EFFECTS OF CALCIUM NITRATE STRESS ON ASCORBATE-GLUTATHIONE CYCLE METABOLISM IN LEAVES OF HYDROPONICALLY-GROWN GRAFTED EGGPLANT SEEDLINGS

 以日本引进的设施专用耐盐茄(Solanum melongena)品种‘Torvum Vigor’为砧木, 栽培茄(S. torvum)品种‘苏崎茄’为接穗, 用营养液栽培, 对80 mmol&;#8226;L^–1 Ca(NO₃)₂胁迫下茄子嫁接苗和自根苗叶片抗坏血酸-谷胱甘肽循环系统中抗氧化酶活性和抗氧化物及H₂O₂含量进行比较。结果表明, Ca(NO₃)₂胁迫下茄子幼苗叶片H₂O₂含量有所增加, 但嫁接苗叶片H₂O₂含量显著低于自根苗。Ca(NO₃)₂胁迫下嫁接苗叶片抗氧化酶(APX、DHAR和GR)活性、AsA和GSH再生率、氧化还原力(AsA/DHA值和GSH/GSSG值)均显著高于自根苗。综上所述, Ca(NO₃)₂胁迫下嫁接苗保持良好的AsA-GSH循环效率, 清除H₂O₂效率较高, 细胞受氧化损伤程度较轻, 表现出较强的耐盐性。     

The effects of grafting on glycolysis and the tricarboxylic acid cycle in Ca(NO<Subscript>3</Subscript>)<Subscript>2</Subscript>-stressed cucumber seedlings with pumpkin as rootstock

In this research, we investigated the effects of grafting on intermediate metabolites and key enzymes of glycolysis and the tricarboxylic acid (TCA) cycle in self-grafted and salt-tolerant pumpkin rootstock-grafted cucumber seedlings supplied with nutrient solution and subjected to 80 mM Ca(NO₃)₂ stress for 6 days. Ca(NO₃)₂ stress induced accumulation of 3-phosphoglycerate (3-PGA) and phosphoenolpyruvate (PEP) in the leaves of self-grafted cucumber seedlings and enhanced the activities of phosphoenolpyruvate carboxylase (PEPC) and enolase (ENO). Succinic acid and malic acid contents and isocitrate dehydrogenase, succinate dehydrogenase (SDH), and malate dehydrogenase (MDH) activities in self-grafted seedlings were significantly decreased by Ca(NO₃)₂ stress. In addition, activities of PEPC, ENO, SDH, and MDH and contents of glycolysis intermediate metabolites (citric, succinic, and malic acids) were significantly higher in leaves of rootstock-grafted seedlings compared with those in self-grafted seedlings under saline conditions. Furthermore, leaf adenosine triphosphate (ATP) content of rootstock-grafted seedlings was relatively higher than that in self-grafted plants under salt stress, with an opposite effect observed on adenosine diphosphate content. These results indicate that rootstock grafting alleviates Ca(NO₃)₂ stress-induced inhibition of the glycolytic pathway and the TCA cycle in cucumber seedling leaves, which may aid the respiratory metabolism of cucumber seedlings and help maintain a high ATP synthesis level, thereby increasing the biomass of cucumber seedlings and enhancing their salt tolerance.     

Effects of Ca(NO<Subscript>3</Subscript>)<Subscript>2</Subscript> stress on oxidative damage,antioxidant enzymes activities and polyamine contents in roots of grafted and non-grafted tomato plants

The effects of Ca(NO₃)₂ stress on biomass production, oxidative damage, antioxidant enzymes activities and polyamine contents in roots of grafted and non-grafted tomato plants were investigated. Results showed that when exposed to 80 mM Ca(NO₃)₂ stress, the biomass production reduction in non-grafted plants was more significant than that of grafted plants. Under Ca(NO₃)₂ stress, superoxide anion radical (O₂•⁻) producing rate, hydrogen peroxide (H₂O₂) and malondialdehyde (MDA) contents of non-grafted plants roots were significantly higher than those of grafted plants, however, nitrate (NO₃ ⁻), ammonium (NH₄ ⁺) and proline contents, superoxide dismutase (SOD, EC1.15.1.1), peroxidase (POD, EC1.11.1.7), catalase (CAT, EC1.11.1.6) and arginine decarboxylase (ADC, EC 4.1.1.19) activities of grafted plants roots were significantly higher than those of non-grafted plants. Regardless of stress, free, conjugated and bound polyamine contents in roots of grafted plants were significantly higher than those of non-grafted plants. The possible roles of antioxidant enzymes, prolines and polyamines in adaptive mechanism of tomato roots to Ca(NO₃)₂ stress were discussed. Gu-Wen Zhang and Zheng-Lu Liu contributed equally to this work.     

