Enhancement of disease resistance to Magnaporthe grisea in rice by accumulation of hydroxy linoleic acid

Linoleic acid (18:2) and linolenic acid (18:3) are sources for various oxidized metabolites called oxylipins, some of which inhibit growth of fungal pathogens. In a previous study, we found disease resistance to rice blast fungus Magnaporthe grisea enhanced in 18:2-accumulating transgenic rice (F78Ri) in which the conversion from 18:2 to 18:3 was suppressed. Here, we demonstrate that 18:2-derived hydroperoxides and hydroxides (HPODEs and HODEs, respectively) inhibit growth of M. grisea more strongly than their 18:3-derived counterparts. Furthermore, in F78Ri plants, the endogenous levels of HPODEs and HODEs increased significantly, compared with wild-type plants. These results suggest that the increased accumulation of antifungal oxylipins, such as HPODEs and HODEs, causes the enhancement of disease resistance against M. grisea.     

Resistance to Magnaporthe grisea in transgenic rice with suppressed expression of genes encoding allene oxide cyclase and phytodienoic acid reductase

Linolenic acid (18:3) and its derivative jasmonic acid (JA) are important molecules in disease resistance in many dicotyledonous plants. We have previously used 18:3- and JA-deficient rice (F78Ri) to investigate the roles of fatty acids and their derivatives in resistance to the blast fungus Magnaporthe grisea [A. Yara, T. Yaeno, J.-L. Montillet, M. Hasegawa, S. Seo, K. Kusumi, K. Iba, Enhancement of disease resistance to Magnaporthe grisea in rice by accumulation of hydroxy linoleic acid, Biochem. Biophys. Res. Commun. 370 (2008) 344-347; A. Yara, T. Yaeno, M. Hasegawa, H. Seto, J.-L. Montillet, K. Kusumi, S. Seo, K. Iba, Disease resistance against Magnaporthe grisea is enhanced in transgenic rice with suppression of ω-3 fatty acid desaturases, Plant Cell Physiol. 48 (2007) 1263-1274]. However, because F78Ri plants are suppressed in the first step of the JA biosynthetic pathway, we could not confirm the specific contribution of JA to disease resistance. In this paper, we generated two JA-deficient rice lines (AOCRi and OPRRi) with suppressed expression of the genes encoding allene oxide cyclase (AOC) and 12-oxo-phytodienoic acid reductase (OPR), which catalyze late steps in the JA biosynthetic pathway. The levels of disease resistance in the AOCRi and OPRRi lines were equal to that in wild-type plants. Our data suggest that resistance to M. grisea is not dependent on JA synthesis.     

Probenazole induces systemic acquired resistance in Arabidopsis with a novel type of action

Probenazole (PBZ; 3-allyloxy-1,2-benzisothiazole-1,1-dioxide), which is the active ingredient in Oryzemate, has been used widely in Asia to protect rice plants against the rice blast fungus Magnaporthe grisea. To study PBZ's mode of action, we analyzed its ability, as well as that of its active metabolite 1, 2-benzisothiazol-3 (2H)-one 1,1-dioxide (BIT) to induce defense gene expression and resistance in Arabidopsis mutants that are defective in various defense signaling pathways. Wild-type Arabidopsis treated with PBZ or BIT exhibited increased expression of several pathogenesis-related genes, increased levels of total salicylic acid (SA), and enhanced resistance to the bacterial pathogen Pseudomonas syringae pv. tomato DC 3000 and the oomycete pathogen Peronospora parasitica Emco5. The role of several defense signaling hormones, such as SA, ethylene and jasmonic acid (JA), in activating resistance following PBZ or BIT treatment was analyzed using NahG transgenic plants and etr1-1 and coi1-1 mutant plants, respectively. In addition, the involvement of NPR1, a key component in the SA signaling pathway leading to defense responses, was assessed. PBZ or BIT treatment did not induce disease resistance or PR-1 expression in NahG transgenic or npr1 mutant plants, but it did activate these phenomena in etr1-1 and coi 1-1 mutant plants. Thus SA and NPR1 appear to be required for PBZ- and BIT-mediated activation of defense responses, while ethylene and JA are not. Furthermore, our data suggest that PBZ and BIT comprise a novel class of defense activators that stimulate the SA/NPR1-mediated defense signaling pathway upstream of SA.     

