Octadecanoid Pathway Research Articles

Molecular cloning and characterization of heat-responsive LcOPR1, a gene encoding oxophytodienoic acid reductase in lentil.

Improving crop plants using biotechnological implications is a promising and modern approach compared to traditional methods. High-temperature exposure to the reproductive stage induces flower abortion and declines grain filling performance, leading to smaller grain production and low yield in lentil and other legumes. Thus, cloning effective candidate genes and their implication in temperature stress tolerance in lentil (Lens culinaris Medik.) using biotechnological tools is highly demandable. The 12-oxophytodienoic acid reductases (OPRs) are flavin mononucleotide-dependent oxidoreductases with vital roles in plants. They are members of the old yellow enzyme (OYE) family. These enzymes are involved in the octadecanoid pathway, which contributes to jasmonic acid biosynthesis and is essential in plant stress responses. Lentil is one of the vital legume crops affected by the temperature fluctuations caused by global warming. Therefore, in this study, the LcOPR1 gene was successfully cloned and isolated from lentils using RT-PCR to evaluate its functional responses in lentil under heat stress. The bioinformatics analysis revealed that the full-length cDNA of LcOPR1 was 1303 bp, containing an 1134 bp open reading frames (ORFs), encoding 377 amino acids with a predicted molecular weight of 41.63 and a theoretical isoelectric point of 5.61. Bioinformatics analyses revealed that the deduced LcOPR1 possesses considerable homology with other plant 12-oxophytodienoic acid reductases (OPRs). Phylogenetic tree analysis showed that LcOPR1 has an evolutionary relationship with other OPRs in different plant species of subgroup I, containing enzymes that are not required for jasmonic acid biosynthesis. The expression analysis of LcOPR1 indicated that this gene is upregulated in response to the heat-stress condition and during recovery in lentil. This study finding might be helpful to plant breeders and biotechnologists in LcOPR1 engineering and/or plant breeding programs in revealing the biological functions of LcOPR1 in lentils and the possibility of enhancing heat stress tolerance by overexpressing LcOPR1 in lentil and other legume plants under high temperature.

Development of a Chiral Supercritical Fluid Chromatography-Tandem Mass Spectrometry and Reversed-Phase Liquid Chromatography-Tandem Mass Spectrometry Platform for the Quantitative Metabolic Profiling of Octadecanoid Oxylipins.

Octadecanoids are broadly defined as oxylipins (i.e., lipid mediators) derived from 18-carbon fatty acids. In contrast to the well-studied eicosanoids, there is a lack of analytical methods for octadecanoids, hampering further investigations in the field. We developed an integrated workflow combining chiral separation by supercritical fluid chromatography (SFC) and reversed-phase liquid chromatography (LC) coupled to tandem mass spectrometry detection for quantification of a broad panel of octadecanoids. The platform includes 70 custom-synthesized analytical and internal standards to extend the coverage of the octadecanoid synthetic pathways. A total of 103 octadecanoids could be separated by chiral SFC and complex enantioseparations could be performed in <13 min, while the achiral LC method separated 67 octadecanoids in 13.5 min. The LC method provided a robust complementary approach with greater sensitivity relative to the SFC method. Both methods were validated in solvent and surrogate matrix in terms of linearity, lower limits of quantification (LLOQ), recovery, accuracy, precision, and matrix effects. Instrumental linearity was good for both methods (R2 > 0.995) and LLOQ ranged from 0.03 to 6.00 ng/mL for SFC and 0.01 to 1.25 ng/mL for LC. The average accuracy in the solvent and surrogate matrix ranged from 89 to 109% in SFC and from 106 to 220% in LC, whereas coefficients of variation (CV) were <14% (at medium and high concentrations) and 26% (at low concentrations). Validation in the surrogate matrix showed negligible matrix effects (<16% for all analytes), and average recoveries ranged from 71 to 83%. The combined methods provide a platform to investigate the biological activity of octadecanoids and expand our understanding of these little-studied compounds.

Identification of the 12-oxophytoeienoic acid reductase (OPR) gene family in pepper (Capsicum annuum L.) and functional characterization of CaOPR6 in pepper fruit development and stress response.