A meta‐analysis of fertilizer‐induced soil NO and combined NO+N2O emissions

Soils are among the important sources of atmospheric nitric oxide (NO) and nitrous oxide (N₂O), acting as a critical role in atmospheric chemistry. Updated data derived from 114 peer‐reviewed publications with 520 field measurements were synthesized using meta‐analysis procedure to examine the N fertilizer‐induced soil NO and the combined NO+N₂O emissions across global soils. Besides factors identified in earlier reviews, additional factors responsible for NO fluxes were fertilizer type, soil C/N ratio, crop residue incorporation, tillage, atmospheric carbon dioxide concentration, drought and biomass burning. When averaged across all measurements, soil NO‐N fluxes were estimated to be 4.06 kg ha^?1 yr^?1, with the greatest (9.75 kg ha^?1 yr^?1) in vegetable croplands and the lowest (0.11 kg ha^?1 yr^?1) in rice paddies. Soil NO emissions were more enhanced by synthetic N fertilizer (+38%), relative to organic (+20%) or mixed N (+18%) sources. Compared with synthetic N fertilizer alone, synthetic N fertilizer combined with nitrification inhibitors substantially reduced soil NO emissions by 81%. The global mean direct emission factors of N fertilizer for NO (EF_NO) and combined NO+N₂O (EF_c) were estimated to be 1.16% and 2.58%, with 95% confidence intervals of 0.71–1.61% and 1.81–3.35%, respectively. Forests had the greatest EF_NO (2.39%). Within the croplands, the EF_NO (1.71%) and EF_c (4.13%) were the greatest in vegetable cropping fields. Among different chemical N fertilizer varieties, ammonium nitrate had the greatest EF_NO (2.93%) and EF_c (5.97%). Some options such as organic instead of synthetic N fertilizer, decreasing N fertilizer input rate, nitrification inhibitor and low irrigation frequency could be adopted to mitigate soil NO emissions. More field measurements over multiyears are highly needed to minimize the estimate uncertainties and mitigate soil NO emissions, particularly in forests and vegetable croplands.     

Proteome Analysis of Roots in Cucumber Seedlings Under Iso-Osmotic NaCl and Ca(NO3)2 Stresses

The growth and productivity of cucumber are severely affected by salinity. To understand the complex salt response mechanism, the physiological and biochemical responses of cucumber seedlings to iso-osmotic NaCl and Ca(NO₃)₂ stresses were investigated. In this study, the biomass was significantly decreased under iso-osmotic NaCl and Ca(NO₃)₂ stresses, and the inhibitory effect of Ca(NO₃)₂ stress was less than that of NaCl stress. The soluble protein contents were increased under Ca(NO₃)₂ stress, whereas they were decreased after 6 days of NaCl stress. A sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis revealed that there were 14 differentially expressed protein bands in roots under iso-osmotic NaCl and Ca(NO₃)₂ stresses at 0, 3, 6, and 9 days, and seven protein bands were little expressed under NaCl stress at 6 and 9 days. Based on these results, 2-D gel electrophoresis was used to separate cucumber root proteins in response to iso-osmotic NaCl and Ca(NO₃)₂ stresses at 3 days. A total of 43 protein spots changed under salt stress. Of these proteins, 33 were successfully identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and liquid chromatography electro-spray ionization tandem mass spectrometry (LC-ESI-MS/MS) and categorized into classes, including those corresponding to antioxidants and defense-related proteins and energy and metabolism. The functions of the significantly differentially expressed root proteins were analyzed, which may facilitate a better understanding of different salt response mechanisms, and we suggest that cucumber seedlings showed a more powerful ability to resist Ca(NO₃)₂ stress.

