Root-derived oxylipins promote green peach aphid performance on Arabidopsis foliage

Oxylipins function as signaling molecules in plant growth and development and contribute to defense against stress. Here, we show that oxylipins also facilitate infestation of Arabidopsis thaliana shoots by the phloem sap-consuming green peach aphid (GPA; Myzus persicae), an agronomically important insect pest. GPAs had difficulty feeding from sieve elements and tapping into the xylem of lipoxygenase5 (lox5) mutant plants defective in LOX activity. These defects in GPA performance in the lox5 mutant were accompanied by reduced water content of GPAs and a smaller population size of GPAs in the mutant compared with the wild-type plant. LOX5 expression was rapidly induced in roots in response to infestation of shoots by GPAs. In parallel, levels of LOX5-derived oxylipins increased in roots and in petiole exudates of GPA-colonized plants. Application of 9-hydroxyoctadecadienoic acid (an oxylipin produced by the LOX5 enzyme) to roots restored water content and GPA population size in lox5 plants, thus confirming that a LOX5-derived oxylipin promotes infestation of the foliage by GPAs. Micrografting experiments demonstrated that GPA performance on foliage is influenced by the LOX5 genotype in roots, thus demonstrating the importance of root-derived oxylipins in colonization of aboveground organs by an insect.     

Evaluation of the antimicrobial activities of plant oxylipins supports their involvement in defense against pathogens

Plant oxylipins are a large family of metabolites derived from polyunsaturated fatty acids. The characterization of mutants or transgenic plants affected in the biosynthesis or perception of oxylipins has recently emphasized the role of the so-called oxylipin pathway in plant defense against pests and pathogens. In this context, presumed functions of oxylipins include direct antimicrobial effect, stimulation of plant defense gene expression, and regulation of plant cell death. However, the precise contribution of individual oxylipins to plant defense remains essentially unknown. To get a better insight into the biological activities of oxylipins, in vitro growth inhibition assays were used to investigate the direct antimicrobial activities of 43 natural oxylipins against a set of 13 plant pathogenic microorganisms including bacteria, oomycetes, and fungi. This study showed unequivocally that most oxylipins are able to impair growth of some plant microbial pathogens, with only two out of 43 oxylipins being completely inactive against all the tested organisms, and 26 oxylipins showing inhibitory activity toward at least three different microbes. Six oxylipins strongly inhibited mycelial growth and spore germination of eukaryotic microbes, including compounds that had not previously been ascribed an antimicrobial activity, such as 13-keto-9(Z),11(E),15(Z)-octadecatrienoic acid and 12-oxo-10,15(Z)-phytodienoic acid. Interestingly, this first large-scale comparative assessment of the antimicrobial effects of oxylipins reveals that regulators of plant defense responses are also the most active oxylipins against eukaryotic microorganisms, suggesting that such oxylipins might contribute to plant defense through their effects both on the plant and on pathogens, possibly through related mechanisms.     

Biosynthesis of oxylipins in non-mammals

Lipid peroxidation is common to all biological systems, appearing in developmentally-regulated processes and as a response to environmental changes. Products derived from lipid peroxidation are collectively named oxylipins. Initial lipid peroxidation may either occur by enzymatic or chemical reactions. An array of alternative reactions further converting lipid hydroperoxides gives rise to a large variety of oxylipin classes, some with reported signaling functions in plants, fungi, algae or animals. The structural diversity of oxylipins is further increased by their occurrence either as esters in complex lipids or as free (non-esterified) fatty acid derivatives. The enzymes involved in oxylipin metabolism are diverse and comprise a multitude of examples with interesting and unusual catalytic properties. This review aims at giving an overview on plant, fungal, algal and bacterial oxylipins and the enzymes responsible for their biosynthesis.     

