for modelling dystroglycanopathies and provides a foundation for therapeutic development. Highlights Generation of FKRP-iPSCs for modelling cortical abnormalities in dystroglycanopathies Precise gene correction by CRISPR/Cas9-mediated genome editing Directed differentiation of isogenic control and FKRP-iPSC to cortical neurons Functional glycosylation of α-dystroglycan is restored in cortical neurons derived from CRISPR/Cas9-corrected iPSCs Targeted gene mutation of FKRP disrupts functional glycosylation of α-dystroglycan in cortical neurons