Regulation of jasmonic acid biosynthesis by silicon application during physical injury to Oryza sativa L.

We investigated the effects of silicon (Si) application on rice plants (Oryza sativa L.) and its responses in the regulation of jasmonic acid (JA) during wounding stress. Endogenous JA was significantly higher in wounded rice plants than in non-wounded. In contrast, Si treatment significantly reduced JA synthesis as compared to non-Si applications under wounding stress. mRNA expression of O. sativa genes showed down-regulation of lipoxygenase, allene oxide synthase 1, allene oxide synthase 2, 12-oxophytodienoate reductase 3, and allene oxide cyclase upon Si application and wounding stress as compared to non-Si-treated wounded rice plants. The physical injury-induced-oxidative stress was modulated by Si treatments, which resulted in higher catalase, peroxidase, and polyphenol oxidase activities as compared with non-Si-treated plants under wounding stress. The higher Si accumulation in rice plants also reduced the level of lipid peroxidation, which helped the rice plants to protect it from wounding stress. In conclusion, Si accumulation in rice plants mitigated the adverse effects of wounding through regulation of antioxidants and JA.     

Identification of the OsOPR7 gene encoding 12-oxophytodienoate reductase involved in the biosynthesis of jasmonic acid in rice

Enzyme 12-oxophytodienoate (OPDA) reductase (EC1.3.1.42), which is involved in the biosynthesis of jasmonic acid (JA), catalyses the reduction of 10, 11-double bonds of OPDA to yield 3-oxo-2-(2′-pentenyl)-cyclopentane-1-octanoic acid (OPC-8:0). The rice OsOPR1 gene encodes OPDA reductase (OPR) converting (−)-cis-OPDA preferentially, rather than (+)-cis-OPDA, a natural precursor of JA. Here, we provide evidence that an OPR family gene in rice chromosome 8, designated OsOPR7, encodes the enzyme involved in the JA biosynthesis. Recombinant OsOPR7-His protein efficiently catalysed the reduction of both enantiomers of cis-OPDA, similar to the OPR3 protein in Arabidopsis thaliana (L.) Heynh. The expression of OsOPR7 mRNA was induced and reached maximum levels within 0.5 h of mechanical wounding and drought stress, and the endogenous JA level started to increase in accordance with the increase in OsOPR7 expression. The GFP-OsOPR7 fusion protein was detected exclusively in peroxisomes in onion epidermal cells. Furthermore, complementation analysis using an Arabidopsis opr3 mutant indicated that the OsOPR7 gene, but not OsOPR1, was able to complement the phenotypes of male sterility in the mutant caused by JA deficiency, and that JA production in the opr3 mutant was also restored by the expression of the OsOPR7 gene. We conclude that the OsOPR7 gene encodes the enzyme catalysing the reduction of natural (+)-cis-OPDA for the JA biosynthesis in rice. Tomoyuki Tani and Hiroyuki Sobajima have equally contributed to this work.     

OsNPR1 negatively regulates herbivore‐induced JA and ethylene signaling and plant resistance to a chewing herbivore in rice

NPR1 (a non‐expressor of pathogenesis‐related genes1) has been reported to play an important role in plant defense by regulating signaling pathways. However, little to nothing is known about its function in herbivore‐induced defense in monocot plants. Here, using suppressive substrate hybridization, we identified a NPR1 gene from rice, OsNPR1, and found that its expression levels were upregulated in response to infestation by the rice striped stem borer (SSB) Chilo suppressalis and rice leaf folder (LF) Cnaphalocrocis medinalis, and to mechanical wounding and treatment with jasmonic acid (JA) and salicylic acid (SA). Moreover, mechanical wounding induced the expression of OsNPR1 quickly, whereas herbivore infestation induced the gene more slowly. The antisense expression of OsNPR1 (as‐npr1), which reduced the expression of the gene by 50%, increased elicited levels of JA and ethylene (ET) as well as of expression of a lipoxygenase gene OsHI‐LOX and an ACC synthase gene OsACS2. The enhanced JA and ET signaling in as‐npr1 plants increased the levels of herbivore‐induced trypsin proteinase inhibitors (TrypPIs) and volatiles, and reduced the performance of SSB. Our results suggest that OsNPR1 is an early responding gene in herbivore‐induced defense and that plants can use it to activate a specific and appropriate defense response against invaders by modulating signaling pathways.     

Tobacco Salicylic Acid Glucosyltransferase Is Active toward Tuberonic Acid (12-Hydroxyjasmonic Acid) and Is Induced by Mechanical Wounding Stress

Recently we reported that rice salicylic acid (SA) glucosyltransferase (OsSGT) is active toward 12-hydroxyjasmonic acid (tuberonic acid, TA) and that OsSGT gene expression is induced by wounding stress. Here we report that tobacco SA glucosyltransferase (NtSGT), which is thought to be an ortholog of OsSGT, is also active toward TA. Although NtSGT expression is known to be induced by biotrophic stress, it was also induced by wounding stress in the same manner as OsSGT. These results indicate that this glucosyltransferase is important not only in biotrophic stress but also for wounding stress. It was found that this enzyme is dually functional, with activity both toward TA and SA.     