The plant growth-promoting fungus Penicillium simplicissimum GP17-2 induces resistance in Arabidopsis thaliana by activation of multiple defense signals

Arabidopsis thaliana grown in soil amended with barley grain inocula of Penicillium simplicissimum GP17-2 or receiving root treatment with its culture filtrate (CF) exhibited clear resistance to Pseudomonas syringae pv. tomato DC3000 (Pst). To assess the contribution of different defense pathways, Arabidopsis genotypes implicated in salicylic acid (SA) signaling expressing the NahG transgene or carrying disruption in NPR1 (npr1), jasmonic acid (JA) signaling (jar1) and ethylene (ET) signaling (ein2) were tested. All genotypes screened were protected by GP17-2 or its CF. However, the level of protection was significantly lower in NahG and npr1 plants than it was in similarly treated wild-type plants, indicating that the SA signaling pathway makes a minor contribution to the GP17-2-mediated resistance and is insufficient for a full response. Examination of local and systemic gene expression revealed that GP17-2 and its CF modulate the expression of genes involved in both the SA and JA/ET signaling pathways. Subsequent challenge of GP17-2-colonized plants with Pst was accompanied by direct activation of SA-inducible PR-2 and PR-5 genes as well as potentiated expression of the JA-inducible Vsp gene. In contrast, CF-treated plants infected with Pst exhibited elevated expression of most defense-related genes (PR-1, PR-2, PR-5, PDF1.2 and Hel) studied. Moreover, an initial elevation of SA responses was followed by late induction of JA responses during Pst infection of induced systemic resistance (ISR)-expressing plants. In conclusion, we hypothesize the involvement of multiple defense mechanisms leading to an ISR of Arabidopsis by GP17-2.     

A rice microsomal delta-12 fatty acid desaturase can enhance resistance to cold stress in yeast and <Emphasis Type="Italic">Oryza sativa</Emphasis>

A full-length cDNA clone of OsFAD2, which encodes a Δ-12 fatty acid desaturase, the key enzyme for the conversion of oleic acid (18:1) into linoleic acid (18:2), was isolated from rice (Oryza sativa ssp. japonica) leaves. The deduced amino acid sequence of OsFAD2 displayed three histidine boxes characteristic of all membrane-bound desaturases, and possessed a C-terminal signal for endoplasmic reticulum retention. Phylogenetic analysis showed that OsFAD2 is grouped within plant housekeeping FAD2 sequences. Expression analysis by real-time PCR showed that the gene is expressed in all tissues of rice tested, including root, seed, stem, and leaf. In situ hybridization showed that OsFAD2 mRNA accumulated in leaf mesophyll cells and in root epidermis cells when exposed to 15°C for 4 days in dark conditions. When OsFAD2 was expressed in Saccharomyces cerevisiae, the cells could convert oleic acid to linoleic acid, which wild-type yeast cells cannot do, suggesting that the isolated gene encoded a functional FAD2 enzyme. Heterologous expression of OsFAD2 enhanced the yeast cells’ cold tolerance capacity compared to wild-type yeast. OsFAD2 was also shown to be a highly active desaturase when expressed in Xenopus oocytes. In addition, when the OsFAD2 gene was transferred into an Arabidopsis thaliana fad2-1 mutant, it effectively restored wild-type fatty acid composition and growth characteristics. Stress tolerance and light regulatory elements were identified in the predicted promoter of the OsFAD2 gene. Exogenously supplied hormone affected the level of FAD2 mRNA accumulation, accompanied by a change of content of di-unsaturated fatty acid species in rice leaves. Furthermore, OsFAD2 enhanced tolerance to low temperature when overexpressed in rice at the vegetative stage. More importantly, the 35S::OsFAD2 plants showed significantly enhanced cold tolerance at the reproductive stage, increasing grain yield by 46% over controls in the greenhouse under cold conditions. These results indicated that OsFAD2 is involved in fatty acid desaturation and maintenance of the membrane lipids balance in cells, and could improve the tolerance of yeast and rice to low temperature stress.     