     

Mechanism of dihydrofolate reductase downregulation in melanoma by 3‐O‐(3,4,5‐trimethoxybenzoyl)‐(−)‐epicatechin

In our search to improve the stability and cellular absorption of tea polyphenols, we synthesized 3‐O‐(3,4,5‐trimethoxybenzoyl)‐(?)‐epicatechin (TMECG), which showed high antiproliferative activity against melanoma. TMECG downregulates dihydrofolate reductase (DHFR) expression in melanoma cells and we detail the sequential mechanisms that result from this even. TMECG is specifically activated in melanoma cells to form a stable quinone methide (TMECG‐QM). TMECG‐QM has a dual action on these cells. First, it acts as a potent antifolate compound, disrupting folate metabolism and increasing intracellular oxidized folate coenzymes, such as dihydrofolate, which is a non‐competitive inhibitor of dihydropterine reductase, an enzyme essential for tetrahydrobiopterin (H₄B) recycling. Such inhibition results in H₄B deficiency, endothelial nitric oxide synthase (eNOS) uncoupling and superoxide production. Second, TMECG‐QM acts as an efficient superoxide scavenger and promotes intra‐cellular H₂O₂ accumulation. Here, we present evidence that TMECG markedly reduces melanoma H₄B and NO bioavailability and that TMECG action is abolished by the eNOS inhibitor N_ω‐nitro‐L ‐arginine methyl ester or the H₂O₂ scavenger catalase, which strongly suggests H₂O₂‐dependent DHFR downregulation. In addition, the data presented here indicate that the simultaneous targeting of important pathways for melanoma survival, such as the folate cycle, H₄B recycling, and the eNOS reaction, could represent an attractive strategy for fighting this malignant skin pathology. J. Cell. Biochem. 110: 1399–1409, 2010. © 2010 Wiley‐Liss, Inc.     

镉胁迫下紫花苜蓿幼苗内源一氧化氮和活性氧的生成

以"甘农三号"紫花苜蓿幼苗为材料,在水培条件下,研究了不同浓度镉(Cd)胁迫下紫花苜蓿根、茎和叶内源一氧化氮(NO)和活性氧(ROS)的生成机制以及根系活力的变化.结果表明:在0~2.0 mmol·L-1范围内,随着Cd浓度的增加,幼苗内NO含量呈现先升高后降低的趋势,最后可维持在略高或持平于对照的水平.幼苗内一氧化氮合成酶(NOS)活性、硝酸还原酶(NR)活性、亚硝酸根离子(NO2-)含量和类胡萝卜素(Car)含量的变化与NO含量变化规律相似却又不全相同.NOS和NR是影响幼苗茎中NO含量的主要因素,NOS、NO2-和NR则是影响叶中NO含量的主要因素,而根中NO含量主要与NOS活性和NO2-含量有较大相关性.随着Cd浓度的增加,幼苗内过氧化氢(H2 O2)含量、丙二醛(MDA)含量、超氧阴离子(O-2·)含量和相对电导率(REC)呈现显著升高趋势,说明高浓度的Cd处理会使ROS大量积累,细胞膜遭破坏,细胞质外流,进而引发膜脂过氧化.随着Cd浓度的增加,紫花苜蓿根系活力的变化为先升高后降低,指示了低浓度Cd处理会促进植物代谢,增强其生命力;而高浓度Cd会致使植株代谢受抑制,细胞受损害.NO和ROS的相关性不大,说明二者虽同为自由基,但它们产生和变化方式大有差别.     

Nitric oxide and hydrogen peroxide are important signals mediating the allelopathic response of Arabidopsis to p‐hydroxybenzoic acid

Both nitric oxide (NO) and hydrogen peroxide (H₂O₂) are important signals that mediate plant response to environmental stimulation. Their role in plants' allelopathic interactions has also been reported, but the underlying mechanism remains little understood. p‐Hydroxybenzoic acid (pHBA) has been proposed to be an allelopathic chemical. Here, we found that pHBA at 0.4 mM efficiently suppressed Arabidopsis growth. Meanwhile, pHBA rapidly induced the accumulation of NO and H₂O₂, where such effect could be reversed by NO or H₂O₂ metabolism inhibitors or scavengers. Also, pHBA‐induced NO and H₂O₂ could be compromised in NO synthesis mutants noa1, nia1 and nia2, or H₂O₂ metabolism mutant rbohD/F, but suppressing NO accumulation with a NO synthesis inhibitor or using NO synthesis‐related mutants did not reduce pHBA‐induced H₂O₂ accumulation. Furthermore, we found that the effect of pHBA on allelopathic inhibition of growth was aggravated in NO/H₂O₂ metabolism‐related mutants or reducing NO/H₂O₂ by different inhibitors, whereas the addition of an NO/H₂O₂ donor could partly relieve the inhibitory effect of pHBA on the growth of wild type. However, adding only an NO donor, but not low concentration of H₂O₂ as the donor, could relieve the inhibitory effect of pHBA on root growth in NO metabolism mutants. On the basis of these results, we propose that both NO and H₂O₂ are important signals that mediate Arabidopsis response to the allelopathic chemical pHBA, where during this process H₂O₂ may work upstream of the NO signal.     