The green peach aphid,Myzus persicae,acquires a LIPOXYGENASE5-derived oxylipin from Arabidopsis thaliana,which promotes colonization of the host plant

Oxylipins derived from lipoxygenase (LOX) activity play important roles in plant growth, development and stress response. In a recent study, we provided evidence that infestation of Arabidopsis thaliana foliage by the green peach aphid (GPA; Myzus persicae), a phloem sap-consuming insect, was promoted by plant LOX5-derived oxylipins. In comparison to the wild-type (WT) plant, GPA population was smaller on the Arabidopsis lox5 mutant. The insect spent less time feeding from the sieve element and xylem of the lox5 mutant compared with the WT plant. In addition, compared with insects feeding on the WT plant, when on the lox5 mutant, the GPA was unable to suppress an antibiotic activity that is present in Arabidopsis vascular sap. Roots are the critical source of a LOX5-derived oxylipin(s) that promotes colonization of the foliage by GPA. Here we show that the 9-hydoxy-10E, 12Z-octadecadienoic acid (9-HOD), a LOX5-derived oxylipin, accumulated in GPA that were reared on the WT, but not the lox5 mutant plant. However, 9-HOD accumulated in insects reared on lox5 mutant plants that were irrigated with 9-HOD, thus indicating that the insect ingests oxylipins from the host plant. We further demonstrate that the host plant requires LOX5 function to promote expression of the defense regulatory gene PHYTOALEXIN-DEFICIENT4 in the foliage. Taken together, our previous observations and results presented here indicate that while the host plant utilizes LOX5-dependent factors for promoting defense mechanisms, GPA has evolved to utilize plant 9-LOX-derived oxylipins as cues to facilitate infestation, thus suggesting a complex involvement of oxylipins in Arabidopsis interaction with GPA.     

Oxylipins and plant abiotic stress resistance

Oxylipins are signaling molecules formed enzymatically or spontaneously from unsaturated fatty acids in all aerobic organisms. Oxylipins regulate growth, development, and responses to environmental stimuli of organisms. The oxylipin biosynthesis pathway in plants includes a few parallel branches named after first enzyme of the corresponding branch as allene oxide synthase, hydroperoxide lyase, divinyl ether synthase, peroxygenase, epoxy alcohol synthase, and others in which various biologically active metabolites are produced. Oxylipins can be formed non-enzymatically as a result of oxygenation of fatty acids by free radicals and reactive oxygen species. Spontaneously formed oxylipins are called phytoprostanes. The role of oxylipins in biotic stress responses has been described in many published works. The role of oxylipins in plant adaptation to abiotic stress conditions is less studied; there is also obvious lack of available data compilation and analysis in this area of research. In this work we analyze data on oxylipins functions in plant adaptation to abiotic stress conditions, such as wounding, suboptimal light and temperature, dehydration and osmotic stress, and effects of ozone and heavy metals. Modern research articles elucidating the molecular mechanisms of oxylipins action by the methods of biochemistry, molecular biology, and genetics are reviewed here. Data on the role of oxylipins in stress signal transduction, stress-inducible gene expression regulation, and interaction of these metabolites with other signal transduction pathways in cells are described. In this review the general oxylipin-mediated mechanisms that help plants to adjust to a broad spectrum of stress factors are considered, followed by analysis of more specific responses regulated by oxylipins only under certain stress conditions. New approaches to improvement of plant resistance to abiotic stresses based on the induction of oxylipin-mediated processes are discussed.     

Disease resistance against Magnaporthe grisea is enhanced in transgenic rice with suppression of omega-3 fatty acid desaturases

