Exogenous ATP triggers antioxidant defense system and alleviates Cd toxicity in maize seedlings.

The role of exogenous adenosine 5'-triphosphate (ATP) in the regulation of antioxidant response in plants under heavy metal stress is unclear. Here, we investigated the effects of exogenous ATP application on plant growth, antioxidant response, and Cd accumulation in maize seedlings. Treatment with 0.1 mM CdCl2 moderately reduced dry weight, decreased chlorophyll content, impaired photosynthesis, and increased lipid peroxidation in maize seedlings compared with controls. However, toxicity due to Cd was alleviated after 10-200 µM ATP treatment. Subsequently, the activity of Cd-regulated antioxidant enzymes, antioxidant metabolite accumulation, and total antioxidant capacity were drastically enhanced after 50 µM ATP treatment. Similar patterns were observed in the ADP-treated group but not in the AMP-treated group under Cd stress. However, the ATP-induced elevation in antioxidant defense ability was decreased by the inhibition of NADPH oxidase (NOX). ATP-induced elevation in NOX activity and H2O2 production was partly reversed by the inhibition of NOX in maize seedlings under Cd stress. Furthermore, ATP promoted Cd accumulation in the roots and shoots of maize seedlings. However, the ATP-induced increase in Cd accumulation was partly abolished by the inhibition of NOX. To our knowledge, this is the first report on the role and mechanism of exogenous ATP in regulating plant growth, antioxidant response, and heavy metal phytoextraction. The study provides a new method based on exogenous ATP for enhancing heavy metal tolerance in plants.

Curcumin treatment enhances bioactive metabolite accumulation and reduces enzymatic browning in soybean sprouts during storage.

Curcumin is a natural polyphenol that is widely used in food and medicine. Here, we investigated the effects of curcumin on the antioxidant accumulation and enzymatic browning of soybean sprouts after storage at 4 °C for 2 weeks. Curcumin drastically reduced the water loss, browning index, and peroxide accumulation, increased the activities of superoxide dismutase, catalase, and peroxidase, decreased the activities of phenylalanine ammonia-lyase and polyphenol oxidase, elevated the contents of ascorbic acid, reduced glutathione, nonprotein thiol, phenolics and isoflavones, and enhanced the total antioxidant capacity of soybean sprouts during storage. These curcumin-induced changes were partly but dramatically attenuated by inhibition of NADPH oxidase (NOX). Curcumin induced NOX activity and H2O2 burst in soybean sprouts during the first 24 h after treatment. The curcumin-induced antioxidants and -inhibited enzymatic browning are closely associated with NOX-dependent H2O2 signaling. The findings provide a new method for improving soybean sprout quality during storage.

Quantification and identification analysis of Ziziphus jujuba Mill. peel pigmentation at different developmental stages.

The fruit peel of a color mutant jujube cultivar, 'Sanbianhong' (SBF), was investigated using an ultra-high performance liquid chromatography quadrupole Orbitrap mass spectrometry (UHPLC-Q-Orbitrap MS) at five ripening stages (S1, Young fruit stage; S2, swelling stage; S3, white-mature stage; S4, pre-mature stage and S5, mature stage). Lutein, ß-carotene, chlorophyll a, chlorophyll b, and 13 anthocyanins were identified. Chlorophyll a and cyanidin 3-O-galactoside were considered key color metabolites in S1 with the content of 1.083 mg/g of fresh weight (FW) and 4.585 mg/g of FW, respectively. Delphinidin (0.488 mg/g FW) and cyanidin (6.259 mg/g FW) were identified as the key pigments in S3. Delphinidin 3-O-glucoside (0.256 mg/g FW) was identified as the key anthocyanin in maturity S5. Herein, the identification and quantitation of pigment-related metabolites of SBF were studied for the first time, and the results provide a theoretical basis for understanding the pigment changes of jujube fruit during ripening.

Metabolic and transcriptomic analyses reveal different metabolite biosynthesis profiles between leaf buds and mature leaves in Ziziphus jujuba mill.

Jujube leaf is well known for its high nutritional value and medicinal benefits. However, a thorough and dynamic assessment of the metabolites present in jujube leaves is lacking. Here, the primary and secondary metabolites from purple leaf buds and green mature leaves were investigated using ultra-high-performance liquid chromatography/tandem mass spectrometry. A total of 778 metabolites were characterized and more than 700 compounds were reported for the first time. Analysis of differentially accumulated metabolites showed that the flavonoids were the major differential metabolites and determined the leaf coloration. The transcriptome data indicated that 20 flavonoid structural genes and three main types of flavonoid regulatory genes were significantly differentially expressed. Moreover, light had a significant influence on flavonoid accumulation. These results improve our understanding of metabolite accumulation and the molecular mechanisms of flavonoid biosynthesis in jujube leaf.

A simple, rapid and efficient method for essential element supplementation based on seed germination.

