Folate synthesis and salvage pathways are relatively well known from classical biochemistry and genetics but they have not been subjected to comparative genomic analysis. The availability of genome sequences from hundreds of diverse bacteria, and from Arabidopsis thaliana, enabled such an analysis using the SEED database and its tools. This study reports the results of the analysis and integrates them with new and existing experimental data.

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.

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. 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. 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.

Summary: Folate (vitamin B9) deficiency causes several health problems globally. However, folate biofortification of major staple crops is one alternative that can be used to improve vitamin intakes in populations at risk. We increased the folate levels in common bean by engineering the pteridine branch required for their biosynthesis. GTP cyclohydrolase I from Arabidopsis (AtGchI) was stably introduced into three common bean Pinto cultivars by particle bombardment. Seed‐specific overexpression of AtGCHI caused significant increases of up to 150‐fold in biosynthetic pteridines in the transformed lines. The pteridine boost enhanced folate levels in raw desiccated seeds by up to threefold (325 μg in a 100 g portion), which would represent 81% of the adult recommended daily allowance. Unexpectedly, the engineering also triggered a general increase in PABA levels, the other folate precursor. This was not observed in previous engineering studies and was probably caused by a feedforward mechanism that remains to be elucidated. Results from this work also show that common bean grains accumulate considerable amounts of oxidized pteridines that might represent products of folate degradation in desiccating seeds. Our study uncovers a probable different regulation of folate homoeostasis in these legume grains than that observed in other engineering works. Legumes are good sources of folates, and this work shows that they can be engineered to accumulate even greater amounts of folate that, when consumed, can improve folate status. Biofortification of common bean with folates and other micronutrients represents a promising strategy to improve the nutritional status of populations around the world.

Lauraceous acetogenins are fatty acid derivatives with an odd-carbon aliphatic chain found in avocado (Persea americana Mill.). These compounds display a wide range of bioactivities that makes them candidates for use as antimicrobial and proapoptotic agents in the food industry and against cancer cells respectively. Existing knowledge about its metabolism in planta is scarce. This work quantifies eight different acetogenins accumulated in fruit tissues (peel, seed, and pulp) from 22 avocado cultivars to sample the existing variation using a targeted metabolomics approach. Multivariate analyses uncovered correlations among acetogenins present in fruit tissues and their chemical backbone that allowed a proposal for classifying them in three families (Avocatins, Pahuatins and Persenins). The seed acetogenin profile differed from that of the pulp and peel, which while different in concentration (peel accumulated low acetogenin amounts), had the same profile. Acetogenins from samples of known origin were also separated by variety using descriptive Linear Discriminant Analysis (LDA), and a chemotaxonomic model was generated via predictive LDA and was tested on samples from unknown origin. This work effectively sampled acetogenin contents and profile variability in seed (1.09–8.33 mg per g FW), peel (0.22–12.5 mg per g FW), and pulp (0.49–9.58 mg per g FW) from avocado fruit, as well as provides a putative classification to seven avocado cultivars. Results from this work show that the eight acetogenins followed are produced in all 22 avocado cultivars, which points to conserved metabolism among avocado plants.