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Background: As a biomarker, elevated serum erythritol concentrations predict type 2 diabetes and cardiovascular disease onset. Erythritol was recently shown to be a product of human glucose metabolism through the pentose phosphate pathway. The regulation of erythritol synthesis from glucose has been explored in cancer cells but not in nontransformed cells. Objective: The kidneys and skeletal muscle have increased erythritol content in response to dietary sucrose, which suggests that they may significantly contribute to circulating erythritol concentrations. In the present study, we evaluated if conditions that promote erythritol synthesis in cancer cells are consistent in skeletal muscle and kidney cells. Methods: C2C12 myotubules were used as a model for skeletal muscle, and human kidney (HK)-2 human proximal tubule cells were used to model kidney. C2C12 cells were exposed to high-or low-glucose conditions. Both C2C12 and HK-2 cells were exposed to the free radical generator menadione, then intracellular reactive oxygen species (ROS) and erythritol concentrations were measured. Intracellular sorbitol concentrations were also measured because increased polyol flux was also observed after exposure to excess glucose and oxidative stress. Results: Intracellular erythritol concentrations were significantly elevated in C2C12 cells following both high-glucose and menadione treatment. In contrast, HK-2 cells did not increase erythritol synthesis in response to oxidative stress. Generation of ROS through hydrogen peroxide exposure elevated sorbitol concentrations in both C2C12 and HK-2 cells, whereas generation of radicals with menadione treatment did not affect sorbitol production in either cell type. Conclusions: These findings highlight that the factors contributing to elevated erythritol synthesis vary between cell types. More specifically, these studies demonstrate that muscle cells increase erythritol synthesis in response to both high glucose in culture medium and oxidative stress, whereas kidney cells increase erythritol synthesis only in response to high glucose. [ABSTRACT FROM AUTHOR]

The article discusses a riboflavin evidence scan to update the riboflavin dietary reference intakes (DRI). Topics covered include components of the evidence scan, the processes involved, the reports on adequacy and chronic disease outcomes, and the limitations identified of studies that lead to exclusion. Also noted is the scan's identification of 35 studies that could be used for a future DRI committee for a full systematic review.

Background: Skeletal muscle progenitor cells (MPCs) repair damaged muscle postinjury. Pyruvate kinase M2 (PKM2) is a glycolytic enzyme (canonical activity) that can also interact with other proteins (noncanonical activity) to modify diverse cellular processes. Recent evidence links PKM2 to MPC proliferation.Objectives: This study aimed to understand cellular roles for PKM2 in MPCs and the necessity of PKM2 in MPCs for muscle regeneration postinjury.Methods: Cultured, proliferating MPCs (C2C12 cells) were treated with a short hairpin RNA targeting PKM2 or small molecules that selectively affect canonical and noncanonical PKM2 activity (shikonin and TEPP-46). Cell number was measured, and RNA-sequencing and metabolic assays were used in follow-up experiments. Immunoprecipitation coupled to proteomics was used to identify binding partners of PKM2. Lastly, an MPC-specific PKM2 knockout mouse was generated and challenged with a muscle injury to determine the impact of PKM2 on regeneration.Results: When the noncanonical activity of PKM2 was blocked or impaired, there was an increase in reactive oxygen species concentrations (1.6-2.0-fold, P < 0.01). Blocking noncanonical PKM2 activity also increased lactate excretion (1.2-1.6-fold, P < 0.05) and suppressed mitochondrial oxygen consumption (1.3-1.6-fold, P < 0.01). Glutamate dehydrogenase 1 (GLUD1) was identified as a PKM2 binding partner and blocking noncanonical PKM2 activity increased GLUD activity (1.5-1.6-fold, P < 0.05). Mice with an MPC-specific PKM2 deletion did not demonstrate impaired muscle regeneration.Conclusions: The results suggest that the noncanonical activity of PKM2 is important for MPC proliferation in vitro and demonstrate GLUD1 as a PKM2 binding partner. Because no impairments in muscle regeneration were detected in a mouse model, the endogenous environment may compensate for loss of PKM2. [ABSTRACT FROM AUTHOR]