Fertilizer application is typically used to increase the availability of essential elements. In this study, an improved method for essential element supplementation based on seed germination was established. Solutions of essential elements (Fe2+, Zn2+, Cu2+, Mn2+, SeO32+, or I- or their combination) were applied to germinating soybean seeds, and the contents of the essential elements in the soybean sprouts were analyzed by atomic absorption spectroscopy. Compared with the control (seeds treated with water), the contents of iron, zinc, copper, manganese, selenium, and iodine in soybean sprouts produced by germinating seeds treated with solutions containing 10 mM essential elements were approximately 10-2000 times higher. Moreover, treatment with essential element solution increased the total antioxidant capacity and content of total thiols in the soybean sprouts. This rapid and simple technique can be used to improve nutrition for humans and livestock in regions deficient in essential elements.

Inhibition of autophagy by hydroxychloroquine enhances antioxidant nutrients and delays postharvest fruit senescence of Ziziphus jujuba.

Autophagy, a mechanism of recycling intracellular constituents, favors plant growth, especially under nutrient starvation. However, autophagy's role in regulating postharvest fruit senescence is unclear. Here, effects of the autophagy inhibitor hydroxychloroquine (HCQ) and activator LiCl on postharvest jujube fruit senescence were investigated. HCQ significantly reduced weight loss and decay incidence, and enhanced firmness compared with those of the control. LiCl had the opposite effects. Protein oxidation and H2O2 increased significantly in LiCl-treated compared with HCQ-treated fruit. The contents of vitamin C, total thiol, and phenolics, and total antioxidant capacity and DPPH-radical scavenging capacity, followed the order: HCQ > control > LiCl. The HCQ-mediated reduction in fruit respiration was significantly enhanced by ATP and partly reversed by 2,4-dinitrophenol, a mitochondrial uncoupler. Thus, jujube fruit senescence may be regulated by autophagy and the antioxidant capacity. A mechanism of autophagy-mediated postharvest fruit senescence involving mitochondrial biogenesis and respiration was proposed.

Mutagen-induced phytotoxicity in maize seed germination is dependent on ROS scavenging capacity.

Ethidium bromide (EB) and acridine orange (AO) bind to nucleic acids and are thus considered as potential mutagens. In this study, the effects of EB and AO on the germination behaviours of white, yellow, red, and purple maize seeds were investigated. The results indicate that low concentrations of EB (50 µg mL-1) and AO (500 µg mL-1) promote germination, particularly for the white and yellow seeds. However, high concentrations of EB (0.5 mg mL-1) and AO (5 mg mL-1) significantly inhibit germination, with the level of inhibition decreasing in the following order: white > yellow > red > purple. In addition, EB and AO induce H2O2 production in a concentration-dependent manner. The effects of these mutagens on seed germination were partly reversed by dimethyl thiourea, a scavenger of reactive oxygen species (ROS), and diphenylene iodonium (DPI), an inhibitor of NADPH oxidase, while the effects were enhanced by treatment with H2O2 and 3-amino-1,2,4-triazole, a specific inhibitor of catalase. In addition, AO and EB profoundly increased NADPH oxidase activities in germinating seeds. The treatment of seeds with EB and AO did not affect the growth or drought tolerance of the resultant seedlings. The findings suggest that the mechanism of mutagen toxicity is related to the induction of ROS production.

Inhibition of NADPH oxidase increases defense enzyme activities and improves maize seed germination under Pb stress.

Contaminated soil accumulated high levels of Pb, which shows great risk to human health and crop growth. To alleviate Pb impaired seed germination and seedling growth, effects of three methods were compared. Here, effects of the heavy metal chelator EDTA, reactive oxygen species (ROS) scavenger (e.g. dimethylthiourea/DMTU, glutathione and melatonin), and specific inhibitors of NADPH oxidase or NOX (e.g. imidazole/IMZ and diphenylene iodonium/DPI), on maize seed germination and seedling growth were examined under Pb stress. IMZ and DPI increased seed germination by 1-2-fold under Pb stress, compared with less than 50% for ROS scavengers, while EDTA decreased germination. Pb-induced H2O2 accumulation was reduced more dramatically by IMZ than DMTU. Compared with DMTU, Pb-impaired SOD and CAT enzyme can be reversed more significantly by IMZ. Thus, inhibiting the NOX was more efficient than using ROS scavengers for improving seed germination under Pb stress. Compared with EDTA and ROS scavenger, IMZ and DPI treatment cannot protect seedling growth under Pb stress. In addition, IMZ is cheap and highly efficient, making it suitable for improving seed germination in Pb-polluted soil.

Omethoate treatment mitigates high salt stress inhibited maize seed germination.