Linolenic acid (18:3) is the most abundant fatty acid in plant membrane lipids and is a source for various oxidized metabolites, called oxylipins. 18:3 and oxylipins play important roles in the induction of defense responses to pathogen infection and wound stress in Arabidopsis. However, in rice, endogenous roles for 18:3 and oxylipins in disease resistance have not been confirmed. We generated 18:3-deficient transgenic rice plants (F78Ri) with co-suppression of two omega-3 fatty acid desaturases, OsFAD7 and OsFAD8. that synthesize 18:3. The F78Ri plants showed enhanced resistance to the phytopathogenic fungus Magnaporthe grisea. A typical 18:3-derived oxylipin, jasmonic acid (JA), acts as a signaling molecule in defense responses to fungal infection in Arabidopsis. However, in F78Ri plants, the expression of JA-responsive pathogenesis-related genes, PBZ1 and PR1b, was induced after inoculation with M. grisea, although the JA-mediated wound response was suppressed. Furthermore, the application of JA methyl ester had no significant effect on the enhanced resistance in F78Ri plants. Taken together, our results indicate that, although suppression of fatty acid desaturases involves the concerted action of varied oxylipins via diverse metabolic pathways, 18:3 or 18:3-derived oxylipins, except for JA, may contribute to signaling on defense responses of rice to M. grisea infection.     

Oxylipins in fungi

In nearly every living organism, metabolites derived from lipid peroxidation, the so-called oxylipins, are involved in regulating developmental processes as well as environmental responses. Among these bioactive lipids, the mammalian and plant oxylipins are the best characterized, and much information about their physiological role and biosynthetic pathways has accumulated during recent years. Although the occurrence of oxylipins and enzymes involved in their biosynthesis has been studied for nearly three decades, knowledge about fungal oxylipins is still scarce as compared with the situation in plants and mammals. However, the research performed so far has shown that the structural diversity of oxylipins produced by fungi is high and, furthermore, that the enzymes involved in oxylipin metabolism are diverse and often exhibit unusual catalytic activities. The aim of this review is to present a synopsis of the oxylipins identified so far in fungi and the enzymes involved in their biosynthesis.     

Methods for the analysis of oxylipins in plants

Plant oxylipins comprise a highly diverse and complex class of molecules that are derived from lipid oxidation. The initial oxidation of unsaturated fatty acids may either occur by enzymatic or chemical reactions. A large variety of oxylipin classes are generated by an array of alternative reactions further converting hydroperoxy fatty acids. The structural diversity of oxylipins is further increased by their occurrence either as free fatty acid derivatives or as esters in complex lipids. Lipid peroxidation is common to all biological systems, appearing in developmentally regulated processes and as a response to environmental changes. The oxylipins formed may perform various biological roles; some of them have signaling functions. In order to elucidate the roles of oxylipins in a given biological context, comprehensive analytical assays are available for determining the oxylipin profiles of plant tissues. This review summarizes indirect methods to estimate the general peroxidation state of a sample and more sophisticated techniques for the identification, structure determination and quantification of oxylipins.     

Organ-dependent oxylipin signature in leaves and roots of salinized tomato plants (Solanum lycopersicum)

Oxylipins have been extensively studied in plant defense mechanisms or as signal molecules. Depending on the stress origin (e.g. wounding, insect, pathogen), and also on the plant species or organ, a specific oxylipin signature can be generated. Salt stress is frequently associated with secondary stress such as oxidative damage. Little is known about the damage caused to lipids under salt stress conditions, especially with respect to oxylipins. In order to determine if an organ-specific oxylipin signature could be observed during salt stress, tomato (Solanum lycopersicum cv. Money Maker) plants were submitted to salt stress (100 mM of NaCl) for a 30-d period. A complete oxylipin profiling and LOX related-gene expression measurement were achieved in leaves and roots. As expected, salt stress provoked premature senescence in leaves, as revealed by a decrease in photosystem II efficiency (F(v)/F(m) ratio) and sodium accumulation in leaves. In roots, a significant decrease in several oxylipins (9- and 13-hydro(pero)xy linole(n)ic acids, keto and divinyl ether derivatives) was initiated at day 5 and intensified at day 21 after salt treatment, whereas jasmonic acid content increased. In leaves, the main changes in oxylipins were observed later (at day 30), with an increase in some 9- and 13-hydro(pero)xy linole(n)ic acids and a decrease in some keto-derivatives and in jasmonic acid. Oxylipin enantiomeric characterization revealed that almost all compounds were formed enzymatically, and therefore a massive auto-oxidation of lipids that can be encountered in abscission processes can be excluded here.     

