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SUMMARY Zea mays(maize) makes phytoalexins such as sesquiterpenoid zealexins, to combat invading pathogens. Zealexins are produced from farnesyl diphosphate in microgram per gram fresh weight quantities. As farnesyl diphosphate is also a precursor for many compounds essential for plant growth, the question arises as to howZ. maysproduces high levels of zealexins without negatively affecting vital plant systems. To examine if specific pools of farnesyl diphosphate are made for zealexin synthesis we made CRISPR/Cas9 knockouts of each of the three farnesyl diphosphate synthases (FPS) inZ. maysand examined the resultant impacts on different farnesyl diphosphate‐derived metabolites. We found that FPS3 (GRMZM2G098569) produced most of the farnesyl diphosphate for zealexins, while FPS1 (GRMZM2G168681) made most of the farnesyl diphosphate for the vital respiratory co‐factor ubiquinone. Indeed,fps1mutants had strong developmental phenotypes such as reduced stature and development of chlorosis. The replication and evolution of thefpsgene family inZ. maysenabled it to produce dedicated FPSs for developmentally related ubiquinone production (FPS1) or defense‐related zealexin production (FPS3). This partitioning of farnesyl diphosphate production between growth and defense could contribute to the ability ofZ. maysto produce high levels of phytoalexins without negatively impacting its growth. 

Abstract Prenylated quinones are membrane-associated metabolites that serve as vital electron carriers for respiration and photosynthesis. The UbiE (EC 2.1.1.201)/MenG (EC 2.1.1.163) C-methyltransferases catalyze pivotal ring methylations in the biosynthetic pathways of many of these quinones. In a puzzling evolutionary pattern, prokaryotic and eukaryotic UbiE/MenG homologs segregate into 2 clades. Clade 1 members occur universally in prokaryotes and eukaryotes, excluding cyanobacteria, and include mitochondrial COQ5 enzymes required for ubiquinone biosynthesis; Clade 2 members are specific to cyanobacteria and plastids. Functional complementation of an Escherichia coli ubiE/menG mutant indicated that Clade 1 members display activity with both demethylbenzoquinols and demethylnaphthoquinols, independently of the quinone profile of their original taxa, while Clade 2 members have evolved strict substrate specificity for demethylnaphthoquinols. Expression of the gene-encoding bifunctional Arabidopsis (Arabidopsis thaliana) COQ5 in the cyanobacterium Synechocystis or its retargeting to Arabidopsis plastids resulted in synthesis of a methylated variant of plastoquinone-9 that does not occur in nature. Accumulation of methylplastoquinone-9 was acutely cytotoxic, leading to the emergence of suppressor mutations in Synechocystis and seedling lethality in Arabidopsis. These data demonstrate that in cyanobacteria and plastids, co-occurrence of phylloquinone and plastoquinone-9 has driven the evolution of monofunctional demethylnaphthoquinol methyltransferases and explains why plants cannot capture the intrinsic bifunctionality of UbiE/MenG to simultaneously synthesize their respiratory and photosynthetic quinones. 

Growing vegetables in controlled environments (CEs), such as hydroponics, aquaponics, and vertical structures, is a rapidly expanding industry in Florida and the United States, especially in nearby urban areas. Although lettuce ( Lactuca sativa ) is still mostly produced in fields, growing in CEs proximal to urban areas has become increasingly popular because it may facilitate reduced transportation time and associated postharvest degradation. Lettuce is among the top-most consumed vegetables in the United States and could provide some of the nutrition missing in the US diet. This research was planned to understand the levels of some vitamins that are key for human health, including vitamin E (tocopherols), vitamin K 1 (phylloquinone), and vitamin C (ascorbic acid), in lettuce grown in greenhouse hydroponics. Lettuce germplasm was grown using the hydroponic nutrient film technique system in three greenhouse experiments: at the beginning, middle, and end of the Florida, USA, growing season (from Aug 2020 to Mar 2021). Genetic variation for these vitamins were found among the germplasm tested in the four morphological types of lettuce, romaine, Boston, Latin, and leaf. In addition, a sugar analysis was conducted in this germplasm, of which fructose was the most abundant sugar. A significant genotype × environment (G × E) interaction was observed, indicating that the levels of these compounds, especially vitamins, was environment dependent. However, the presence of certain non-crossover G × E interactions indicates that selecting lettuce in a representative environment could result in new cultivars with higher vitamin content. This research marks the initial steps to improve lettuce for these vitamins, which can contribute to better health of US consumers, not for the highest amount of these compounds in lettuce but for the offset due to its high consumption. 

Coenzyme Q (CoQ) is an essential component of the mitochondrial electron transport chain and an important antioxidant present in all cellular membranes. CoQ deficiencies are frequent in aging and in age-related diseases, and current treatments are limited to CoQ supplementation. Strategies that rely on CoQ supplementation suffer from poor uptake and trafficking of this very hydrophobic molecule. In a previous study, the dietary flavonol kaempferol was reported to serve as a CoQ ring precursor and to increase the CoQ content in kidney cells, but neither the part of the molecule entering CoQ biosynthesis nor the mechanism were described. In this study, kaempferol labeled specifically in the B-ring was isolated from Arabidopsis plants. Kidney cells treated with this compound incorporated the B-ring of kaempferol into newly synthesized CoQ, suggesting that the B-ring is metabolized via a mechanism described in plant cells. Kaempferol is a natural flavonoid present in fruits and vegetables and possesses antioxidant, anticancer, and anti-inflammatory therapeutic properties. A better understanding of the role of kaempferol as a CoQ ring precursor makes this bioactive compound a potential candidate for the design of interventions aiming to increase endogenous CoQ biosynthesis and may improve CoQ deficient phenotypes in aging and disease. 

Plants have evolved the ability to derive the benzenoid moiety of the respiratory cofactor and antioxidant, ubiquinone (coenzyme Q), either from the β-oxidative metabolism of p-coumarate or from the peroxidative cleavage of kaempferol. Here, isotopic feeding assays, gene co-expression analysis and reverse genetics identified Arabidopsis 4-COUMARATE-COA LIGASE 8 (4-CL8; At5g38120) as a contributor to the β-oxidation of p-coumarate for ubiquinone biosynthesis. The enzyme is part of the same clade (V) of acyl-activating enzymes than At4g19010, a p-coumarate CoA ligase known to play a central role in the conversion of p-coumarate into 4-hydroxybenzoate. A 4-cl8 T-DNA knockout displayed a 20% decrease in ubiquinone content compared with wild-type plants, while 4-CL8 overexpression boosted ubiquinone content up to 150% of the control level. Similarly, the isotopic enrichment of ubiquinone's ring was decreased by 28% in the 4-cl8 knockout as compared with wild-type controls when Phe-[Ring-13C6] was fed to the plants. This metabolic blockage could be bypassed via the exogenous supply of 4-hydroxybenzoate, the product of p-coumarate β-oxidation. Arabidopsis 4-CL8 displays a canonical peroxisomal targeting sequence type 1, and confocal microscopy experiments using fused fluorescent reporters demonstrated that this enzyme is imported into peroxisomes. Time course feeding assays using Phe-[Ring-13C6] in a series of Arabidopsis single and double knockouts blocked in the β-oxidative metabolism of p-coumarate (4-cl8; at4g19010; at4g19010 × 4-cl8), flavonol biosynthesis (flavanone-3-hydroxylase), or both (at4g19010 × flavanone-3-hydroxylase) indicated that continuous high light treatments (500 µE m−2 s−1; 24 h) markedly stimulated the de novo biosynthesis of ubiquinone independently of kaempferol catabolism.