Background: Adequate cellular thymidylate (dTMP) pools are essential for preservation of nuclear and mitochondrial genome stability. Previous studies have indicated that disruption in nuclear dTMP synthesis leads to increased uracil misincorporation into DNA, affecting genome stability. To date, the effects of impaired mitochondrial dTMP synthesis in nontransformed tissues have been understudied.Objectives: This study aimed to determine the effects of decreased serine hydroxymethyltransferase 2 (Shmt2) expression and dietary folate deficiency on mitochondrial DNA (mtDNA) integrity and mitochondrial function in mouse tissues.Methods: Liver mtDNA content, and uracil content in liver mtDNA, were measured in Shmt2+/- and Shmt2+/+ mice weaned onto either a folate-sufficient control diet (2 mg/kg folic acid; C) or a modified diet lacking folic acid (0 mg/kg folic acid) for 7 wk. Shmt2+/- and Shmt2+/+ mouse embryonic fibroblast (MEF) cells were cultured in defined culture medium containing either 0 or 25 nM folate (6S-5-formyl-tetrahydrofolate, folinate) to assess proliferative capacity and mitochondrial function. Chi-square tests, linear mixed models, and 2-factor ANOVA with Tukey post hoc analyses were used to analyze data.Results: Shmt2 +/- mice exhibited a 48%-67% reduction in SHMT2 protein concentrations in tissues. Interestingly, Shmt2+/- mice consuming the folate-sufficient C diet exhibited a 25% reduction in total folate in liver mitochondria. There was also a >20-fold increase in uracil in liver mtDNA in Shmt2+/- mice consuming the C diet, and dietary folate deficiency also increased uracil content in mouse liver mtDNA from both Shmt2+/+ and Shmt2+/- mice. Furthermore, decreased Shmt2 expression in MEF cells reduced cell proliferation, mitochondrial membrane potential, and oxygen consumption rate.Conclusions: This study demonstrates that Shmt2 heterozygosity and dietary folate deficiency impair mitochondrial dTMP synthesis in mice, as evidenced by the increased uracil in mtDNA. In addition, Shmt2 heterozygosity impairs mitochondrial function in MEF cells. These findings suggest that elevated uracil in mtDNA may impair mitochondrial function. [ABSTRACT FROM AUTHOR]

Background: Erythritol is both a common nonnutritive sweetener and an endogenous product of glucose metabolism. Recent reports suggest that elevated plasma erythritol is a predictive biomarker of cardiometabolic disease onset and complications.Objectives: Although short-term erythritol consumption has been evaluated, the effect of chronically elevated circulating erythritol on adiposity and glucose metabolism has not. This study investigated the effect of longer-term erythritol consumption on weight gain and glucose tolerance in young/adolescent mice.Methods: Four erythritol supplementation experiments were completed and analyzed separately in male C57BL/6J mice. In experiments 1 and 2, mice aged 8 wk or 20 wk, respectively, were randomly allocated to consume 16% fat diet (LFD) or LFD with 40 g/kg erythritol. In experiments 3 and 4, mice aged 8 wk or 20 wk were fed 45% fat diet (HFD) or HFD with 40 g/kg erythritol (HFD + ERY). In each experiment, we compared the effect of erythritol consumption on plasma erythritol, body weight and composition, glucose tolerance, and brown adipose tissue (BAT) uncoupling protein 1 (UCP1) expression. We also investigated relative endogenous tissue erythritol concentrations in a subset of control (LFD or HFD) mice in experiments 1 and 3.Results: There was no effect of erythritol supplementation on body weight or glucose tolerance in experiments 1-3. In experiment 4, in the 20-wk-old mice fed HFD or HFD + ERY, there was a significant interaction of time and erythritol on body weight (P < 0.0001), but the main effect of diet was not significant. Plasma erythritol was elevated 40-fold in mice consuming erythritol-supplemented diets relative to mice consuming LFD or HFD controls. We found no effect of chronic erythritol consumption on BAT UCP1 protein concentrations. Liver and kidney tissue contained significantly higher endogenous erythritol than quadriceps and visceral adipose (P < 0.001) in young mice fed LFD and HFD.Conclusions: In young/adolescent mice, prolonged erythritol consumption did not significantly affect body weight, composition, or glucose tolerance. [ABSTRACT FROM AUTHOR]

Folate, an essential nutrient found naturally in foods in a reduced form, is present in dietary supplements and fortified foods in an oxidized synthetic form (folic acid). There is widespread agreement that maintaining adequate folate status is critical to prevent diseases due to folate inadequacy (e.g. anemia, birth defects, and cancer). However, there are concerns of potential adverse effects of excess folic acid intake and/or elevated folate status, with the original concern focused on exacerbation of clinical effects of vitamin B-12 deficiency and its role in neurocognitive health. More recently, animal and observational studies have suggested potential adverse effects on cancer risk, birth outcomes, and other diseases. Observations indicating adverse effects from excess folic acid intake, elevated folate status, and unmetabolized folic acid (UMFA) remain inconclusive; the data do not provide the evidence needed to affect public health recommendations. Moreover, strong biological and mechanistic premises connecting elevated folic acid intake, UMFA, and/or high folate status to adverse health outcomes are lacking. However, the body of evidence on potential adverse health outcomes indicates the need for comprehensive research to clarify these issues and bridge knowledge gaps. Three key research questions encompass the additional research needed to establish whether high folic acid or total folate intake contributes to disease risk. 1) Does UMFA affect biological pathways leading to adverse health effects? 2) Does elevated folate status resulting from any form of folate intake affect vitamin B-12 function and its roles in sustaining health? 3) Does elevated folate intake, regardless of form, affect biological pathways leading to adverse health effects other than those linked to vitamin B-12 function? This article summarizes the proceedings of an August 2019 NIH expert workshop focused on addressing these research areas. [ABSTRACT FROM AUTHOR]