Phytochrome chromophore deficiency leads to overproduction of jasmonic acid and elevated expression of jasmonate-responsive genes in Arabidopsis

An Arabidopsis mutant line named hy1-101 was isolated because it shows stunted root growth on medium containing low concentrations of jasmonic acid (JA). Subsequent investigation indicated that even in the absence of JA, hy1-101 plants exhibit shorter roots and express higher levels of a group of JA-inducible defense genes. Here, we show that the hy1-101 mutant has increased production of JA and its jasmonate-related phenotype is suppressed by the coi1-1 mutation that interrupts JA signaling. Gene cloning and genetic complementation analyses revealed that the hy1-101 mutant contains a mutation in the HY1 gene, which encodes a heme oxygenase essential for phytochrome chromophore biosynthesis. These results support a hypothesis that phytochrome chromophore deficiency leads to overproduction of JA and activates COI1-dependent JA responses. Indeed, we show that, like hy1-101, independent alleles of the phytochrome chromophore-deficient mutants, including hy1-100 and hy2 (CS68), also show elevated JA levels and constant expression of JA-inducible defense genes. We further provide evidence showing that, on the other hand, JA inhibits the expression of a group of light-inducible and photosynthesis-related genes. Together, these data imply that the JA-signaled defense pathway and phytochrome chromophore-mediated light signaling might have antagonistic effects on each other.     

Structure,chromosomal location and expression of a rice gene encoding the microsome ω-3 fatty acid desaturase

The -3 fatty acid desaturases are membrane-bound enzymes catalyzing the conversion of linoleic acid to linolenic acid in lipids, and are located both in the microsome and plastid envelopes as two different isoforms. A cDNA encoding the microsome -3 fatty acid desaturase (OsFAD3) and the corresponding genomic clone were isolated from rice (Oryza sativa L.). The OsFAD3 gene was composed of 8 exons and 7 introns. A microsatellite was present in the second exon of the OsFAD3 gene, showing polymorphism between Indica and Japonica rice varieties. The mapping of this microsatellite showed that the OsFAD3 gene was located on chromosome 11. Expression of the OsFAD3 cDNA in tobacco hairy root tissues and subsequent analysis of fatty acid compositions demonstrated the activity of the microsome -3 fatty acid desaturase. The OsFAD3 mRNA was abundant in root tissues, but was hardly detectable in leaves. In root tissues, a high level of the OsFAD3 mRNA was observed at 15 °C and 20 °C, with its level decreasing markedly at temperatures below 10 °C. The accumulation of the OsFAD3 mRNA in leaf tissues remained at quite low levels, both at normal growth temperatures and at chilling temperatures. Similar temperature responses of the OsFAD3 gene were observed both in chilling- tolerant and in chilling-intolerant rice cultivars.     

LOX genes in blast fungus (Magnaporthe grisea) resistance in rice

Plant Lipoxygenases (LOX) are known to play major role in plant immunity by providing front-line defense against pathogen-induced injury. To verify this, we isolated a full-length OsLOX3 gene and also 12 OsLOX cDNA clones from Oryza sativa indica (cultivar Pusa Basmati 1). We have examined the role played by LOXs in plant development and during attack by blast pathogen Magnaporthe grisea. Gene expression, promoter region analysis, and biochemical and protein structure analysis of isolated OsLOX3 revealed significant homology with LOX super family. Protein sequence comparison of OsLOXs revealed high levels of homology when compared with japonica rice (up to100%) and Arabidopsis (up to 64%). Isolated LOX3 gene and 12 OsLOX cDNAs contained the catalytic LOX domains much required for oxygen binding and synthesis of oxylipins. Amino acid composition, protein secondary structure, and promoter region analysis (with abundance of motifs CGTCA and TGACG) support the role of OsLOX3 gene in providing resistance to diseases in rice plants. OsLOX3 gene expression analysis of root, shoot, flag leaf, and developing and mature seed revealed organ specific patterns during rice plant development and gave evidence to association between tissue location and physiological roles played by individual OsLOXs. Increased defense activity of oxylipins was observed as demonstrated by PCR amplification of OsLOX3 gene and upon inoculation with virulent strains of M. grisea and ectopic application of methyl jasmonate in the injured leaf tissue in adult rice plants.     