Omethoate (OM) is a highly toxic organophophate insecticide, which is resistant to biodegradation in the environment and is widely used for pest control in agriculture. The effect of OM on maize seed germination was evaluated under salt stress. Salt (800mM) greatly reduced germination of maize seed and this could be reversed by OM. Additionally, H2O2 treatment further improved the effect of OM on seed germination. Higher H2O2 content was measured in OM treated seed compared to those with salt stress alone. Dimethylthiourea (DTMU), a specific scavenger of reactive oxygen species (ROS), inhibited the effect of OM on seed germination, as did IMZ (imidazole), an inhibitor of NADPH oxidase. Abscisic acid (ABA) inhibited the effect of OM on seed germination, whereas fluridone, a specific inhibitor of ABA biosynthesis, enhanced the effect of OM. Taken together, these findings suggest a role of ROS and ABA in the promotion of maize seed germination by OM under salt stress.

Can antioxidant's reactive oxygen species (ROS) scavenging capacity contribute to aged seed recovery? Contrasting effect of melatonin, ascorbate and glutathione on germination ability of aged maize seeds.

It is well known that antioxidants such as AA (reduced ascorbate), glutathione (GSH) (reduced glutathione) and melatonin can delay seed ageing. Can they recover aged seed? Artificial aged maize seeds were obtained and their reduced germination rate (GR) and high lipid peroxidation were recorded. Exogenous melatonin was applied on these aged seeds and enhanced GR was observed. However, treatment with other antioxidants such as AA, GSH or DMTU (dimethyl thiourea) did not significantly improve or even reduce the GR of aged seeds. In addition, melatonin improved germination ability of theses aged seeds can be significantly impaired by DDC (diethyldithiocarbamic acid, a specific inhibitor of superoxide dismutase or superoxide dismutase (SOD)) and ATZ (aminotriazol, a specific inhibitor of catalase or CAT). In a further study, we found that melatonin but not other antioxidants (AA, GSH and DMTU) significantly induced CAT and SOD activities of aged seeds after imbibition. Accordingly, melatonin significantly reduced lipid peroxidation in aged seeds than that of other antioxidants. Taken together, these data suggest that melatonin induced antioxidant enzyme but not its direct reactive oxygen species (ROS) scavenging capacity contributing to recovery of aged maize seeds.

Can heavy metal pollution defend seed germination against heat stress? Effect of heavy metals (Cu(2+), Cd(2+) and Hg(2+)) on maize seed germination under high temperature.

Heavy metal pollution, as well as greenhouse effect, has become a serious threat today. Both heavy metal and heat stresses can arrest seed germination. What response can be expected for seed germination under both stress conditions? Here, the effects of heavy metals (Cu(2+), Cd(2+) and Hg(2+)) on maize seed germination were investigated at 20 °C and 40 °C. Compared with 20 °C, heat stress induced thermodormancy. However, this thermodormancy could be significantly alleviated by the addition of a low concentration of heavy metals. Heavy metals, as well as heat stress induced H2O2 accumulation in germinating seeds. Interestingly, this low concentration of heavy metal that promoted seed germination could be partly blocked by DMTU (a specific ROS scavenger), irrespective of temperature. Accordingly, H2O2 addition reinforced this promoting effect on seed germination, which was induced by a low concentration of heavy metal. Furthermore, we found that the NADPH oxidase derived ROS was required for seed germination promoted by the heavy metals. Subsequently, treatment of seeds with fluridone (a specific inhibitor of ABA) or ABA significantly alleviated or aggravated thermodormancy, respectively. However, this alleviation or aggravation could be partly attenuated by a low concentration of heavy metals. In addition, germination that was inhibited by high concentrations of heavy metals was also partly reversed by fluridone. The obtained results support the idea that heavy metal-mediated ROS and hormone interaction can finally affect the thermodormancy release or not.

Down-regulation of free riboflavin content induces hydrogen peroxide and a pathogen defense in Arabidopsis.

Riboflavin mediates many bioprocesses associated with the generation of hydrogen peroxide (H2O2), a cellular signal that regulates defense responses in plants. Although plants can synthesize riboflavin, the levels vary widely in different organs and during different stages of development, indicating that changes in riboflavin levels may have physiological effects. Here, we show that changing riboflavin content affects H2O2 accumulation and a pathogen defense in Arabidopsis thaliana. Leaf content of free riboflavin was modulated by ectopic expression of the turtle gene encoding riboflavin-binding protein (RfBP). The RfBP-expressing Arabidopsis thaliana (REAT) plants produced the RfBP protein that possessed riboflavin-binding activity. Compared with the wild-type plant, several tested REAT lines had >70% less flavins of free form. This change accompanied an elevation in the level of H2O2 and an enhancement of plant resistance to a bacterial pathogen. All the observed REAT characters were eliminated due to RfBP silencing (RfBPi) under REAT background. When an H2O2 scavenger was applied, H2O2 level declined in all the plants, and REAT no longer exhibited the phenotype of resistance enhancement. However, treatment with an NADPH oxidase inhibitor diminished H2O2 content and pathogen defense in wild-type and RfBPi but not in REAT. Our results suggest that the intrinsic down-regulation of free flavins is responsible for NADPH oxidase-independent H2O2 accumulation and the pathogen defense.