Oxylipins: Structurally diverse metabolites from fatty acid oxidation

Oxylipins are lipophilic signaling molecules derived from the oxidation of polyunsaturated fatty acids. Initial fatty acid oxidation occurs mainly by the enzymatic or chemical formation of fatty acid hydroperoxides. An array of alternative reactions further converting fatty acid hydroperoxides gives rise to a multitude of oxylipin classes, many with reported signaling functions in plants. Oxylipins include the phytohormone, jasmonic acid, and a number of other molecules including hydroxy-, oxo- or keto-fatty acids or volatile aldehydes that may perform various biological roles as second messengers, messengers in inter-organismic signaling, or even as bactericidal agents. The structural diversity of oxylipins is further increased by esterification of the compounds in plastidial glycolipids, for instance the Arabidopsides, or by conjugation of oxylipins to amino acids or other metabolites. The enzymes involved in oxylipin metabolism are diverse and comprise a multitude of examples with interesting and unusual catalytic properties. In addition, the interplay of different subcellular compartments during oxylipin biosynthesis suggests complex mechanisms of regulation that are not well understood. This review aims at giving an overview of plant oxylipins and the multitude of enzymes responsible for their biosynthesis.     

Wounding stimulates the accumulation of glycerolipids containing oxophytodienoic acid and dinor-oxophytodienoic acid in Arabidopsis leaves

Although oxylipins can be synthesized from free fatty acids, recent evidence suggests that oxylipins are components of plastid-localized polar complex lipids in Arabidopsis (Arabidopsis thaliana). Using a combination of electrospray ionization (ESI) collisionally induced dissociation time-of-flight mass spectrometry (MS) to identify acyl chains, ESI triple-quadrupole (Q) MS in the precursor mode to identify the nominal masses of complex polar lipids containing each acyl chain, and ESI Q-time-of-flight MS to confirm the identifications of the complex polar lipid species, 17 species of oxylipin-containing phosphatidylglycerols, monogalactosyldiacylglycerols (MGDG), and digalactosyldiacylglycerols (DGDG) were identified. The oxylipins of these polar complex lipid species include oxophytodienoic acid (OPDA), dinor-OPDA (dnOPDA), 18-carbon ketol acids, and 16-carbon ketol acids. Using ESI triple-Q MS in the precursor mode, the accumulation of five OPDA- and/or dnOPDA-containing MGDG and two OPDA-containing DGDG species were monitored as a function of time in mechanically wounded leaves. In unwounded leaves, the levels of these oxylipin-containing complex lipid species were low, between 0.001 and 0.023 nmol/mg dry weight. However, within the first 15 min after wounding, the levels of OPDA-dnOPDA MGDG, OPDA-OPDA MGDG, and OPDA-OPDA DGDG, each containing two oxylipin chains, increased 200- to 1,000-fold. In contrast, levels of OPDA-hexadecatrienoic acid MGDG, linolenic acid (18:3)-dnOPDA MGDG, OPDA-18:3 MGDG, and OPDA-18:3 DGDG, each containing a single oxylipin chain, rose 2- to 9-fold. The rapid accumulation of high levels of galactolipid species containing OPDA-OPDA and OPDA-dnOPDA in wounded leaves is consistent with these lipids being the primary products of plastidic oxylipin biosynthesis.     

Auxin-oxylipin crosstalk: relationship of antagonists

Stephan Pollmann (Corresponding author) Phytohormones regulate a wide array of developmental processes throughout the life cycle of plants. Herein, the various plant hormones may interact additively, synergistically, or antagonistically. By their cooperation they create a delicate regulatory network whose net output largely depends on the action of specific phytohormone combinations rather than on the independent activities of separate hormones. While most classical studies of plant hormonal control have focused mainly on the action of single hormones or on the synergistic interaction of hormones in regulating various developmental processes, recent work is beginning to shed light on the crosstalk of nominally antagonistic plant hormones, such as gibberellins and auxins with oxylipins or abscisic acid. In this review, we summarize our current understanding of how two of the first sight antagonistic plant hormones, i.e. auxins and oxylipins, interact in controlling plant responses and development.     