Background: Neural tube defects (NTDs) occur in nervous tissue during embryogenesis when the neural tube fails to close. Approximately 70% of all human NTDs can be prevented by folic acid (FA). Altered expression and/or function of the tumor suppressor protein p53 can lead to NTDs in mouse models.Objectives: The aim of this study was to determine if dietary FA could rescue p53-/--induced NTDs in mice, and to determine the effect loss of p53 has on pathways in folate 1-carbon metabolism.Methods: p53+/- female mice were randomly allocated and weaned onto either an FA-sufficient diet (2 mg/kg folic acid; +FA), or an FA-deficient diet (-FA). After 8 wk, the females were time-mated to p53-/- males. Embryos were examined at E12.5 for NTDs. Folate enzyme concentrations, nucleotide synthesis, uracil accumulation in DNA, and proliferation were measured in primary murine embryonic fibroblasts (MEFs). The "n - 1" chi-square test was used to compare NTD percentages, whereas all other data were analyzed by Student t test, except where noted a multilevel-fit model was used.Results: NTD rates of litters from dams consuming the +FA diet (20/46; 43%) did not differ from those of litters from dams consuming the -FA diet (14/35; 40%) (P > 0.05). p53-/- MEFs had 55% higher rates of folate-dependent de novo dTMP synthesis, a ∼2-fold higher accumulation of uracil in DNA, and a ∼30% higher rate of proliferation (P ≤ 0.05) than p53+/- MEFs independent of folate.Conclusions: p53-related NTDs are not FA responsive. Increased dTMP synthesis in p53-/- MEFs might not have been sufficient to meet the demands for thymidine triphosphate (dTTP) synthesis as evidenced by the elevated amounts of uracil in DNA. This study provides additional evidence that elevated uracil in DNA is a risk factor for NTDs. [ABSTRACT FROM AUTHOR]

Abstract Genomic instability is implicated in the etiology of several deleterious health outcomes including megaloblastic anemia, neural tube defects, and neurodegeneration. Uracil misincorporation and its repair are known to cause genomic instability by inducing DNA strand breaks leading to apoptosis, but there is emerging evidence that uracil incorporation may also result in broader modifications of gene expression, including: changes in transcriptional stalling, strand break-mediated transcriptional upregulation, and direct promoter inhibition. The factors that influence uracil levels in DNA are cytosine deamination, de novo thymidylate (dTMP) biosynthesis, salvage dTMP biosynthesis, dUTPase, and DNA repair. There is evidence that the nuclear localization of the enzymes in these pathways in mammalian cells may modify and/or control the levels of uracil accumulation into nuclear DNA. Uracil sequencing technologies demonstrate that uracil in DNA is not distributed stochastically across the genome, but instead shows patterns of enrichment. Nuclear localization of the enzymes that modify uracil in DNA may serve to change these patterns of enrichment in a tissue-specific manner, and thereby signal the genome in response to metabolic and/or nutritional state of the cell. [ABSTRACT FROM AUTHOR]

Public trust in nutrition science is the foundation on which nutrition and health progress is based, including sound public health. An ASN-commissioned, independent Advisory Committee comprehensively reviewed the literature and available public surveys about the public's trust in nutrition science and the factors that influence it and conducted stakeholder outreach regarding publicly available information. The Committee selected 7 overlapping domains projected to significantly influence public trust: 1) conflict of interest and objectivity; 2) public benefit; 3) standards of scientific rigor and reproducibility; 4) transparency; 5) equity; 6) information dissemination (education, communication, and marketing); and 7) accountability. The literature review comprehensively explored current practices and threats to public trust in nutrition science, including gaps that erode trust. Unfortunately, there is a paucity of peer-reviewed material specifically focused on nutrition science. Available material was examined, and its analysis informed the development of priority best practices. The Committee proposed best practices to support public trust, appropriate to ASN and other food and nutrition organizations motivated by the conviction that public trust remains key to the realization of the benefits of past, present, and future scientific advances. The adoption of the best practices by food and nutrition organizations, such as ASN, other stakeholder organizations, researchers, food and nutrition professionals, companies, government officials, and individuals working in the food and nutrition space would strengthen and help ensure earning and keeping the public's continued trust in nutrition science [ABSTRACT FROM AUTHOR]

The article comments on a study on riboflavin deficiency and vitamin B-6 status. It identifies strengths of the study including its use of erythrocyte glutathione reductase activation (EGRac), inclusion of data on dietary intake and riboflavin marker and determination of the effect of methylenetetrahydrofolate reductase (MTHFR) C677T polymorphism on riboflavin requirements. Also noted is the significance of the study to defining nutrient reference intake.

