Maternal Vitamin C Deficiency and Genetic Risk Factors Contribute to Congenital Malformations through Dysregulation of DNA Methylation. [RNA-Seq E8.5]
Maternal dietary insufficiencies can reshape the offspring epigenome during gestation, contributing to birth defects and developmental disorders. Vitamin C (VitC) is a critical co-factor for Ten-Eleven-Translocation (TET) DNA demethylases, but the impact of its deficiency on embryonic development remains unclear. Here, we show that insufficient maternal VitC combined with genetic susceptibility can give rise to congenital malformations, including neural tube defects (NTDs), through dysregulation of DNA methylation. We previously reported NTDs in Tet1 knockout (KO) mice at low penetrance in C57BL/6J (B6) congenic inbred strains, but at two- to three-fold higher rates when outbred or highly backcrossed (incipient congenic) on a 129S6.B6 background. Similarly, maternal VitC deficiency in L-gulonolactone oxidase (Gulo) KO mice, which like humans are unable to synthesize VitC, resulted in highly penetrant congenital malformations in non-inbred mice, during a vulnerable window coinciding with gastrulation. DNA hypermethylation is a signature of VitC-deprived 129S6.B6-Gulo-/- embryonic headfold tissues, being absent in B6 embryos which are grossly normal. In outbred Gulo-/- embryonic brains, genomic regions harboring hypermethylation – hallmarks of TET dysfunction - increased with the severity of embryonic pathologies. A moderate reduction in VitC status is sufficient to induce hypermethylated regions and cause NTDs. Severe embryonic defects in VitC-deprived embryos can be rescued by timely re-supplementation of VitC at the onset of gastrulation, which normalized DNA methylation at most loci. Our results suggest that promoting timely VitC supplementation by at-risk pregnant mothers may prevent many birth defects currently refractory to folic acid supplementation and enhance the “health-span” of future generations. Overall design: RNA-seq in E8.5 headfolds, collected from B6 and 129S6.B6-Gulo<-/-> embryos, with or without VitC supplementation.