The 12-oxophytoeienoic acid reductase (OPR) is a kind of enzyme in the octadecanoid biosynthesis pathway that determines the biosynthesis of jasmonic acid. Although the roles of OPRs have been extensively studied in several crop plants, little is known about the biological functions of OPR-encoding genes in Capsicum annuum plants. In this study, seven OPR family genes (CaOPR1-7) were identified from the C. annuum genome. The physical and chemical properties of CaOPR1-7 were further analyzed, including gene expression patterns, promoter elements, and chromosomal locations. The results showed that the seven CaOPR homologues could be divided into two subgroups, and CaOPR6 was highly similar to AtOPR3 in Arabidopsis. The expression of CaOPR6 was significantly induced by various stresses such as cold, salt, and pathogen infection, indicating that CaOPR6 plays important roles in response to abiotic and biotic stresses. Overall, these findings improve the understanding of the biological functions of CaOPR6 in the development of pepper fruit and stress response of pepper plants, and facilitate further studies on the molecular biology of OPR proteins in Solanaceae vegetables.

Central role of Methyl jasmonate in resistance of wheat against crown and root rot caused by Fusarium culmorum

The Fusarium crown and root rot pathogen Fusarium culmorum (Fc) is a hemibiotrophic pathogen, with a short biotrophic phase preceding necrotrophic life-style. In this study, for the first time, a comparative proteomic analysis was made in seedling roots of two wheat cultivars Falat (susceptible) and Sumai3 (resistant) subjected to Methyl jasmonate (MeJA) at 2 days post Fc-inoculation (2 dpi) to understand defense responses induced by treatment during the necrotrophic stage. Using a high-resolution 2-D gel, a total of 176 proteins were identified at 48 h after MeJA treatment. Among the 176 matched spots, the expression abundances (vol%) of 30 protein spots were changed more than 2-fold (p < 0.05), and thus were treated as differentially expressed proteins (DEPs). Most DEPs from both cultivars were assigned to the 4 categories: metabolic process, energy, defense, and cell structure. Along with the increased resistance induced by MeJA during the later stage of infection which leads to decrease in disease index of both cultivars particularly much greater in susceptible cv. Falat, we observed increased accumulation of isocitrate dehydrogenase, malate dehydrogenase, enolase, glyceraldehyde-3-phosphate dehydrogenase, lipoxygenase (LOX), ATP synthase, peroxidase and l-ascorbate peroxidase compared to untreated Fc-inoculated control in both cultivars. Moreover, the photosynthesis responsive DEPs like RuBisCO down-regulated, to adjust the metabolic status and reserve energy to confront the pathogen attack. Consequently, the soil application of MeJA could be one of a suitable and healthier solution in integrated management to decease the incidence of Fc-infection by triggering plant defense responses via octadecanoid pathway.

An evolutionarily ancient fatty acid desaturase is required for the synthesis of hexadecatrienoic acid, which is the main source of the bioactive jasmonate in Marchantia polymorpha.

Jasmonates are fatty acid-derived hormones that regulate multiple aspects of plant development, growth and stress responses. Bioactive jasmonates, defined as the ligands of the conserved COI1 receptor, differ between vascular plants and bryophytes (jasmonoyl-l-isoleucine (JA-Ile) and dinor-12-oxo-10,15(Z)-phytodienoic acid (dn-OPDA), respectively). The biosynthetic pathways of JA-Ile in the model vascular plant Arabidopsis thaliana have been elucidated. However, the details of dn-OPDA biosynthesis in bryophytes are still unclear. Here, we identify an orthologue of Arabidopsis fatty-acid-desaturase 5 (AtFAD5) in the model liverwort Marchantia polymorpha and show that FAD5 function is ancient and conserved between species separated by more than 450 million years (Myr) of independent evolution. Similar to AtFAD5, MpFAD5 is required for the synthesis of 7Z-hexadecenoic acid. Consequently, in Mpfad5 mutants, the hexadecanoid pathway is blocked, dn-OPDA concentrations are almost completely depleted and normal chloroplast development is impaired. Our results demonstrate that the main source of wounding-induced dn-OPDA in Marchantia is the hexadecanoid pathway and the contribution of the octadecanoid pathway (i.e. from OPDA) is minimal. Remarkably, despite extremely low concentrations of dn-OPDA, MpCOI1-mediated responses to wounding and insect feeding can still be activated in Mpfad5, suggesting that dn-OPDA may not be the only bioactive jasmonate and COI1 ligand in Marchantia.