Silencing genes encoding omega-3 fatty acid desaturase alters seed size and accumulation of Bean pod mottle virus in soybean

Omega-3 fatty acid desaturase (FAD3)-catalyzed conversion of linoleic acid to linolenic acid (18:3) is an important step for the biosynthesis of fatty acids as well as the phytohormone jasmonic acid (JA) in plants. We report that silencing three microsomal isoforms of GmFAD3 enhanced the accumulation of Bean pod mottle virus (BPMV) in soybean. The GmFAD3-silenced plants also accumulated higher levels of JA, even though they contained slightly reduced levels of 18:3. Consequently, the GmFAD3-silenced plants expressed JA-responsive pathogenesis-related genes constitutively and exhibited enhanced susceptibility to virulent Pseudomonas syringae. Increased accumulation of BPMV in GmFAD3-silenced plants was likely associated with their JA levels, because exogenous JA application also increased BPMV accumulation. The JA-derived increase in BPMV levels was likely not due to repression of salicylic acid (SA)-derived signaling because the GmFAD3-silenced plants were enhanced in SA-dependent defenses. Furthermore, neither exogenous SA application nor silencing the SA-synthesizing phenylalanine ammonia lyase gene altered BPMV levels in soybean. In addition to the altered defense responses, the GmFAD3-silenced plants also produced significantly larger and heavier seed. Our results indicate that loss of GmFAD3 enhances JA accumulation and, thereby, susceptibility to BPMV in soybean.     

Fusarium oxysporum hijacks COI1-mediated jasmonate signaling to promote disease development in Arabidopsis

Although defense responses mediated by the plant oxylipin jasmonic acid (JA) are often necessary for resistance against pathogens with necrotrophic lifestyles, in this report we demonstrate that jasmonate signaling mediated through COI1 in Arabidopsis thaliana is responsible for susceptibility to wilt disease caused by the root-infecting fungal pathogen Fusarium oxysporum . Despite compromised JA-dependent defense responses, the JA perception mutant coronatine insensitive 1 ( coi1 ), but not JA biosynthesis mutants, exhibited a high level of resistance to wilt disease caused by F. oxysporum . This response was independent from salicylic acid-dependent defenses, as coi1/NahG plants showed similar disease resistance to coi1 plants. Inoculation of reciprocal grafts made between coi1 and wild-type plants revealed that coi1 -mediated resistance occurred primarily through the coi1 rootstock tissues. Furthermore, microscopy and quantification of fungal DNA during infection indicated that coi1 -mediated resistance was not associated with reduced fungal penetration and colonization until a late stage of infection, when leaf necrosis was highly developed in wild-type plants. In contrast to wild-type leaves, coi1 leaves showed no necrosis following the application of F. oxysporum culture filtrate, and showed reduced expression of senescence-associated genes during disease development, suggesting that coi1 resistance is most likely achieved through the inhibition of F. oxysporum -incited lesion development and plant senescence. Together, our results indicate that F. oxysporum hijacks non-defensive aspects of the JA-signaling pathway to cause wilt-disease symptoms that lead to plant death in Arabidopsis.     

