MTHFD1 controls DNA methylation in Arabidopsis.

DNA methylation is an epigenetic mechanism that has important functions in transcriptional silencing and is associated with repressive histone methylation (H3K9me). To further investigate silencing mechanisms, we screened a mutagenized Arabidopsis thaliana population for expression of SDCpro-GFP, redundantly controlled by DNA methyltransferases DRM2 and CMT3. Here, we identify the hypomorphic mutant mthfd1-1, carrying a mutation (R175Q) in the cytoplasmic bifunctional methylenetetrahydrofolate dehydrogenase/methenyltetrahydrofolate cyclohydrolase (MTHFD1). Decreased levels of oxidized tetrahydrofolates in mthfd1-1 and lethality of loss-of-function demonstrate the essential enzymatic role of MTHFD1 in Arabidopsis. Accumulation of homocysteine and S-adenosylhomocysteine, genome-wide DNA hypomethylation, loss of H3K9me and transposon derepression indicate that S-adenosylmethionine-dependent transmethylation is inhibited in mthfd1-1. Comparative analysis of DNA methylation revealed that the CMT3 and CMT2 pathways involving positive feedback with H3K9me are mostly affected. Our work highlights the sensitivity of epigenetic networks to one-carbon metabolism due to their common S-adenosylmethionine-dependent transmethylation and has implications for human MTHFD1-associated diseases

Effects of Substrate Salinity and Nutrient Levels on Physiological Response, Yield, and Fruit Quality of Habanero Pepper

Although habanero peppers (Capsicum chinense, Jacq.) are highly appreciated as a result of their organoleptic and pungency properties, the crop faces edaphic stresses throughout Mexico. A study was conducted to determine how the photosynthetic parameters, vegetative growth, yield, and fruit quality of the plant change in response to suboptimal conditions in the substrate. Habanero plants were grown in an inert substrate (perlite) and exposed to increased salinity levels (4 and 7 dS·m−1) and reduced nitrogen and phosphorus conditions. Plants grown with a Hoagland-based solution were used as controls. High salinity conditions reduced the light-saturated photosynthetic rates (64% of the control) but did not compromise yield or fruit quality. This effect was possibly the result of the addition of Ca2+, which reduced salinity-induced calcium deficiency. Although comparable low nitrogen levels in previous studies were shown to cause a severe reduction in plant viability, in our study, low nitrogen reduced the light-saturated photosynthetic rates (47% of the control) and shoot:root ratio (67% of the control) but did not significantly affect yield or fruit quality. Low nitrogen and 7-dS·m−1\ua0treatments increased fructose and glucose content (increases of 27% and 21%, respectively). Low phosphorus significantly affected plant growth and yield and reduced fructose content (73% of the control). Plants were not sensitive to low nitrogen and high salinity, possibly as a result of the use of nitrate-based fertilizers and the addition of calcium, respectively. These results provide guidelines for habanero pepper production under suboptimal edaphic conditions

Quantification of folate in food using deconjugase of plant origin combined with LC-MS/MS: A method comparison of a large and diverse sample set

A round robin comparison was performed in order to test the performance of a recently developed LC-MS/MS method for quantification of 6 folate forms. Eighty-nine samples representing the food groups of fruits, vegetables, legumes, cereals, dairy products, meat, and offal were analyzed by two LC-MS/MS methods and a microbiological assay (MA). A plant-origin deconjugase enzyme (Arabidopsis thaliana) for deconjugation of folates (PE-LC-MS/MS), or animal-origin deconjugase (rat serum and chicken pancreas) (AE-LC-MS/MS) was used in the LC-MS/MS methods, each in a single enzymatic step. In contrast, the MA involved tri-enzyme extraction including human plasma as a deconjugase. A significant bias of 17% lower and 25% higher results was found when PE-LC-MS/MS was compared to MA and AE-LC-MS/MS, respectively. The PE-LC-MS/MS provides fast quantification of various folate vitamers and total folate content, which could be a proper substitute to the currently standardized but imprecise and time-consuming microbiological assay in the future

Avocado fruit maturation and ripening: dynamics of aliphatic acetogenins and lipidomic profiles from mesocarp, idioblasts and seed