WD40‐REPEAT 5a functions in drought stress tolerance by regulating nitric oxide accumulation in Arabidopsis

Nitric oxide (NO) generation by NO synthase (NOS) in guard cells plays a vital role in stomatal closure for adaptive plant response to drought stress. However, the mechanism underlying the regulation of NOS activity in plants is unclear. Here, by screening yeast deletion mutants with decreased NO accumulation and NOS‐like activity when subjected to H₂O₂ stress, we identified TUP1 as a novel regulator of NOS‐like activity in yeast. Arabidopsis WD40‐REPEAT 5a (WDR5a), a homolog of yeast TUP1, complemented H₂O₂‐induced NO accumulation of a yeast mutant Δtup1, suggesting the conserved role of WDR5a in regulating NO accumulation and NOS‐like activity. This note was further confirmed by using an Arabidopsis RNAi line wdr5a‐1 and two T‐DNA insertion mutants of WDR5a with reduced WDR5a expression, in which both H₂O₂‐induced NO accumulation and stomatal closure were repressed. This was because H₂O₂‐induced NOS‐like activity was inhibited in the mutants compared with that of the wild type. Furthermore, these wdr5a mutants were more sensitive to drought stress as they had reduced stomatal closure and decreased expression of drought‐related genes. Together, our results revealed that WDR5a functions as a novel factor to modulate NOS‐like activity for changes of NO accumulation and stomatal closure in drought stress tolerance.     

Proteomics reveals the overlapping roles of hydrogen peroxide and nitric oxide in the acclimation of citrus plants to salinity

Hydrogen peroxide (H₂O₂) and nitric oxide (˙NO) are key reactive species in signal transduction pathways leading to activation of plant defense against biotic or abiotic stress. Here, we investigated the effect of pre‐treating citrus plants (Citrus aurantium L.) with either of these two molecules on plant acclimation to salinity and show that both pre‐treatments strongly reduced the detrimental phenotypical and physiological effects accompanying this stress. A proteomic analysis disclosed 85 leaf proteins that underwent significant quantitative variations in plants directly exposed to salt stress. A large part of these changes was not observed with salt‐stressed plants pre‐treated with either H₂O₂ or sodium nitroprusside (SNP; a ˙NO‐releasing chemical). We also identified several proteins undergoing changes either in their oxidation (carbonylation; 40 proteins) and/or S‐nitrosylation (49 proteins) status in response to salinity stress. Both H₂O₂ and SNP pre‐treatments before salinity stress alleviated salinity‐induced protein carbonylation and shifted the accumulation levels of leaf S‐nitrosylated proteins to those of unstressed control plants. Altogether, the results indicate an overlap between H₂O₂‐ and ˙NO‐signaling pathways in acclimation to salinity and suggest that the oxidation and S‐nitrosylation patterns of leaf proteins are specific molecular signatures of citrus plant vigour under stressful conditions.     

Hydrogen peroxide is involved in nitric oxide‐induced cell death in maize leaves

Recent evidence indicates that nitric oxide (NO) plays an important role in plant hypersensitive cell death. Here, we report that NO treatment led to rapid cell death and induced hydrogen peroxide (H₂O₂) accumulation in maize leaves. We also show that NO induced the expression of Zmrboh genes. Pharmacological study suggests that NO‐induced cell death is in part mediated via H₂O₂. In addition, semi‐quantitative RT‐PCR revealed that NO induced expression of the systemic acquired resistance (SAR) genes, ZmPR1 and ZmPR5.     

