Patient-specific iPSC models of neural tube defects identify underlying deficiencies in neuroepithelial cell shape regulation and differentiation

Spina bifida and anencephaly are neural tube defects caused by failure of embryonic neural tube closure. Successful closure requires elongation and apical constriction of neuroepithelial cells. These critical behaviours have not been studied in human, patient-specific models. We characterise a human iPSC-derived neuroepithelial morphogenesis model which is highly reproducible between lines. Differentiated neuroepithelial cells actively undergo ROCK-dependent apical constriction. ROCK acts downstream of planar cell polarity/VANGL2 in other species. We find that VANGL2 is up-regulated and phosphorylated during iPSCs-to-neuroepithelial differentiation. The patient-identifiedVANGL2-R353Cmutation does not alter its expression, localisation or phosphorylation, but reduces myosin-II phosphorylation and apical constriction relative to congenic control iPSCs. Non-congenic comparisons and forward genetic testing are also informative in this reproducible model. We compare lines reprogrammed from amniocytes of two patients with spina bifida, versus two controls. One patient-derived line forms a morphologically normal neuroepithelium, but fails to differentiate into neurons. The second fails to undergo apicobasal elongation, and harbours compound heterozygous mutations in theMED24gene previously implicated in neuroepithelial elongation in mice. Thus, iPSC-derived neuroepithelial modelling provides mechanistic insights into conserved cell behaviours, links genetically-impaired apical constriction with a human disease, and establishes patient-specific models which recapitulate failures of heterogenous neuroepithelial functions.