Constitutive expression of an inducible lipoxygenase in transgenic tobacco decreases susceptibility to Phytophthora parasitica var. nicotianae

Plant lipoxygenases (LOXs) are key enzymes involved in the generation of fatty acid derivatives, called oxylipins. In tobacco, LOX gene expression and activity are very low in healthy tissues and are highly enhanced in response to infection by Phytophthora parasitica nicotianae and to elicitor treatment. We previously showed, using antisense-LOX1 plants, that expression of the tobacco LOX1 gene is required for the race-cultivar specific resistance of tobacco to Phytophthora parasitica nicotianae. In order to investigate the effect of over-expressing a LOX gene on plant resistance, we transformed tobacco plants with the LOX1 coding sequence fused to the CaMV 35S promoter. Four transgenic lines with enhanced levels of LOX protein and specific activity over control plants were selected for further analysis. These plants were macroscopically indistinguishable from WT plants. Upon stem inoculation, the sense-LOX1 plants displayed a significantly decreased susceptibility to virulent races of Phytophthora parasitica nicotianae, stem lesions being 2- to 3-fold shorter in the transgenic lines than in WT plants. Using a root inoculation assay, the survival rate of sense-LOX1 seedlings was increased about 4-fold compared to their WT counterparts, with 60 to 80% of transgenic plants vs 15 to 20% of WT controls remaining healthy following inoculation with Phytophthora parasitica nicotianae. This is the first demonstration that the over-expression of a LOX gene is sufficient to reduce the susceptibility of a host plant to an oomycete pathogen.     

Jasmonates and Related Compounds in Plant-Insect Interactions

Herbivore attack elicits defense responses in host plants by a complex chain of events that starts with the introduction of herbivore-specific elicitors into the wounds at the feeding or oviposition site, their recognition by the plant, and activation of several signaling cascades that trigger defense responses that finally increase resistance. Oxylipin signaling plays a central role in the activation of these herbivore-induced responses. Wounding activates some but not all of these defense responses, but herbivore attack frequently amplifies the oxylipin responses well beyond that elicited by wounding alone, suggesting recognition of herbivore attack. In addition to their signaling role within the plant, oxylipins can also directly influence the performance of herbivores or attract natural enemies to feeding herbivores. Here we review the literature on the regulation and function of herbivore-specific oxylipin signaling and the direct effects of oxylipins on herbivore performance. Online publication: 13 January 2005     

Growth, nitrogen fixation and ammonium assimilation in common bean (Phaseolus vulgaris): effect of phosphorus

The impact of phosphorous nutrition on plant growth, symbiotic N₂ fixation, ammonium assimilation, carbohydrate and amino-acid accumulation, as well as on nitrogen, phosphorus and ATP content in tissues in common bean ( Phaseolus vulgaris ) plants was investigated. Plants inoculated with Rhizobium tropici CIAT899 were grown in Leonard jars under controlled conditions, with P-deficient (0 and 0.1 m M ), P-medium (0.5, 1 and 1.5 m M ) and P-high (2 m M ) conditions in a N-free nutrient solution. The P application, increased leaf area, whole plant DW (67%), nodule biomass (4-fold), and shoot and root P content (4- and 6-fold, respectively) in plant harvested at the onset of flowering (28-days-old). However, P treatments decreased the total soluble sugar and amino acid content in vegetative organs (leaf, root and nodules). The root growth proved less sensitive to P deficiency than did shoot growth, and the leaf area was significantly reduced at low P-application. The absence of a relationship between shoot N content, and P levels in the growth medium could indicate that nitrogen fixation requires more P than does plant growth. The optimal amount for the P. vulgaris – R. tropici CIAT899 symbiosis was 1.5 m M P, this treatment augmented nodule-ARA 20-fold, and ARA per plant 70-fold compared with plants without P application.     