Cross-talk between salicylic acid and NaCl-generated reactive oxygen species and nitric oxide in tomato during acclimation to high salinity

Hydrogen peroxide (H₂O₂) and nitric oxide (NO) generated by salicylic acid (SA) are considered to be functional links of cross‐tolerance to various stressors. SA‐stimulated pre‐adaptation state was beneficial in the acclimation to subsequent salt stress in tomato (Solanum lycopersicum cv. Rio Fuego). At the whole‐plant level, SA‐induced massive H₂O₂ accumulation only at high concentrations (10^?3–10^?2M), which later caused the death of plants. The excess accumulation of H₂O₂ as compared with plants exposed to 100 mM NaCl was not associated with salt stress response after SA pre‐treatments. In the root tips, 10^?3–10^?2M SA triggered the production of reactive oxygen species (ROS) and NO with a concomitant decline in the cell viability. Sublethal concentrations of SA, however, decreased the effect of salt stress on ROS and NO production in the root apex. The attenuation of oxidative stress because of high salinity occurred not only in pre‐adapted plants but also at cell level. When protoplasts prepared from control leaves were exposed to SA in the presence of 100 mM NaCl, the production of NO and ROS was much lower and the viability of the cells was higher than in salt‐treated samples. This suggests that, the cross‐talk of signalling pathways induced by SA and high salinity may occur at the level of ROS and NO production. Abscisic acid (ABA), polyamines and 1‐aminocyclopropane‐1‐carboxylic acid, the compounds accumulating in pre‐treated plants, enhanced the diphenylene iodonium‐sensitive ROS and NO levels, but, in contrast to others, ABA and putrescine preserved the viability of protoplasts.     

Grape seed extract enhances eNOS expression and NO production through regulating calcium‐mediated AKT phosphorylation in H2O2‐treated endothelium

GSE (grape seed extract) has been shown to exhibit protective effects against cardiovascular events and atherosclerosis, although the underlying molecular mechanisms of action are unknown. Herein, we assessed the ability of GSE to enhance eNOS (endothelial nitric oxide synthase) expression and NO (nitric oxide) production in H₂O₂ (hydrogen peroxide)‐treated HUVECs (human umbilical vein endothelial cells). GSE enhanced eNOS expression and NO release in H₂O₂‐treated cells in a dose‐dependent manner. GSE inhibited intracellular ROS (reactive oxygen species) and reduced intracellular calcium in a dose‐dependent manner in H₂O₂‐treated cells, as shown by confocal microscopy. ROS was inhibited in cells pretreated with 5.0 μM GSE, 2.0 μM TG (thapsigargin) and 20.0 μM 2‐APB (2‐aminoethoxydiphenyl borate) instead of 0.25 μM extracellular calcium. In addition, GSE enhanced eNOS expression and reduced ROS production via increasing p‐AKT (AKT phosphorylation) with high extracellular calcium (13 mM). In conclusion, GSE protected against endothelial injury by up‐regulation of eNOS and NO expression via inhibiting InsP₃Rs (inositol 1,4,5‐trisphosphate receptors)‐mediated intracellular excessive calcium release and by activating p‐AKT in endothelial cells.     

Effects of root and foliar applications of exogenous NO on alleviating cadmium toxicity in lettuce seedlings

The effects of sodium nitroprusside (SNP, a donor of NO) on cadmium (Cd) toxicity in lettuce seedlings were studied. SNP was added into hydroponic systems or sprayed directly on the leaves of plants grown with and without Cd. Excess supply of Cd (100 μM) caused growth inhibition, dramatically increased Cd accumulation in both leaves and roots, and inhibited the absorption of Ca, Mg, Fe and Cu. Excess Cd also decreased activities of superoxide dismutase peroxidase and catalase in leaves and roots, and increased the accumulation of superoxide anion (O ₂ ^·? ), hydrogen peroxide (H₂O₂) and malondialdehyde (MDA). Root or foliar applications of exogenous NO alleviated Cd-induced growth suppression, especially root application of 250 μM SNP and foliar addition of 500 μM SNP. Addition of SNP promoted the chlorophyll synthesis suggesting that the photosynthesis was up-regulated. Exogenous NO increased Cd-decreased activities of antioxidant enzymes and markedly diminished Cd-induced reactive oxygen species (ROS) and MDA accumulation. Moreover, the absorption of Ca, Mg, Fe and Cu was increased, indicating that exogenous NO stimulated H⁺-ATPase activity to promote sequestration or uptake of ions. In addition, exogenous NO also inhibited Cd transfer from roots to shoots, which may indicate that Cd retention in roots induced by NO plays a significant role in Cd tolerance in lettuce seedlings. These data suggest that under Cd stress, exogenous NO improves photosynthesis by increasing chlorophyll synthesis, protects lettuce seedlings against oxidative damage by scavenging ROS, helps to maintain the uptake of nutrient elements, and inhibits Cd transferred to shoots effectively.     