The Algal Polysaccharide Ulvan Induces Resistance in Wheat Against Zymoseptoria tritici Without Major Alteration of Leaf Metabolome

This study aimed to examine the ability of ulvan, a water-soluble polysaccharide from the green seaweed Ulva fasciata, to provide protection and induce resistance in wheat against the hemibiotrophic fungus Zymoseptoria tritici. Matrix-assisted laser desorption/ionization-time-of-flight-mass spectrometry (MALDI-TOF-MS) analysis indicated that ulvan is mainly composed of unsaturated monosaccharides (rhamnose, rhamnose-3-sulfate, and xylose) and numerous uronic acid residues. In the greenhouse, foliar application of ulvan at 10 mg.ml–1 2 days before fungal inoculation reduced disease severity and pycnidium density by 45 and 50%, respectively. Ulvan did not exhibit any direct antifungal activity toward Z. tritici, neither in vitro nor in planta. However, ulvan treatment significantly reduced substomatal colonization and pycnidium formation within the mesophyll of treated leaves. Molecular assays revealed that ulvan spraying elicits, but does not prime, the expression of genes involved in several wheat defense pathways, including pathogenesis-related proteins (β-1,3-endoglucanase and chitinase), reactive oxygen species metabolism (oxalate oxidase), and the octadecanoid pathway (lipoxygenase and allene oxide synthase), while no upregulation was recorded for gene markers of the phenylpropanoid pathway (phenylalanine ammonia-lyase and chalcone synthase). Interestingly, the quantification of 83 metabolites from major chemical families using ultra-high-performance liquid chromatography-mass spectrometry (UHPLC-MS) in both non-infectious and infectious conditions showed no substantial changes in wheat metabolome upon ulvan treatment, suggesting a low metabolic cost associated with ulvan-induced resistance. Our findings provide evidence that ulvan confers protection and triggers defense mechanisms in wheat against Z. tritici without major modification of the plant physiology.

Avoidance of the Plant Hormone Cis-Jasmone by Aedes aegypti Depends On Mosquito Age in Both Plant and Human Odor Backgrounds

Adults of many mosquito species feed on plants to obtain metabolic energy and to enhance reproduction. Mosquitoes primarily rely on olfaction to locate plants and are known to respond to a range of plant volatiles. We studied the olfactory response of the yellow fever mosquito Aedes aegypti to methyl jasmonate (MeJA) and cis-jasmone (CiJA), volatile compounds originating from the octadecanoid signaling pathway that plays a key role in plant defense against herbivores. Specifically, we investigated how Ae. aegypti of different ages responded to elevated levels of CiJA in two attractive odor contexts, either derived from Lima bean plants or human skin. Aedes aegypti females landed significantly less often on a surface with CiJA and MeJA compared to the solvent control, CiJA exerting a stronger reduction in landing than MeJA. Odor context (plant or human) had no significant main effect on the olfactory responses of Ae. aegypti females to CiJA. Mosquito age significantly affected the olfactory response, older females (7–9 d) responding more strongly to elevated levels of CiJA than young females (1–3 d) in either odor context. Our results show that avoidance of CiJA by Ae. aegypti is independent of odor background, suggesting that jasmonates are inherently aversive cues to these mosquitoes. We propose that avoidance of plants with elevated levels of jasmonates is adaptive to mosquitoes to reduce the risk of encountering predators that is higher on these plants, i.e. by avoiding ‘enemy-dense-space’.