Background: Formate provides one-carbon units for de novo purine and thymidylate (dTMP) synthesis and is produced via both folate-dependent and folate-independent pathways. Folate-independent pathways are mediated by cytosolic alcohol dehydrogenase 5 (ADH5) and mitochondrial aldehyde dehydrogenase 2 (ALDH2), which generate formate by oxidizing formaldehyde. Formate is a potential biomarker of B-vitamin-dependent one-carbon metabolism.Objective: This study investigated the contributions of ADH5 and ALDH2 to formate production and folate-dependent de novo purine and dTMP synthesis in HepG2 cells.Methods:ADH5 knockout and ALDH2 knockdown HepG2 cells were cultured in folate-deficient [0 nM (6S) 5-formyltetrahydrofolate] or folate-sufficient [25 nM (6S) 5-formyltetrahydrofolate] medium. Purine biosynthesis was quantified as the ratio of [14C]-formate to [3H]-hypoxanthine incorporated into genomic DNA, which indicates the contribution of the de novo purine synthesis pathway relative to salvage synthesis. dTMP synthesis was quantified as the ratio of [14C]-deoxyuridine to [3H]-thymidine incorporation into genomic DNA, which indicates the capacity of de novo dTMP synthesis relative to salvage synthesis.Results: The [14C]-formate-to-[3H]-hypoxanthine ratio was greater in ADH5 knockout than in wild-type HepG2 cells, under conditions of both folate deficiency (+30%; P < 0.001) and folate sufficiency (+22%; P = 0.02). These data indicate that ADH5 deficiency increases the use of exogenous formate for de novo purine biosynthesis. The [14C]-deoxyuridine-to-[3H]-thymidine ratio did not differ between ADH5 knockout and wild-type cells, indicating that ADH5 deficiency does not affect de novo dTMP synthesis capacity relative to salvage synthesis. Under folate deficiency, ALDH2 knockdown cells exhibited a 37% lower ratio of [14C]-formate to [3H]-hypoxanthine (P < 0.001) compared with wild-type HepG2 cells, indicating decreased use of exogenous formate, or increased endogenous formate synthesis, for de novo purine biosynthesis.Conclusions: In HepG2 cells, ADH5 is a source of formate for de novo purine biosynthesis, especially during folate deficiency when folate-dependent formate production is limited. Formate is also shown to be limiting in the growth of HepG2 cells. [ABSTRACT FROM AUTHOR]

Disruptions in folate-mediated one-carbon metabolism (FOCM) are associated with risk for several pathologies including developmental anomalies such as neural tube defects and congenital heart defects, diseases of aging including cognitive decline, neurodegeneration and epithelial cancers, and hematopoietic disorders including megaloblastic anemia. However, the causal pathways and mechanisms that underlie these pathologies remain unresolved. Because folate-dependent anabolic pathways are tightly interconnected and best described as a metabolic network, the identification of causal pathways and associated mechanisms of pathophysiology remains a major challenge in identifying the contribution of individual pathways to disease phenotypes. Investigations of genetic mouse models and human inborn errors of metabolism enable a more precise dissection of the pathways that constitute the FOCM network and enable elucidation of causal pathways associated with NTDs. In this overview, we summarize recent evidence that the enzyme MTHFD1 plays an essential role in FOCM in humans and in mice, and that it determines the partitioning of folate-activated one carbon units between the folate-dependent de novo thymidylate and homocysteine remethylation pathways through its regulated nuclear localization. We demonstrate that impairments in MTHFD1 activity compromise both homocysteine remethylation and de novo thymidylate biosynthesis, and provide evidence that MTHFD1-associated disruptions in de novo thymidylate biosynthesis lead to genome instability that may underlie folate-associated immunodeficiency and birth defects. [ABSTRACT FROM AUTHOR]