Oxylipin profiling of the hypersensitive response in Arabidopsis thaliana. Formation of a novel oxo-phytodienoic acid-containing galactolipid, arabidopside E

Oxidation products of unsaturated fatty acids, collectively known as oxylipins, function as signaling molecules in plants during development, wounding, and insect and pathogen attack. Certain oxylipins are also known to have direct cytotoxic effects on pathogens. We used inducible expression of bacterial avirulence proteins in planta to study the involvement of oxylipins in race-specific defense against bacterial pathogens. We demonstrate that recognition of the Pseudomonas syringae avirulence protein AvrRpm1 induces 9- and 13-lipoxygenase-dependent oxylipin synthesis in Arabidopsis thaliana. The major oxylipins accumulated were jasmonic acid, 12-oxo-phytodienoic acid, and dinor-oxo-phytodienoic acid. The majority of the newly formed oxylipins (>90%) was found to be esterified to glycerolipids, whereby 12-oxo-phytodienoic acid and dinor-oxo-phytodienoic acid were found to be esterified to a novel galactolipid. The structure of the substance was determined as a monogalactosyldiacylglycerol containing two 12-oxo-phytodienoic acids and one dinor-oxo-phytodienoic acid acyl chain and was given the trivial name arabidopside E. This substance accumulated to surprisingly high levels, 7-8% of total lipid content, and was shown to inhibit growth of a bacterial pathogen in vitro. Arabidopside E was formed also after recognition of the avirulence protein AvrRpt2, suggesting that this could be a conserved feature of defense reactions against bacterial pathogens. In conclusion, the data presented suggest a role of enzymatically formed oxylipins, especially the octadecanoids and arabidopside E in race-specific resistance against bacterial pathogens.     

Regulation of oxylipin synthesis

Two very common groups of oxylipins formed in plants involve the conversion of fatty acid hydroperoxides, such as hydroperoxy-octadecatrienoic acid, into further metabolites by allene oxide synthase and hydroperoxide lyase. Both of these oxylipin branch pathways appear to be ubiquitous or nearly so in plants, but the relative activities of these two branches vary among plant species. In most plants examined, including Arabidopsis, product formation from either of these pathways is minimal until elicited by wounding or some other means, upon which products from both pathways, such as jasmonic acid and C(6) aldehydes and alcohols, can increase by orders of magnitude. In some plant species such as Artemisia and Jasminum spp. oxylipin product formation is heavily skewed towards allene oxide synthase products. Others such as watermelon (Citrullus lanatus) produce 10-fold higher amounts or more of hydroperoxide lyase than allene oxide synthase products. Arabidopsis and tobacco are intermediate between these extremes. Artemisia and Jasminum are also unusual in that they do not require wounding or other types of induction for high oxylipin product formation. Release of non-esterified fatty acids appears to be correlated with oxylipin formation, but phospholipase A(2) appears not to be involved with oxylipin production, at least in the case of Artemisia leaves.     

Production of cross-kingdom oxylipins by pathogenic fungi: An update on their role in development and pathogenicity

Oxylipins are a class of molecules derived from the incorporation of oxygen into polyunsaturated fatty acid substrates through the action of oxygenases. While extensively investigated in the context of mammalian immune responses, over the last decade it has become apparent that oxylipins are a common means of communication among and between plants, animals, and fungi to control development and alter hostmicrobe interactions. In fungi, some oxylipins are derived nonenzymatically while others are produced by lipoxygenases, cyclooxygenases, and monooxygenases with homology to plant and human enzymes. Recent investigations of numerous plant and human fungal pathogens have revealed oxylipins to be involved in the establishment and progression of disease. This review highlights oxylipin production by pathogenic fungi and their role in fungal development and pathogen/host interactions.