Verification of the versatility of the in vitro enzymatic reaction giving (+)-cis-12-Oxo-phytodienoic acid

Jasmonic acid (JA) is a plant hormone involved in the defense response against insects and fungi. JA is synthesized from α-linolenic acid (LA) by the octadecanoid pathway in plants. 12-oxo-Phytodienoic acid (OPDA) is one of the biosynthetic intermediates in this pathway. The reported stereo selective total synthesis of cis-(+)-OPDA is not very efficient due to the many steps involved in the reaction as well as the use of water sensitive reactions. Therefore, we developed an enzymatic method for the synthesis of OPDA using acetone powder of flax seed and allene oxide cyclase (PpAOC2) from Physcomitrella patens. From this method, natural cis-(+)-OPDA can be synthesized in the high yield of approximately 40%. In this study, we investigated the substrate specificity of the enzymatic synthesis of other OPDA analogs with successions to afford OPDA amino acid conjugates, dinor-OPDA (dn-OPDA), and OPDA monoglyceride, and it was suggested that the biosynthetic pathway of arabidopsides could occur via MGDG.

OPDAT1, a plastid envelope protein involved in 12-oxo-phytodienoic acid export for jasmonic acid biosynthesis in Populus.

Twelve-oxo-phytodienoic acid (OPDA), the cyclopentenone precursor of jasmonic acid (JA), is required for the wounding response of plants. OPDA is derived from plastid-localized α-linolenic acid (α-LeA; 18:3) via the octadecanoid pathway, and is further exported from plastids to the cytosol for JA biosynthesis. However, the mechanism of OPDA transport from plastids has yet to be elucidated. In the current study, a plastid inner envelope-localized protein, designated 12-oxo-Phtyodienoic Acid Transporter 1 (OPDAT1), was identified and shown to potentially be involved in OPDA export from plastids, in Populus trichocarpa. Torr. OPDAT1 is expressed predominantly in young leaves of P. trichocarpa. Functional expression of OPDAT1 in yeast cells revealed that OPDAT1 is involved in OPDA transport. Loss-of-function of OPDAT1 in poplar resulted in increased accumulation of OPDA in the extracted plastids and a reduction in JA concentration, whereas an OPDAT1-overexpressing line showed a reverse tendency in OPDA accumulation and JA biosynthesis. OPDAT1 transcripts were rapidly induced by mechanical wounding of leaves, and an opdat1 mutant transgenic plant displayed increased susceptibility to spider mite (Tetranychus urticae) infestation. Collectively, these data suggest that OPDAT1 is an inner envelope transporter for OPDA, and this has potential implications for JA biosynthesis in poplar under environmental stresses.

Fatty acid desaturases modulated octadecanoid pathway in Sesame

Omega 3 fatty acid desaturases are involved in the production of α-linolenic acid (LNA) an essential omega 3 fatty acid, which is present in only traces in sesame seeds. LNA is the precursor for jasmonic acid which is the end product of octadecanoid pathway. This study was undertaken to analyze the key components of octadecanoid pathway and its relationship with fatty acid content in sesame. Fatty acid desaturation and membrane fluidity are modulated differentially in various stresses. Sesame seedlings were subjected to transient stress to analyse the octadecanoid pathway and its impact on fatty acid desaturation. The mRNA levels of omega 3 desaturases and LNA content were higher in cold stressed sesame seedlings than heat, drought and salinity stresses. The LOX activity and MDA content were higher in heat stressed sesame seedlings. Jasmonic acid content was higher in salinity stressed seedlings while abscisic acid registered the highest in drought stressed seedlings. Chloroplast fatty acid desaturase genes expression was found to increase the LNA content in cold stressed seedlings. The level of membrane damage measured by lipid peroxidation in terms of LOX activity and MDA content were found to be minimal in cold stressed seedling. This suggests a possible role of LNA in membrane fluidity and cold acclimation in sesame. A synergistic role of JA and ABA is also suspected in abiotic stress tolerance in sesame.

A Plant Nutrient- and Microbial Protein-Based Resistance Inducer Elicits Wheat Cultivar-Dependent Resistance Against Zymoseptoria tritici.