The Nicotiana attenuata GLA1 lipase controls the accumulation of Phytophthora parasitica‐induced oxylipins and defensive secondary metabolites

Nicotiana attenuata plants silenced in the expression of GLYCEROLIPASE A1 (ir‐gla1 plants) are compromised in the herbivore‐ and wound‐induced accumulation of jasmonic acid (JA). However, these plants accumulate wild‐type (WT) levels of JA and divinyl‐ethers during Phytophthora parasitica infection. By profiling oxylipin‐enriched fractions with targeted and untargeted liquid chromatography‐tandem time‐of‐flight mass spectrometry approaches, we demonstrate that the accumulation of 9‐hydroxy‐10E,12Z‐octadecadienoic acid (9‐OH‐18:2) and additional C₁₈ and C₁₉ oxylipins is reduced by ca. 20‐fold in P. parasitica‐infected ir‐gla1 leaves compared with WT. This reduced accumulation of oxylipins was accompanied by a reduced accumulation of unsaturated free fatty acids and specific lysolipid species. Untargeted metabolic profiling of total leaf extracts showed that 87 metabolites accumulated differentially in leaves of P. parasitica‐infected ir‐gla1 plants with glycerolipids, hydroxylated‐diterpene glycosides and phenylpropanoid derivatives accounting together for ca. 20% of these 87 metabolites. Thus, P. parasitica‐induced oxylipins may participate in the regulation of metabolic changes during infection. Together, the results demonstrate that GLA1 plays a distinct role in the production of oxylipins during biotic stress responses, supplying substrates for 9‐OH‐18:2 and additional C₁₈ and C₁₉ oxylipin formation during P. parasitica infection, whereas supplying substrates for the biogenesis of JA during herbivory and mechanical wounding.     

Fatty acid unsaturation,mobilization, and regulation in the response of plants to stress

Stress acclimating plants respond to abiotic and biotic stress by remodeling membrane fluidity and by releasing α-linolenic (18:3) from membrane lipids. The modification of membrane fluidity is mediated by changes in unsaturated fatty acid levels, a function provided in part by the regulated activity of fatty acid desaturases. Adjustment of membrane fluidity maintains an environment suitable for the function of critical integral proteins during stress. α-Linolenic acid, released from membrane lipid by regulated lipase activity, is the precursor molecule for phyto-oxylipin biosynthesis. The modulation of chloroplast oleic acid (18:1) levels is central to the normal expression of defense responses to pathogens in Arabidopsis. Oleic (18:1) and linolenic (18:2) acid levels, in part, regulate development, seed colonization, and mycotoxin production by Aspergillus spp.     

Low oleic acid-derived repression of jasmonic acid-inducible defense responses requires the WRKY50 and WRKY51 proteins

Signaling induced upon a reduction in oleic acid (18:1) levels simultaneously up-regulates salicylic acid (SA)-mediated responses and inhibits jasmonic acid (JA)-inducible defenses, resulting in enhanced resistance to biotrophs but increased susceptibility to necrotrophs. SA and the signaling component Enhanced Disease Susceptibility1 function redundantly in this low-18:1-derived pathway to induce SA signaling but do not function in the repression of JA responses. We show that repression of JA-mediated signaling under low-18:1 conditions is mediated via the WRKY50 and WRKY51 proteins. Knockout mutations in WRKY50 and WRKY51 lowered SA levels but did not restore pathogenesis-related gene expression or pathogen resistance to basal levels in the low-18:1-containing Arabidopsis (Arabidopsis thaliana) mutant, suppressor of SA insensitivity2 (ssi2). In contrast, both JA-inducible PDF1.2 (defensin) expression and basal resistance to Botrytis cinerea were restored. Simultaneous mutations in both WRKY genes (ssi2 wrky50 wrky51) did not further enhance the JA or Botrytis-related responses. The ssi2 wrky50 and ssi2 wrky51 plants contained high levels of reactive oxygen species and exhibited enhanced cell death, the same as ssi2 plants. This suggested that high reactive oxygen species levels or increased cell death were not responsible for the enhanced susceptibility of ssi2 plants to B. cinerea. Exogenous SA inhibited JA-inducible PDF1.2 expression in the wild type but not in wrky50 or wrky51 mutant plants. These results show that the WRKY50 and WRKY51 proteins mediate both SA- and low-18:1-dependent repression of JA signaling.     