NO serves as a signaling intermediate downstream of H2O2 to modulate dynamic microtubule cytoskeleton during responses to VD-toxins in Arabidopsis

Although hydrogen peroxide (H₂O₂) and nitric oxide (NO) can act as an upstream signaling molecule to modulate the dynamic microtubule cytoskeleton during the defense responses to Verticillium dahliae (VD) toxins in Arabidopsis, it is not known the relationship between these two signaling molecules. Here, we show that VD-toxin-induced NO accumulation was dependent on prior H₂O₂ production, NO is downstream of H₂O₂ in the signaling process, and that H₂O₂ acted synergistically with NO to modulate the dynamic microtubule cytoskeleton responses to VD-toxins in Arabidopsis.     

Hydrogen sulphide may be a novel downstream signal molecule in nitric oxide‐induced heat tolerance of maize (Zea mays L.) seedlings

Nitric oxide (NO) is a second messenger with multifunction that is involved in plant growth, development and the acquisition of stress tolerance. In recent years, hydrogen sulphide (H₂S) has been found to have similar functions, but crosstalk between NO and H₂S in the acquisition of heat tolerance is not clear. In this study, pretreatment with the NO donor sodium nitroprusside (SNP) improved the survival percentage of maize seedlings and alleviated an increase in electrolyte leakage and a decrease in tissue vitality as well as accumulation of malondialdehyde, indicating that pretreatment with SNP improved the heat tolerance of maize seedlings. In addition, pretreatment with SNP enhanced the activity of L‐cystine desulfhydrase, which, in turn, induced accumulation of endogenous H₂S, while application of H₂S donors, NaHS and GYY4137, increased endogenous H₂S content, followed by mitigating increase in electrolyte leakage and enhanced survival percentage of seedlings under heat stress. Interestingly, SNP‐induced heat tolerance was enhanced by application of NaHS and GYY4137, but was eliminated by inhibitors of H₂S synthesis DL‐propargylglycine, aminooxyacetic acid, potassium pyruvate and hydroxylamine, and the H₂S scavenger hypotaurine. All of the above‐mentioned results suggest that SNP pretreatment could improve heat tolerance, and H₂S may be a downstream signal molecule in NO‐induced heat tolerance of maize seedlings.     

Glucose-6-phosphate dehydrogenase plays critical role in artemisinin production of <Emphasis Type="Italic">Artemisia annua</Emphasis> under salt stress

Artemisinin, a natural sesquiterpenoid isolated from Artemisia annua L., is regarded as the most efficient drug against malaria in the world. Artemsinin production in NaCl-treated A. annua seedlings and its relationships with the glucose-6-phosphate dehydrogenase (G6PDH) activity and generation of H₂O₂ and nitric oxide (NO) were investigated. Results revealed that artemisinin content in the seedlings was increased by 79.3 % over the control after 1-month treatment with 68 mM NaCl. The G6PDH activity was enhanced in the presence of NaCl together with stimulated generation of H₂O₂ and NO. Application of 1.0 mM glucosamine (GlcN), an inhibitor of G6PDH, blocked the increase of NADPH oxidase and nitrate reductase (NR) activities, as well as H₂O₂ and NO production in A. annua seedlings under the salt stress. The induced H₂O₂ was found to be involved in the upgrading gene expression of two key enzymes in the later stage of artemisinin biosynthetic pathway: amorphadiene synthase (ADS) and amorpha-4,11-diene monooxygenase (CYP71AV1). The released NO being attributed mainly to the increase of NR activity, negatively interacted with H₂O₂ production and enhanced gene expression of 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGR). Inhibition of NO generation partly blocked NaCl-induced artemisinin accumulation, and NO donor strongly rescued the decreased content of artemisinin caused by GlcN. These results suggest that G6PDH could play a critical role in NaCl-induced responses and artemisinin biosynthesis in A. annua.