The induction of plant defense mechanisms by resistance inducers is an attractive and innovative alternative to reduce the use of fungicides on wheat against Zymoseptoria tritici, the responsible agent of Septoria tritici blotch (STB). Under controlled conditions, we investigated the resistance induction in three wheat cultivars with different susceptible levels to STB as a response to a treatment with a sulfur, manganese sulfate, and protein-based resistance inducer (NECTAR Céréales). While no direct antigermination effect of the product was observed in planta, more than 50% reduction of both symptoms and sporulation were recorded on the three tested cultivars. However, an impact of the wheat genotype on resistance induction was highlighted, which affects host penetration, cell colonization, and the production of cell-wall degrading enzymes by the fungus. Moreover, in the most susceptible cultivar Alixan, the product upregulated POX2, PAL, PR1, and GLUC gene expression in both noninoculated and inoculated plants and CHIT2 in noninoculated plants only. In contrast, defense responses induced in Altigo, the most resistant cultivar, seem to be more specifically mediated by the phenylpropanoid pathway in noninoculated as well as inoculated plants, since PAL and CHS were most specifically upregulated in this cultivar. In Premio, the moderate resistant cultivar, NECTAR Céréales elicits mainly the octadecanoid pathway, via LOX and AOS induction in noninoculated plants. We concluded that this complex resistance-inducing product protects wheat against Z. tritici by stimulating the cultivar-dependent plant defense mechanisms.

The Octadecanoid Pathway, but Not COI1, Is Required for Nectar Secretion in Arabidopsis thaliana.

Over 75% of crop species produce nectar and are dependent on pollinators to achieve maximum seed set, yet little is known about the mechanisms regulating nectar secretion. The phytohormone jasmonic acid (JA) is recognized to be involved in several plant processes including development and defense. JA was also recently shown to positively influence nectar secretion in both floral and extrafloral nectaries. For example, endogenous JA levels peak in flowers just prior to nectar secretion, but the details of how JA regulates nectar secretion have yet to be elucidated. We have found that the octadecanoid pathway does indeed play a role in the production and regulation of floral nectar in Arabidopsis. Null alleles for several JA biosynthesis and response genes had significantly reduced amounts of nectar, as well as altered expression of genes known to be involved in nectar production. We additionally identified crosstalk between the JA and auxin response pathways in nectaries. For example, the nectar-less JA synthesis mutant aos-2 showed no auxin response in nectaries, but both nectar production and the auxin response were restored upon exogenous JA and auxin treatment. Conversely, coi1-1, a JA-Ile-insensitive receptor mutant, displayed no auxin response in nectaries under any circumstance, even in older flowers that produced nectar. Surprisingly, opr3-1, a mutant for 12-oxophytodienoate reductase 3 [an enzyme further down the JA biosynthetic pathway that reduces 12-oxo phytodienoic acid (OPDA)], produced no nectar in newly opened flowers, but did secrete nectar in older flowers. Furthermore, a similar phenotype was observed in coi1-1. Cumulatively, these observations strongly suggest an indispensable role for an octadecanoic acid- and auxin-dependent, but JA- and COI1-dispensible, pathway in regulating nectar production in Arabidopsis.

Effects of Methyl Jasmonate on the Composition of Volatile Compounds in Pyropia yezoensis

Volatile organic compounds in marine algae have been reported to comprise characteristic flavor of algae and play an important role in their growth, development and defensive response. Yet their biogeneration remain largely unknown. Here we studied the composition of volatile compouds in Pyropia yezoensis and their variations in response to methyl jasmonate (MeJA) and diethyldithiocarbamic acid (DIECA) treatment using gas chromatography-mass spectrometry (GC-MS). A total of 44 compounds belonging to the following chemical classes (n) were identified, including aldehydes (11), alcohols (8), acids and esters (6), alkanes (5), ketones (5), alkenes (3), and S- or N-containing miscellaneous compounds (6). External treatment with plant hormone MeJA increased the content of 1-dodecanol, 4-heptenal, and 2-propenoic acid-2-methyl dodecylester, but decreased the content of phytol, 3-heptadecene, 2-pentadecanone, and isophytol. When pretreated with DIECA, an inhibitor of the octadecanoid pathway leading to the biosynthesis of endogeneous jasmonates and some secondary metabolites, phytol and isophytol were increased, while 4-heptenal, 1-dodecanol, and 2-propenoic acid-2-methyl dodecylester were decreased, both of which were negatively correlated with their variations under MeJA treatment. Collectively, these results suggest that MeJA does affect the volatile composition of P. yezoensis, and the octadecanoid pathway together with endogenous jasmonate pathway may be involved in the biosynthesis of volatile compounds, thereby providing some preliminary envision on the composition and biogeneration of volatile compounds in P. yezoensis.