The tomato suppressor of prosystemin-mediated responses2 gene encodes a fatty acid desaturase required for the biosynthesis of jasmonic acid and the production of a systemic wound signal for defense gene expression

Genetic analysis of the wound response pathway in tomato indicates that systemin and its precursor protein, prosystemin, are upstream components of a defensive signaling cascade that involves the synthesis and subsequent action of the octadecatrienoic acid (18:3)-derived plant hormone jasmonic acid (JA). The suppressor of prosystemin-mediated responses2 (spr2) mutation, which was isolated previously as a suppressor of (pro)systemin-mediated signaling, impairs wound-induced JA biosynthesis and the production of a long-distance signal for the expression of defensive Proteinase inhibitor genes. Using a map-based cloning approach, we demonstrate here that Spr2 encodes a chloroplast fatty acid desaturase involved in JA biosynthesis. Loss of Spr2 function reduced the 18:3 content of leaves to <10% of wild-type levels, abolished the accumulation of hexadecatrienoic acid, and caused a corresponding increase in the level of dienoic fatty acids. The effect of spr2 on the fatty acyl content of various classes of glycerolipids indicated that the Spr2 gene product catalyzes most, if not all, omega3 fatty acid desaturation within the "prokaryotic pathway" for lipid synthesis in tomato leaves. Despite the reduced levels of trienoic fatty acids, spr2 plants exhibited normal growth, development, and reproduction. However, the mutant was compromised in defense against attack by tobacco hornworm larvae. These results indicate that jasmonate synthesis from chloroplast pools of 18:3 is required for wound- and systemin-induced defense responses and support a role for systemin in the production of a transmissible signal that is derived from the octadecanoid pathway.     

Probenazole-induced accumulation of salicylic acid confers resistance to Magnaporthe grisea in adult rice plants

Probenazole (PBZ) is the active ingredient of Oryzemate, an agrochemical which is used for the protection of rice plants from Magnaporthe grisea (blast fungus). While PBZ was reported to function upstream of salicylic acid (SA) in Arabidopsis, little is known about the mechanism of PBZ-induced resistance in rice. The role of SA in blast fungus resistance is also unclear. The recommended application period for Oryzemate is just before the Japanese rainy season, at which time rice plants in the field have reached the 8-leaf stage with adult traits. Thus, the involvement of SA in PBZ-induced resistance was studied in compatible and incompatible blast fungus-rice interactions at two developmentally different leaf morphology stages. Pre-treatment of inoculated fourth leaves of young wild-type rice plants at the 4-leaf stage with PBZ did not influence the development of whitish expanding lesions (ELs) in the susceptible interaction without the accumulation of SA and pathogenesis-related (PR) proteins. However, PBZ pre-treatment increased accumulation of SA and PR proteins in the eighth leaves of adult plants at the 8-leaf stage, resulting in the formation of hypersensitive reaction (HR) lesions (HRLs). Exogenous SA induced resistance in adult but not young plants. SA concentrations in blast fungus-inoculated young leaves were essentially the same in compatible and incompatible interactions, suggesting that PBZ-induced resistance in rice is age-dependently regulated via SA accumulation.     

Rice OsPAD4 functions differently from Arabidopsis AtPAD4 in host‐pathogen interactions

