Deletion of neural tube defect-associated gene Mthfd1l causes reduced cranial mesenchyme density.

BACKGROUND: Periconceptional intake of supplemental folic acid can reduce the incidence of neural tube defects by as much as 70%, but the mechanisms by which folic acid supports cellular processes during neural tube closure are unknown. The mitochondrial 10-formyl-tetrahydrofolate synthetase MTHFD1L catalyzes production of formate, thus generating one-carbon units for cytoplasmic processes. Deletion of Mthfd1l causes embryonic lethality, developmental delay, and neural tube defects in mice. METHODS: To investigate the role of mitochondrial one-carbon metabolism during cranial neural tube closure, we have analyzed cellular morphology and function in neural tissues in Mthfd1l knockout embryos. RESULTS: The head mesenchyme showed significantly lower cellular density in Mthfd1l nullizygous embryos compared to wildtype embryos during the process of neural tube closure. Apoptosis and neural crest cell specification were not affected by deletion of Mthfd1l. Sections from the cranial region of Mthfd1l knockout embryos exhibited decreased cellular proliferation, but only after completion of neural tube closure. Supplementation of pregnant dams with formate improved mesenchymal density and corrected cell proliferation in the nullizygous embryos. CONCLUSIONS: Deletion of Mthfd1l causes decreased density in the cranial mesenchyme and this defect is improved with formate supplementation. This study reveals a mechanistic link between folate-dependent mitochondrially produced formate, head mesenchyme formation and neural tube defects.

Deletion of the neural tube defect-associated gene Mthfd1l disrupts one-carbon and central energy metabolism in mouse embryos.

One-carbon (1C) metabolism is a universal folate-dependent pathway essential for de novo purine and thymidylate synthesis, amino acid interconversion, universal methyl-donor production, and regeneration of redox cofactors. Homozygous deletion of the 1C pathway gene Mthfd1l encoding methylenetetrahydrofolate dehydrogenase (NADP+-dependent) 1-like, which catalyzes mitochondrial formate production from 10-formyltetrahydrofolate, results in 100% penetrant embryonic neural tube defects (NTDs), underscoring the central role of mitochondrially derived formate in embryonic development and providing a mechanistic link between folate and NTDs. However, the specific metabolic processes that are perturbed by Mthfd1l deletion are not known. Here, we performed untargeted metabolomics on whole Mthfd1l-null and wildtype mouse embryos in combination with isotope tracer analysis in mouse embryonic fibroblast (MEF) cell lines to identify Mthfd1l deletion-induced disruptions in 1C metabolism, glycolysis, and the TCA cycle. We found that maternal formate supplementation largely corrects these disruptions in Mthfd1l-null embryos. Serine tracer experiments revealed that Mthfd1l-null MEFs have altered methionine synthesis, indicating that Mthfd1l deletion impairs the methyl cycle. Supplementation of Mthfd1l-null MEFs with formate, hypoxanthine, or combined hypoxanthine and thymidine restored their growth to wildtype levels. Thymidine addition alone was ineffective, suggesting a purine synthesis defect in Mthfd1l-null MEFs. Tracer experiments also revealed lower proportions of labeled hypoxanthine and inosine monophosphate in Mthfd1l-null than in wildtype MEFs, suggesting that Mthfd1l deletion results in increased reliance on the purine salvage pathway. These results indicate that disruptions of mitochondrial 1C metabolism have wide-ranging consequences for many metabolic processes, including those that may not directly interact with 1C metabolism.

Mitochondrial one-carbon metabolism and neural tube defects.

BACKGROUND: Neural tube defects (NTDs) are one of the most common birth defects in humans. Maternal intake of folic acid was linked to prevention of NTDs in the 1970s. This realization led to the establishment of mandatory and/or voluntary food folic acid fortification programs in many countries that have reduced the incidence of NTDs by up to 70% in humans. Despite 40 years of intensive research, the biochemical mechanisms underlying the protective effects of folic acid remain unknown. RESULTS: Recent research reveals a role for mitochondrial folate-dependent one-carbon metabolism in neural tube closure. CONCLUSION: In this article, we review the evidence linking NTDs to aberrant mitochondrial one-carbon metabolism in humans and mouse models. The potential of formate, a product of mitochondrial one-carbon metabolism, to prevent NTDs is also discussed.

Deletion of Mthfd1l causes embryonic lethality and neural tube and craniofacial defects in mice.

Maternal supplementation with folic acid is known to reduce the incidence of neural tube defects (NTDs) by as much as 70%. Despite the strong clinical link between folate and NTDs, the biochemical mechanisms through which folic acid acts during neural tube development remain undefined. The Mthfd1l gene encodes a mitochondrial monofunctional 10-formyl-tetrahydrofolate synthetase, termed MTHFD1L. This gene is expressed in adults and at all stages of mammalian embryogenesis with localized regions of higher expression along the neural tube, developing brain, craniofacial structures, limb buds, and tail bud. In both embryos and adults, MTHFD1L catalyzes the last step in the flow of one-carbon units from mitochondria to cytoplasm, producing formate from 10-formyl-THF. To investigate the role of mitochondrial formate production during embryonic development, we have analyzed Mthfd1l knockout mice. All embryos lacking Mthfd1l exhibit aberrant neural tube closure including craniorachischisis and exencephaly and/or a wavy neural tube. This fully penetrant folate-pathway mouse model does not require feeding a folate-deficient diet to cause this phenotype. Maternal supplementation with sodium formate decreases the incidence of NTDs and partially rescues the growth defect in embryos lacking Mthfd1l. These results reveal the critical role of mitochondrially derived formate in mammalian development, providing a mechanistic link between folic acid and NTDs. In light of previous studies linking a common splice variant in the human MTHFD1L gene with increased risk for NTDs, this mouse model provides a powerful system to help elucidate the specific metabolic mechanisms that underlie folate-associated birth defects, including NTDs.