Cis-Jasmone treatments affect pests and beneficial insects of wheat (Triticum aestivum L.): the influence of doses and plant growth stages

cis-Jasmone (CJ) is a natural plant volatile derived from linolenic acid via the octadecanoid pathway, and acts as a semiochemical and plant defense elicitor. In the present study, the effects of three different CJ doses (25 g.ha−1; 50 g.ha−1; 100 g.ha−1), applied at two different wheat growth stages (flag leaf just visible and emergence of inflorescence completed), on wheat insect pests and beneficial insects were investigated using three different sampling methods (direct visual counting, sweep-netting, colored sticky traps) under semi-arid conditions during 2011–2013.Depending on the insect species, CJ dose-effects varied with study years, plant growth stages and/or sampling methods. Among the pest species, cereal aphids (apterous and alate) were the most prominently affected by CJ treatment, irrespective of the dose; fewer aphids were detected on the CJ-treated plants than on untreated control plants by direct visual counting, sweep net and yellow sticky trap samplings. CJ treatment also had deterrent effects on phytophagous thrips species and wheat stem sawflies (Hymenoptera: Cephidae), whereas CJ was an attractant for aphid parasitoids, coccinellid species and the wheat stem sawfly parasitoid Collyria coxator (Hymenoptera: Ichneumonidae). CJ treatment had no effect on syrphid species (Diptera: Syrphidae). There was no significant effect of CJ treatment on wheat yield; however, CJ-treated plants were taller than untreated control plants.Although the behavioral responses of the studied species to CJ treatment varied with the plant growth stage and treatment dose, and were not consistent across all sampling methods, our findings indicate CJ treatment may be used to reduce some insect pest populations and manipulate some natural enemy species under field conditions.

HPLC Analysis and Biochemical Characterization of LOX from Eschscholtzia californica Cham.

Background: Plant lipoxygenases (LOXs, EC 1.13.11.12) are involved in lipid degradation, regulation of growth and development, senescence, and defence reactions. LOX represents the starting enzyme of the octadecanoid pathway. The aim of the work was to purify LOX from California poppy (Eschscholtzia californica Cham.), to determine its biochemical properties and to identify and quantify the products of LOX reaction with unsaturated fatty acids. Methods: LOX from California poppy seedlings was purified by hydrophobic chromatography (Phenyl-Sepharose CL-4B) and by ion-exchange chromatography (Q-Sepharose). The isolated LOX was incubated with linoleic acid used as a substrate. The HPLC experiments were performed with the Agilent Technologies 1050 series HPLC system. For the preparative separation of a mixture of hydroxy fatty acids from the sample matrix, the RP-HPLC method was used (column 120-5 Nucleosil C18). Then, the NP-HPLC analysis (separation, identification, and determination) of hydroxy fatty acid isomers was carried out on a Zorbax Rx-SIL column. Results: The purified LOX indicates the presence of a nontraditional plant enzyme with dual positional specificity (a ratio of 9- and 13-hydroperoxide products 1:1), a relative molecular mass of 85 kDa, a pH optimum of 6.5, an increasing activity stimulation by CaCl2 till 2 mM, and a high substrate reactivity to linoleic acid with kinetic values of KM 2.6 mM and Vmax 3.14 μM/min/mg. Conclusions: For the first time, the LOX from California poppy seedlings was partially purified and the biochemical properties of the enzyme were analyzed. A dual positional specificity of the LOX found from California poppy seedlings is in agreement with the results obtained for LOXs isolated from other Papaveraceaes. A 1:1 ratio of 9-/13-HODE is attractive for the simultaneous investigation of both biotic stress responses (indicated by the 9-HODE marker) and the biosynthesis of jasmonic acid and jasmonates (indicated by the 13-HODE marker).