Abstract Background Avocado fruit contains aliphatic acetogenins (oft-acetylated, odd-chain fatty alcohols) with promising bioactivities for both medical and food industries. However, we have scarce knowledge about their metabolism. The present work aimed to study changes in acetogenin profiles from mesocarp, lipid-containing idioblasts, and seeds from ‘Hass’ cultivar during fruit development, germination, and three harvesting years. An untargeted LC-MS based lipidomic analysis was also conducted to profile the lipidome of avocado fruit in each tissue. Results The targeted analysis showed that acetogenin profiles and contents remained unchanged in avocado mesocarp during maturation and postharvest ripening, germination, and different harvesting years. However, a shift in the acetogenin profile distribution, accompanied with a sharp increase in concentration, was observed in seed during early maturation. Untargeted lipidomics showed that this shift was accompanied with remodeling of glycerolipids: TAGs and DAGs decreased during fruit growing in seed. Remarkably, the majority of the lipidome in mature seed was composed by acetogenins; we suggest that this tissue is able to synthesize them independently from mesocarp. On the other hand, lipid-containing idioblasts accumulated almost the entire acetogenin pool measured in the whole mesocarp, while only having 4% of the total fatty acids. The lipidome of this cell type changed the most when the fruit was ripening after harvesting, TAGs decreased while odd-chain DAGs increased. Notably, idioblast lipidome was more diverse than that from mesocarp. Conclusions Evidence shown here suggests that idioblasts are the main site of acetogenin biosynthesis in avocado mesocarp. This work unveiled the prevalence of aliphatic acetogenins in the avocado fruit lipidome and evidenced TAGs as initial donors of the acetogenin backbones in its biosynthesis. It also sets evidence for acetogenins being included in future works aimed at characterizing the avocado seed, as they are a main component of their lipidome

Insights into Drivers of Liking for Avocado Pulp (Persea americana): Integration of Descriptive Variables and Predictive Modeling

Trends in new food products focus on low-carbohydrate ingredients rich in healthy fats, proteins, and micronutrients; thus, avocado has gained worldwide attention. This study aimed to use predictive modeling to identify the potential sensory drivers of liking for avocado pulp by evaluating acceptability scores and sensory descriptive profiles of two commercial and five non-commercial cultivars. Macronutrient composition, instrumental texture, and color were also characterized. Trained panelists performed a descriptive profile of nineteen sensory attributes. Affective data from frequent avocado adult consumers (n = 116) were collected for predictive modeling of an external preference map (R2 = 0.98), which provided insight into sensory descriptors that drove preference for particular avocado pulps. The descriptive map explained 67.6% of the variance in sensory profiles. Most accepted pulps were from Hass and Colin V-33; the latter had sweet and green flavor notes. Descriptive flavor attributes related to liking were global impact, oily, and creamy. Sensory drivers of texture liking included creamy/oily, lipid residue, firmness, and cohesiveness. Instrumental stickiness was disliked and inversely correlated to dry-matter and lipids (r = −0.87 and −0.79, respectively). Color differences (∆Eab*) also contributed to dislike. Sensory-guided selection of avocado fruits and ingredients can develop products with high acceptability in breeding and industrialization strategies

In Vivo Rate of Formaldehyde Condensation with Tetrahydrofolate

Formaldehyde is a highly reactive compound that participates in multiple spontaneous reactions, but these are mostly deleterious and damage cellular components. In contrast, the spontaneous condensation of formaldehyde with tetrahydrofolate (THF) has been proposed to contribute to the assimilation of this intermediate during growth on C1 carbon sources such as methanol. However, the in vivo rate of this condensation reaction is unknown and its possible contribution to growth remains elusive. Here, we used microbial platforms to assess the rate of this condensation in the cellular environment. We constructed Escherichia coli strains lacking the enzymes that naturally produce 5,10-methylene-THF. These strains were able to grow on minimal medium only when equipped with a sarcosine (N-methyl-glycine) oxidation pathway that sustained a high cellular concentration of formaldehyde, which spontaneously reacts with THF to produce 5,10-methylene-THF. We used flux balance analysis to derive the rate of the spontaneous condensation from the observed growth rate. According to this, we calculated that a microorganism obtaining its entire biomass via the spontaneous condensation of formaldehyde with THF would have a doubling time of more than three weeks. Hence, this spontaneous reaction is unlikely to serve as an effective route for formaldehyde assimilation

The Folylpolyglutamate Synthetase Plastidial Isoform Is Required for Postembryonic Root Development in Arabidopsis1[W][OA]

A recessive Arabidopsis (Arabidopsis thaliana) mutant with short primary roots and root hairs was identified from a forward genetic screen. The disrupted gene in the mutant encoded the plastidial isoform of folylpolyglutamate synthetase (FPGS), previously designated as AtDFB, an enzyme that catalyzes the addition of glutamate residues to the folate molecule to form folylpolyglutamates. The short primary root of atdfb was associated with a disorganized quiescent center, dissipated auxin gradient in the root cap, bundled actin cytoskeleton, and reduced cell division and expansion. The accumulation of monoglutamylated forms of some folate classes in atdfb was consistent with impaired FPGS function. The observed cellular defects in roots of atdfb underscore the essential role of folylpolyglutamates in the highly compartmentalized one-carbon transfer reactions (C1 metabolism) that lead to the biosynthesis of compounds required for metabolically active cells found in the growing root apex. Indeed, metabolic profiling uncovered a depletion of several amino acids and nucleotides in atdfb indicative of broad alterations in metabolism. Methionine and purines, which are synthesized de novo in plastids via C1 enzymatic reactions, were particularly depleted. The root growth and quiescent center defects of atdfb were rescued by exogenous application of 5-formyl-tetrahydrofolate, a stable folate that was readily converted to metabolically active folates. Collectively, our results indicate that AtDFB is the predominant FPGS isoform that generates polyglutamylated folate cofactors to support C1 metabolism required for meristem maintenance and cell expansion during postembryonic root development in Arabidopsis