Background: Folic acid prevents neural tube closure defects (NTDs), but the causal metabolic pathways have not been established. Serine hydroxymethyltransferase 1 (SHMT1) is an essential scaffold protein in folate-dependent de novo thymidylate synthesis in the nucleus. SHMT1-deficient mice provide a model to investigate folic acid-responsive NTDs wherein disruption of de novo thymidylate synthesis impairs neural tube closure. Objective: We examined the effects of maternal supplementation with the pyrimidine nucleosides uridine, thymidine, or deoxyuridine with and without folate deficiency on NTD incidence in the Shmt1 mouse model. Design: Shmt1+y+ and Shmt1~y~ female mice fed folate-replete or folate-deficient diets and supplemented with uridine, thymidine, or deoxyuridine were bred, and litters (n = 10-23 per group) were examined for the presence of NTDs. Biomarkers of impaired folate status and metabolism were measured, including plasma nucleosides, hepatic uracil content, maternal plasma folate concentrations, and incorporation of nucleoside precursors into DNA. Results: Shmt1+I~ and Shmt1~'~ embryos from dams fed the folatedeficient diet were susceptible to NTDs. No NTDs were observed in litters from dams fed the folate-deficient diet supplemented with deoxyuridine. Surprisingly, uridine supplementation increased NTD incidence, independent of embryo genotype and dietary folic acid. These dietary nucleosides did not affect maternal hepatic uracil accumulation in DNA but did affect plasma folate concentrations. Conclusions: Maternal deoxyuridine supplementation prevented NTDs in dams fed the folate-deficient diet, whereas maternal uridine supplementation increased NTD incidence, independent of folate and embryo genotype. These findings provide new insights into the metabolic impairments and mechanisms of folate-responsive NTDs resulting from decreased Shmt1 expression. Am J Clin Nutr 2015;101:860-9. [ABSTRACT FROM AUTHOR]

Folate-mediated one-carbon metabolism is a metabolic network of interconnected pathways that is required for the de novo synthesis of three of the fourDNAbases and the remethylation of homocysteine to methionine. Previous studies have indicated that the thymidylate synthesis and homocysteine remethylation pathways compete for a limiting pool of methylenetetrahydrofolate cofactors and that thymidylate biosynthesis is preserved in folate deficiency at the expense of homocysteine remethylation, but the mechanisms are unknown. Recently, it was shown that thymidylate synthesis occurs in the nucleus, whereas homocysteine remethylation occurs in the cytosol. In this study we demonstrate that methylenetetrahydrofolate dehydrogenase 1 (MTHFD1), an enzyme that generates methylenetetrahydrofolate from formate, ATP, and NADPH, functions in the nucleus to support de novo thymidylate biosynthesis. MTHFD1 translocates to the nucleus in S-phase MCF-7 and HeLa cells. During folate deficiency mouse liver MTHFD1levels are enriched in the nucleus>2-fold at the expense of levels in the cytosol. Furthermore, nuclear folate levels are resistant to folate depletion when total cellular folate levels are reduced by >50% in mouse liver. The enrichment of folate cofactors and MTHFD1protein in the nucleus during folate deficiency in mouse liver and human cell lines accounts for previous metabolic studies that indicated 5,10-methylenetetrahydrofolate is preferentially directed toward de novo thymidylate biosynthesis at the expense of homocysteine remethylation during folate deficiency. [ABSTRACT FROM AUTHOR]

Impaired folate-mediated one-carbon metabolism (OCM) has emerged as a risk factor for several diseases associated with age-related cognitive decline, but the underlying mechanisms remain unknown and thus hinder the identification of subpopulations most vulnerable to OCM disruption. Here we investigated the role of serine hydroxymethyltransferase 1 (SHMT1), a folate-dependent enzyme regulating de novo thymidylate biosynthesis, in influencing neuronal and cognitive function in the adult mouse. We observed Shmt1 expression in the hippocampus, including the granule cell layer of the dentate gyrus (DG), and examined hippocampal neurogenesis and hippocampal-dependent fear conditioning in mice deficient for Shmt1. We used a 3 x 3 design in which adult male Shmt1+/+, Shmt1+/-, and Shmt1-/- mice were fed folic acid control (2 mg/kg), folic acid-deficient (0 mg/kg), or folic acid-supplemented (8 mg/kg) diets from weaning through the duration of the study. Proliferation within the DG was elevated by 70% in Shmt1+/- mice, yet the number of newborn mature neurons was reduced by 98% compared with that in Shmt1+/+ mice. Concomitant with these alterations, Shmt1+/- mice showed a 45% reduction in mnemonic recall during trace fear conditioning. Dietary folate manipulations alone did not influence neural outcomes. Together, these data identify SHMT1 as one of the first enzymes within the OCM pathway to regulate neuronal and cognitive profiles and implicate impaired thymidylate biosynthesis in the etiology of folate-related neuropathogenesis. [ABSTRACT FROM AUTHOR]