Covalent immobilization of oxylipin biosynthetic enzymes on nanoporous rice husk silica for production of cis(+)-12-oxophytodienoic acid

Soybean lipoxygenase, recombinant rice allene oxide synthase-1 and rice allene oxide cyclase were covalently immobilized on nanoporous rice husk silica using two types of linkers: glutardialdehyde and polyethylene glycol. The immobilization efficiency achieved using glutardialdehyde-linked rice husk silica was higher than that achieved using polyethylene glycol-linked rice husk silica (50–92% and 25–50%, respectively). Immobilization on both types of matrices significantly decreased the specific activities of the immobilized enzymes. Solid-phase reaction yields of the enzymes were determined relative to the yields observed for the solution-phase reactions. Yields of the solid-phase reactions catalyzed by immobilized soybean lipoxygenase, rice allene oxide synthase-1, and rice allene oxide cyclase ranged from 50% to 230% and were dependent on both the enzymes and linkers used. Production of cis(+)-12-oxophytodienoic acid from α-linolenic acid by consecutive reactions using all three enzymes in a co-immobilization system resulted in 83.6% and 65.1% yields on glutardialdehyde-linked and epichlorohydrin-polyethylene glycol-linked rice husk silica, respectively. Our results suggest that immobilization of biosynthetic enzymes of the octadecanoid pathway on rice husk silica may be an efficient method for the in vitro production of oxylipins. Additionally, enzyme immobilizations on rice husk silica matrices may be more broadly applicable for producing physiologically important compounds in other biosynthetic pathways.

Homologs of old yellow enzyme in plants

Old Yellow Enzyme (OYE) is a flavin mononucleotide-dependent oxidoreductase. 12-oxophytodienoic acid reductases (OPRs) are OYE homologs and are represented by multigene families in plants. OPRs catalyze the reduction of double bonds in α,β-unsaturated aldehydes or ketones. They belong to the octadecanoid pathway, which converts linolenic acid to jasmonic acid (JA). Individual OPR family members may have distinct functions due to their different substrate specificity, subcellular localization, tissue distribution, and differential regulation of their expression in response to specific environmental cues. Based on their differential preferences for 12-oxophytodienoate (OPDA) stereoisomers, these enzymes are classified into two subgroups: OPRI and OPRII. OPRI enzymes preferentially catalyze the reduction of cis-(−)OPDA over the JA precursor cis-(+)OPDA , and therefore, they are not involved in JA biosynthesis. Although a significant progress has been made to understand the exact physiological roles of OPR enzymes, the function of OPRI subgroup members in plants remains largely unknown. Enzymes belonging to this subgroup are possibly involved in defense responses and signaling. The members of the OPRII subgroup are required for JA biosynthesis because they catalyze the reduction of the JA precursor cis-(+)OPDA. This review will highlight some characteristics of this family in Arabidopsis and other species and discuss the physiological role of OPR family members in plants.

Phosphatidic acid synthesis, octadecanoic pathway and fatty acids content as lipid markers of exogeneous salicylic acid-induced elicitation in wheat.

Activators of plant defence responses against pathogens are a potential alternative to fungicides, and the well-known resistance inducer salicylic acid (SA) protects wheat (Triticum aestivum L.) against powdery mildew. The aim of our work was to investigate through biochemical and molecular approaches whether lipid metabolism alteration could be considered as a characteristic feature of induced resistance in wheat upon SA infiltration. Expression levels of lox, PI-PLC2 and ltp genes encoding for a lipoxygenase (LOX), a phospholipase C2 and a lipid-transfer protein, respectively, were investigated. Increase of phosphatidic acid (PA) content 48h after SA infiltration in wheat leaves, upregulation of PI-PLC2 gene expression and increased diacylglycerol content were recorded, indicating the involvement of the PLC pathway in the PA synthesis. The wheat octadecanoid pathway was shown to be highly responsive to SA infiltration through simultaneous increases in lox gene expression and LOX activity, as well as a reduction in the content of linolenic acid. Changes in several FA contents and increases of the ltp gene expression were also recorded during the latest hours after SA infiltration. The status of lipid metabolism, as well as the connections between its components as markers of SA-induced resistance in wheat, are discussed.