Chemical Profile and Safety Assessment of a Food-Grade Acetogenin-Enriched Antimicrobial Extract from Avocado Seed.

Acetogenins are bioactive fatty acid derivatives found in avocado tissues. Their efficacy as antimicrobials has been documented and initiated interest to use them as replacements of synthetic food additives. The present work focused on evaluation of multiple analytical methodologies for detection and quantification of organic solids present in a food-grade acetogenin-enriched extract (Avosafe®), and on its safety evaluations using bacterial reverse mutation (AMES) tests and acute oral toxicity to rat assays. Results confirmed chemical structures of two acetogenins as present in Avosafe® (AcO-avocadyne-(0) and AcO-avocadiene B-(3)), and together with seven other previously known compounds, quantified 94.74 ± 5.77% w/w of its solids as acetogenins. Safety evaluations indicated that Avosafe® was non-mutagenic and had an acute median lethal oral dose (LD50) to rats higher than the maximum concentration tested (>2000 mg·kg-1), with no signs of macroscopic abnormalities in organs. Mean body weight and hematological and biochemical parameters were normal after 14 days of a single oral dose of 2000 mg·kg-1. The results advance scientific information on the safety of avocado seed acetogenins and also generate new knowledge on profiles and concentrations of individual acetogenins found in avocado tissues (seed, pulp, and leaves) and in Avosafe®

Chemical Profile and Safety Assessment of a Food-Grade Acetogenin-Enriched Antimicrobial Extract from Avocado Seed

Acetogenins are bioactive fatty acid derivatives found in avocado tissues. Their efficacy as antimicrobials has been documented and initiated interest to use them as replacements of synthetic food additives. The present work focused on evaluation of multiple analytical methodologies for detection and quantification of organic solids present in a food-grade acetogenin-enriched extract (Avosafe®), and on its safety evaluations using bacterial reverse mutation (AMES) tests and acute oral toxicity to rat assays. Results confirmed chemical structures of two acetogenins as present in Avosafe® (AcO-avocadyne-(0) and AcO-avocadiene B-(3)), and together with seven other previously known compounds, quantified 94.74 ± 5.77% w/w of its solids as acetogenins. Safety evaluations indicated that Avosafe® was non-mutagenic and had an acute median lethal oral dose (LD50) to rats higher than the maximum concentration tested (>2000 mg·kg−1), with no signs of macroscopic abnormalities in organs. Mean body weight and hematological and biochemical parameters were normal after 14 days of a single oral dose of 2000 mg·kg−1. The results advance scientific information on the safety of avocado seed acetogenins and also generate new knowledge on profiles and concentrations of individual acetogenins found in avocado tissues (seed, pulp, and leaves) and in Avosafe®

The plastidial folylpolyglutamate synthetase and root apical meristem maintenance

Folylpolyglutamate synthetase (FPGS) catalyzes the attachment of glutamate residues to the folate molecule in plants. Three isoforms of FPGS have been identified in Arabidopsis and these are localized in the plastid (AtDFB), mitochondria (AtDFC) and cytosol (AtDFD). We recently determined that mutants in the AtDFB (At5G05980) gene disrupt primary root development in Arabidopsis thaliana seedlings. Transient expression of AtDFB-green fluorescent protein (GFP) fusion under the control of the native AtDFB promoter in Nicotiana tabacum leaf epidermal cells verified the plastid localization of AtDFB. Furthermore, low concentrations of methotrexate (MTX), a compound commonly used as a folate antagonist in plant and mammalian cells induced primary root defects in wild type seedlings that were similar to atdfb. In addition, atdfb seedlings were more sensitive to MTX when compared to wild type. Quantitative (q) RT-PCR showed lower transcript levels of the mitochondrial and cytosolic FPGS in roots of 7-day-old atdfb seedling suggesting feedback regulation of AtDFB on the expression of other FPGS isoforms during early seedling development. The primary root defects of atdfb, which can be traced in part to altered quiescent center (QC) identity, pave the way for future studies that could link cell type specific folate and FPGS isoform requirements to whole organ development