The extensively studied Arabidopsis phytoalexin deficient 4 (AtPAD4) gene plays an important role in Arabidopsis disease resistance; however, the function of its sequence ortholog in rice is unknown. Here, we show that rice OsPAD4 appears not to be the functional ortholog of AtPAD4 in host‐pathogen interactions, and that the OsPAD4 encodes a plasma membrane protein but that AtPAD4 encodes a cytoplasmic and nuclear protein. Suppression of OsPAD4 by RNA interference (RNAi) increased rice susceptibility to the biotrophic pathogen Xanthomonas oryzae pv. oryzae (Xoo), which causes bacteria blight disease in local tissue. OsPAD4‐RNAi plants also show compromised wound‐induced systemic resistance to Xoo. The increased susceptibility to Xoo was associated with reduced accumulation of jasmonic acid (JA) and phytoalexin momilactone A (MOA). Exogenous application of JA complemented the phenotype of OsPAD4‐RNAi plants in response to Xoo. The following results suggest that OsPAD4 functions differently than AtPAD4 in response to pathogen infection. First, OsPAD4 plays an important role in wound‐induced systemic resistance, whereas AtPAD4 mediates systemic acquired resistance. Second, OsPAD4‐involved defense signaling against Xoo is JA‐dependent, but AtPAD4‐involved defense signaling against biotrophic pathogens is salicylic acid‐dependent. Finally, OsPAD4 is required for the accumulation of terpenoid‐type phytoalexin MOA in rice‐bacterium interactions, but AtPAD4‐mediated resistance is associated with the accumulation of indole‐type phytoalexin camalexin.     

Analysis of the subcellular localisation of lipoxygenase in legume and actinorhizal nodules

Plant lipoxygenases (LOXs; EC 1.13.11.12) catalyse the oxygenation of polyunsaturated fatty acids, linoleic (18:2) and α-linolenic acid (18:3(n-3)) and are involved in processes such as stress responses and development. Depending on the regio-specificity of a LOX, the incorporation of molecular oxygen leads to formation of 9- or 13-fatty acid hydroperoxides, which are used by LOX itself as well as by members of at least six different enzyme families to form a series of biologically active molecules, collectively called oxylipins. The best characterised oxylipins are the jasmonates: jasmonic acid (JA) and its isoleucine conjugate that are signalling compounds in vegetative and propagative plant development. In several types of nitrogen-fixing root nodules, LOX expression and/or activity is induced during nodule development. Allene oxide cyclase (AOC), a committed enzyme of the JA biosynthetic pathway, has been shown to localise to plastids of nodules of one legume and two actinorhizal plants, Medicago truncatula, Datisca glomerata and Casuarina glauca, respectively. Using an antibody that recognises several types of LOX interspecifically, LOX protein levels were compared in roots and nodules of these plants, showing no significant differences and no obvious nodule-specific isoforms. A comparison of the cell-specific localisation of LOXs and AOC led to the conclusion that (i) only cytosolic LOXs were detected although it is generally assumed that the (13S)-hydroperoxy α-linolenic acid for JA biosynthesis is produced in the plastids, and (ii) in cells of the nodule vascular tissue that contain AOC, no LOX protein could be detected.     

Jasmonic acid carboxyl methyltransferase regulates development and herbivory‐induced defense response in rice

Jasmonic acid(JA) and related metabolites play a key role in plant defense and growth. JA carboxyl methyltransferase(JMT) may be involved in plant defense and development by methylating JA to methyl jasmonate(Me JA) and thus influencing the concentrations of JA and related metabolites. However, no JMT gene has been well characterized in monocotyledon defense and development at the molecular level. After we cloned a rice JMT gene,Os JMT1, whose encoding protein was localized in the cytosol, we found that the recombinant Os JMT1 protein catalyzed JA to Me JA. Os JMT1 is up-regulated in response to infestation with the brown planthopper(BPH; Nilaparvata lugens). Plants in which Os JMT1 had been overexpressed(oeJMT plants) showed reduced height and yield. These oe-JMT plants also exhibited increased Me JA levels but reduced levels of herbivore-induced JA and jasmonoyl-isoleucine(JAIle). The oe-JMT plants were more attractive to BPH female adults but showed increased resistance to BPH nymphs,probably owing to the different responses of BPH female adults and nymphs to the changes in levels of H_2O_2 and Me JA in oe-JMT plants. These results indicate that Os JMT1,by altering levels of JA and related metabolites, plays a role in regulating plant development and herbivore-induced defense responses in rice.