Impaired utilization of folate is caused by insufficient dietary intake and/or genetic variation and has been shown to prompt changes in related pathways, including choline and methionine metabolism. These pathways have been shown to be sensitive to variation within the Mthfd1 gene, which codes for a folate-metabolizing enzyme responsible for generating 1-carbon (1-C)--substituted folate derivatives. The Mthfd1gt/+ mouse serves as a potential model of human Mthfd1 loss-of-function genetic variants that impair MTHFD1 function. This study investigated the effects of the Mthfd1gt/+ genotype and folate intake on markers of choline, folate, methionine, and transsulfuration metabolism. Male Mthfd1gt/+ and Mthfd1+/+ mice were randomly assigned at weaning (3 wk of age) to either a control (2 mg/kg folic acid) or folate-deficient (0 mg/kg folic acid) diet for 5 wk. Mice were killed at 8 wk of age following 12 h of food deprivation; blood and liver samples were analyzed for choline, methionine, and transsulfuration biomarkers. Independent of folate intake, mice with the Mthfd1gt/+ genotype had higher hepatic concentrations of choline (P = 0.005), betaine (P= 0.013), and dimethylglycine (P = 0.004) and lower hepatic concentrations of glycerophosphocholine (P = 0.002) relative to Mthfd1+/+ mice. Mthfd1gt/+ mice also had higher plasma concentrations of homocysteine (P = 0.0016) and cysteine (P < 0.001) as well as lower plasma concentrations of methionine (P = 0.0003) and cystathionine (P = 0.011). The metabolic alterations observed in Mthfd1gt/+ mice indicate perturbed choline and folate-dependent 1 -C metabolism and support the future use of Mthfd1gt/+ mice as a tool to investigate the impact of impaired 1-C metabolism on disease outcomes. [ABSTRACT FROM AUTHOR]

10-Formyltetrahydrofolate dehydrogenase (FDH) catalyzes the NADP+-dependent conversion of 10-formyltetrahydrofolate to CO2 and tetrahydrofolate (THF) and is an abundant high affinity folate-binding protein. Although several activities have been ascribed to FDH, its metabolic role in folate-mediated one-carbon metabolism is not well understood. FDH has been proposed to: 1) inhibit purine biosynthesis by depleting 10-formyl-THF pools, 2) maintain cellular folate concentrations by sequestering THF, 3) deplete the supply of folate- activated one-carbon units, and 4) stimulate the generation of THE- activated one-carbon unit synthesis by channeling folate cofactors to other folate-dependent enzymes. The metabolic functions of FDH were investigated in neuroblastoma, which do not contain detectable levels of FDH. Both low and high FDH expression reduced total cellular folate concentrations by 60%, elevated rates of folate catabolism, and depleted cellular 5-methyl-THE and S-adenosylmethionine levels. Low FDH expression increased the formyl-THF/THF ratio nearly 10-fold, whereas THF accounted for nearly 50% of total folate in neuroblastoma with high FDH expression. FDH expression did not affect the enrichment of exogenous formate into methionine, serine, or purines and did not suppress de novo purine nucleotide biosynthesis. We conclude that low FDH expression facilitates the incorporation of one-carbon units into the one-carbon pool, whereas high levels of FDH expression deplete the folate-activated one-carbon pool by catalyzing the conversion of 10-formyl-THF to THF. Furthermore, FDH does not increase cellular folate concentrations by sequestering THE in neuroblastoma nor does it inhibit or regulate de novo purine biosynthesis. FDH expression does deplete cellular 5-methyl-THF and S-adenosyl-methionine levels indicating that FDH impairs the folate-dependent homocysteine remethylation cycle. [ABSTRACT FROM AUTHOR]