Hydrogen Peroxide Acts as a Signaling Molecule for the Methyl Jasmonate-Induced Antioxidant Defense in Wheat Callus to Promote Enhanced Drought Tolerance

Drought stress is a major challenge in agriculture, causing severe loss of crops throughout the world. Methyl jasmonate (MeJA) regulates a variety of plant developmental process and responds to many biotic and abiotic stresses. To explore the physiological mechanisms of drought stress mitigated by exogenous MeJA, we subjected callus of wheat (Triticum aestivum L.) to drought induced medium after pretreatment with 10 ml sterile solution that contained the following singly or in combinations: MeJA (0, 0.25, and 2.5µM) and the lipoxygenase (LOX) inhibitor salicylhydroxamic acid (SHAM; 0 and 0.5 mM) for 24 hours. The endogenous jasmonic acid (JA) content, activity of LOX, hydrogen peroxide (H2O2) content, antioxidant enzymes activities, levels of reactive oxygen species (ROS), content of malondialdehyde (MDA), and cell viability were measured. The results showed that exogenous MeJA induced the activity of LOX and resulted in rapid increase in endogenous JA levels in a concentration-dependent manner; JA declined gradually after reaching its maximum. The level of H2O2 increased with increases in LOX activity and endogenous JA level, which are involved in the octadecanoid signaling pathway. However, SHAM inhibited the LOX activity, the endogenous JA level, and the accumulation of H2O2 in wheat callus. In addition, the activities of several antioxidant enzymes increased after MeJA-pretreatment, but this effect was also inhibited by SHAM. The cumulative results suggest that H2O2 was generated through the octadecanoid signaling pathway, and some antioxidant defense genes were provoked by the accumulation of H2O2. Therefore, we inferred that H2O2 might act as a signaling molecule for the MeJA-induced antioxidant defense. Drought stress can be improved by MeJA-pretreatment, leading to decrease in endogenous MDA and ROS contents, which are normally induced by drought conditions. Both drought stress and exogenous MeJA-pretreatment increased in LOX activity, endogenous JA level, and antioxidant enzymes. Combinatorial treatments showed an apparent synergistic effect on the activities of these antioxidant enzymes and resulted in improved cell viability. These results confirmed the hypothesis that H2O2 acts as a signaling molecule for the MeJA-induced antioxidant defense and could alleviate the negative effects of drought stress on wheat callus.

Nitric oxide: a signaling molecule which activates cell wall-associated defense of tomato against Rhizoctonia solani

We investigated the role of sodium nitroprusside (SNP), as a NO donor, in activating cell wall-related defense responses of tomato against R. solani. Based on previous knowledge on the cross talk of NO with various pathways and its effect on cell wall components, experiments were carried out to investigate the function of cell wall-related defense responses, octadecanoid and phenylpropanoid pathways (which are associated with cell wall modification) not only in SNP-activated defense but also in basal resistance of tomato to the pathogen. In detached leaves and intact plants, we observed a considerable decrease in disease progress on both partially resistant (CH Falat) and susceptible (Mobil) tomato cultivars treated with SNP. The SNP treatment regulated malondialdehyde (MDA), H2O2 and O2 −levels in plant cells, whereas SNP primed callose depsition, phenolics and lignification, as defense responses related to cell wall modification in the inoculated tomato plants. Priming in the activity and expression of lipoxygenase (LOX) and phenylalanine ammonia-lyase (PAL) as key markers of octadecanoid and phenylpropanoid pathways, respectively, was observed in SNP-treated inoculated plants. Co-application of LOX or PAL inhibitors and SNP completely suppressed SNP-activated defense responses. Using DEA-NONOate, as an independent NO releaser, supported the data obtained by SNP on disease progress and the role of PAL and LOX in NO-related protection of tomato against the pathogen. These findings highlight the role of phenylpropanoid and octadecanoid pathways and involvement of cell wall modifications in NO-associated induction of defense as novel mechanisms of SNP-IR in tomato–R. solani pathosystem.