Skeletal muscle regeneration relies on muscle-specific adult stem cells (MuSCs), MuSC progeny, muscle progenitor cells (MPCs), and a coordinated myogenic program that is influenced by the extracellular environment. Following injury, MPCs undergo a transient and rapid period of population expansion, which is necessary to repair damaged myofibers and restore muscle homeostasis. Certain pathologies (e.g., metabolic diseases and muscle dystrophies) and advanced age are associated with dysregulated muscle regeneration. The availability of serine and glycine, two nutritionally non-essential amino acids, is altered in humans with these pathologies, and these amino acids have been shown to influence the proliferative state of non-muscle cells. Our objective was to determine the role of serine/glycine in MuSC/MPC function. Primary human MPCs (h MPCs) were used for in vitro experiments, and young (4–6 mo) and old (>20 mo) mice were used for in vivo experiments. Serine/glycine availability was manipulated using specially formulated media in vitro or dietary restriction in vivo followed by downstream metabolic and cell proliferation analyses. We identified that serine/glycine are essential for h MPC proliferation. Dietary restriction of serine/glycine in a mouse model of skeletal muscle regeneration lowered the abundance of MuSCs 3 days post-injury. Stable isotope-tracing studies showed that h MPCs rely on extracellular serine/glycine for population expansion because they exhibit a limited capacity for de novo serine/glycine biosynthesis. Restriction of serine/glycine to h MPCs resulted in cell cycle arrest in G0/G1. Extracellular serine/glycine was necessary to support glutathione and global protein synthesis in h MPCs. Using an aged mouse model, we found that reduced serine/glycine availability augmented intermyocellular adipocytes 28 days post-injury. These studies demonstrated that despite an absolute serine/glycine requirement for MuSC/MPC proliferation, de novo synthesis was inadequate to support these demands, making extracellular serine and glycine conditionally essential for efficient skeletal muscle regeneration. • Extracellular serine and glycine are necessary for muscle stem/progenitor cell population expansion. • Human muscle progenitor cells possess limited capacity for de novo serine/glycine biosynthesis. • Extracellular serine/glycine restriction increases reactive oxygen species and reduces intracellular glutathione. • Extracellular serine/glycine inhibits protein synthesis and cell cycle arrest. • Reduced extracellular serine and glycine in old mice impairs skeletal muscle regeneration. [ABSTRACT FROM AUTHOR]

Dietary reference intakes (DRIs) are quantitative, nutrient intake-based standards used for assessing the diets and specific nutrient intakes of healthy individuals and populations and for informing national nutrition policy and nutrition programs. Because nutrition needs vary by age, sex, and physiological state, DRIs are often specified for healthy subgroups within a population. Diet is known to be the leading modifiable risk factor for chronic disease, and the prevalence of chronic disease is growing in all populations globally and across all subgroups, but especially in older adults. It is known that nutrient needs can change in some chronic disease and other clinical states. Disease states and/or disease treatment can cause whole-body or tissue-specific nutrient depletion or excess, resulting in the need for altered nutrient intakes. In other cases, disease-related biochemical dysfunction can result in a requirement for a nonessential nutrient, rendering it as conditionally essential, or result in toxicity for a food component at levels usually tolerated by healthy people, as seen in inborn errors of metabolism. Here we summarize examples from a growing body of literature of disease-altering nutrient requirements, supporting the need to give more consideration to special nutrient requirements in disease states. [ABSTRACT FROM AUTHOR]

The author discusses guidelines released by the World Health Organization (WHO ) regarding the implementation, monitoring, and impact evaluation of interventions for preventing neural tube defects (NTDs). Topics include the factors that interact to determine folate status in humans such as dietary intake of folate and a polymorphism in the gen encoding the enzyme methylenetetrahydrofolate reductase (MTHFR), and the use of folic acid supplements to prevent NTDs.

Mitochondrial inner membrane protein MPV17 is a protein of unknown function that is associated with mitochondrialDNA (mtDNA)-depletion syndrome (MDS). MPV17 loss-of-function has been reported to result in tissue-specific nucleotide pool imbalances, which can occur in states of perturbed folate-mediated one-carbon metabolism (FOCM), but MPV17 has not been directly linked to FOCM. FOCM is a metabolic network that provides one-carbon units for the de novo synthesis of purine and thymidylate nucleotides (e.g. dTMP) for both nuclear DNA (nuDNA) and mtDNA replication. In this study, we investigated the impact of reduced MPV17 expression on markers of impaired FOCM in HeLa cells. Depressed MPV17 expression reduced mitochondrial folate levels by 43% and increased uracil levels, a marker of impaired dTMP synthesis, in mtDNA by 3-fold. The capacity of mitochondrial de novo and salvage pathway dTMP biosynthesis was unchanged by the reduced MPV17 expression, but the elevated levels of uracil inmtDNAsuggested that other sources of mitochondrial dTMP are compromised in MPV17-deficient cells. These results indicate that MPV17 provides a third dTMP source, potentially by serving as a transporter that transfers dTMP from the cytosol to mitochondria to sustain mtDNA synthesis. We propose that MPV17 loss-offunction and related hepatocerebralMDSare linked to impaired FOCMin mitochondria by providing insufficient access to cytosolic dTMP pools and by severely reducing mitochondrial folate pools. [ABSTRACT FROM AUTHOR]

Mammals require essential nutrients from dietary sources to support normal metabolic, physiological and neuronal functions, to prevent diseases of nutritional deficiency as well as to prevent chronic disease. Disease and/or its treatment can modify fundamental biological processes including cellular nutrient accretion, stability and function in cells. These effects can be isolated to a specific diseased organ in the absence of whole-body alterations in nutrient status or biochemistry. Loss of blood–brain barrier function, which occurs in in-born errors of metabolism and in chronic disease, can cause brain-specific folate deficiency and contribute to disease co-morbidity. The role of brain folate deficiency in neuropsychiatric disorders is reviewed, as well as emerging diagnostic and nutritional strategies to identify and address brain folate deficiency in blood–brain barrier dysfunction. [ABSTRACT FROM AUTHOR]

Thymidylate (dTMP) biosynthesis plays an essential and exclusive function in DNA synthesis and proper cell division, and therefore has been an attractive therapeutic target. Folate analogs, known as antifolates, and nucleotide analogs that inhibit the enzymatic action of the de novo thymidylate biosynthesis pathway and are commonly used in cancer treatment. In this review, we examine the mechanisms by which the antifolate 5-fluorouracil, as well as other dTMP synthesis inhibitors, function in cancer treatment in light of emerging evidence that dTMP synthesis occurs in the nucleus. Nuclear localization of the de novo dTMP synthesis pathway requires modification of the pathway enzymes by the small ubiquitin-like modifier (SUMO) protein. SUMOylation is required for nuclear localization of the de novo dTMP biosynthesis pathway, and disruption in the SUMO pathway inhibits cell proliferation in several cancer models. We summarize evidence that the nuclear localization of the dTMP biosynthesis pathway is a critical factor in the efficacy of antifolate-based therapies that target dTMP synthesis. [ABSTRACT FROM AUTHOR]

Folate is an essential B vitamin required for the de novo synthesis of purines, thymidylate and methionine. Folate deficiency can lead to mutations and genome instability, and has been shown to exacerbate the genotoxic potential of environmental toxins. We hypothesized that a folic acid (FA) deficient diet would induce genotoxicity in mice as measured by the Pig-a mutant phenotype (CD24-) and micronuclei (MN) in reticulocytes (RET) and red blood cells/normochromatic erythrocytes (RBC/NCE). Male Balb/c mice were fed a FA deficient (0 mg/kg), control (2 mg/kg) or supplemented (6 mg/kg) diet from weaning for 18 wk. Mice fed the deficient diet had 70% lower liver folate ( p < 0.001), 1.8 fold higher MN-RET ( p < 0.001), and 1.5 fold higher MN-NCE ( p < 0.001) than mice fed the control diet. RET CD24− and RBC CD24− frequencies were not different between mice fed the deficient and control diets. Compared to mice fed the FA supplemented diet, mice fed the deficient diet had 73% lower liver folate ( p < 0.001), a 2.0 fold increase in MN-RET ( p < 0.001), a 1.6 fold increase in MN-NCE ( p < 0.001) and 3.8 fold increase in RBC CD24− frequency ( p = 0.011). RET CD24− frequency did not differ between mice fed the deficient and supplemented diets. Our data suggest that FA adequacy protects against mutagenesis at the Pig-a locus and MN induction in the red blood cells of mice. [ABSTRACT FROM AUTHOR]

The low-calorie sweetener erythritol is endogenously produced from glucose through the pentose phosphate pathway in humans. Erythritol is of medical interest because elevated plasma levels of this polyol are predictive for visceral adiposity gain and development of type 2 diabetes. However, the mechanisms behind these associations remain unknown because the erythritol biosynthesis pathway, particularly the enzyme catalyzing the final step of erythritol synthesis (reduction of erythrose to erythritol), is not characterized. In this study, we purified two enzymes from rabbit liver capable of catalyzing the conversion of erythrose to erythritol: alcohol dehydrogenase 1 (ADH1) and sorbitol dehydrogenase (SORD). Both recombinant human ADH1 and SORD reduce erythrose to erythritol, using NADPH as a co-factor, and cell culture studies indicate that this activity is primarily NADPH-dependent. We found that ADH1 variants vary markedly in both their affinity for erythrose and their catalytic capacity (turnover number). Interestingly, the recombinant protein produced from the ADH1B2 variant, common in Asian populations, is not active when NADPH is used as a co-factor in vitro. We also confirmed SORD contributes to intracellular erythritol production in human A549 lung cancer cells, where ADH1 is minimally expressed. In summary, human ADH1 and SORD catalyze the conversion of erythrose to erythritol, pointing to novel roles for two dehydrogenase proteins in human glucose metabolism that may contribute to individual responses to diet. Proteomics data are available via ProteomeXchange with identifier PXD015178. [ABSTRACT FROM AUTHOR]