OsJAR1 and OsJAR2 are jasmonyl-l-isoleucine synthases involved in wound- and pathogen-induced jasmonic acid signalling

The synthesis of JA-Ile was catalysed by JA-Ile synthase, which is a member of the group I GH3 family of proteins. Here, we showed evidence that OsGH3.5 (OsJAR1) and OsGH3.3 (OsJAR2) are the functional JA-Ile synthases in rice, using recombinant proteins. The expression levels of OsJAR1 and OsJAR2 were induced in response to wounding with the concomitant accumulation of JA-Ile. In contrast, only the expression of OsJAR1 was associated with the accumulation of JA-Ile after blast infection. Our data suggest that these two JA-Ile synthases are differentially involved in the activation of JA signalling in response to wounding and pathogen challenge in rice.     

Light induces jasmonate‐isoleucine conjugation via OsJAR1‐dependent and ‐independent pathways in rice

The bioactive form of jasmonate is the conjugate of the amino acid isoleucine (Ile) with jasmonic acid (JA), which is biosynthesized in a reaction catalysed by the GH3 enzyme JASMONATE RESISTANT 1 (JAR1). We examined the biochemical properties of OsJAR1 and its involvement in photomorphogenesis of rice (Oryza sativa). OsJAR1 has a similar substrate specificities as its orthologue in Arabidopsis. However, osjar1 loss‐of‐function mutants did not show as severe coleoptile phenotypes as the JA‐deficient mutants coleoptile photomorphogenesis 2 (cpm2) and hebiba, which develop long coleoptiles in all light qualities we examined. Analysis of hormonal contents in the young seedling stage revealed that osjar1 mutants are still able to synthesize JA‐Ile conjugate in response to blue light, suggesting that a redundantly active enzyme can conjugate JA and Ile in rice seedlings. A good candidate for this enzyme is OsJAR2, which was found to be able to catalyse the conjugation of JA with Ile as well as with some additional amino acids. In contrast, if plants in the vegetative stage were mechanically wounded, the content of JA‐Ile was severely reduced in osjar1, demonstrating that OsJAR1 is the most important JA‐Ile conjugating enzyme in the wounding response during the vegetative stage.     

Kinetics of the Accumulation of Jasmonic Acid and Its Derivatives in Systemic Leaves of Tobacco (Nicotiana tabacum cv. Xanthi nc) and Translocation of Deuterium-Labeled Jasmonic Acid from the Wounding Site to the Systemic Site

In plants, the mobile signal needed for wound-induced systemic acquired resistance (WSR) has been elusive. The signal compound involved in WSR is supposed to be JA or its derivatives. On the basis of kinetic study of the accumulation of JA or its derivatives, it was discovered that JA, JA-Ile, tuberonic acid (TA, 12-OH epi-JA), and tuberonic acid glucoside (TAG) accumulated in systemic tissues in response to mechanical wounding stress in the tobacco plant (Nicotiana tabacum). Attempts to recover deuterium-labeled JA in systemic leaves after feeding the wounded leaves with deuterium-labeled JA were successfully done. It was also found that the translocated deuterium-labeled JA was metabolized to TA in systemic leaves under feeding of deuterium-labeled JA to the wounding leaves.     

Jasmonoyl isoleucine accumulation is needed for abscisic acid build‐up in roots of Arabidopsis under water stress conditions

Phytohormones are central players in sensing and signalling numerous environmental conditions like drought. In this work, hormone profiling together with gene expression of key enzymes involved in abscisic acid (ABA) and jasmonate biosynthesis were studied in desiccating Arabidopsis roots. Jasmonic acid (JA) content transiently increased after stress imposition whereas progressive and concomitant ABA and Jasmonoyl Isoleucine (JA‐Ile) accumulations were detected. Molecular data suggest that, at least, part of the hormonal regulation takes place at the biosynthetic level. These observations also point to a possible involvement of jasmonates on ABA biosynthesis under stress. To test this hypothesis, mutants impaired in jasmonate biosynthesis (opr3, lox6 and jar1‐1) and in JA‐dependent signalling (coi1) were employed. Results showed that the early JA accumulation leading to JA‐Ile build up was necessary for an ABA increase in roots under two different water stress conditions. Signal transduction between water stress‐induced JA‐Ile accumulation and COI1 is necessary for a full induction of the ABA biosynthesis pathway and subsequent hormone accumulation in roots of Arabidopsis plants. The present work adds a level of interaction between jasmonates and ABA at the biosynthetic level.     

Overexpression of <Emphasis Type="Italic">OsiSAP8</Emphasis>, a member of stress associated protein (SAP) gene family of rice confers tolerance to salt,drought and cold stress in transgenic tobacco and rice

We describe here the isolation and characterization of OsiSAP8, a member of stress Associated protein (SAP) gene family from rice characterized by the presence of A20 and AN1 type Zinc finger domains. OsiSAP8 is a multiple stress inducible gene, induced by various stresses, namely heat, cold, salt, desiccation, submergence, wounding, heavy metals as well as stress hormone Abscisic acid. OsiSAP8 protein fused to GFP was localized towards the periphery of the cells in the epidermal cells of infiltrated Nicotiana benthamiana leaves. Yeast two hybrid analysis revealed that A20 and AN1 type zinc-finger domains of OsiSAP8 interact with each other. Overexpression of the gene in both transgenic tobacco and rice conferred tolerance to salt, drought and cold stress at seed germination/seedling stage as reflected by percentage of germination and gain in fresh weight after stress recovery. Transgenic rice plants were tolerant to salt and drought during anthesis stage without any yield penalty as compared to unstressed transgenic plants. OsiSAP8 is deposited in the Genbank with the Accession number AY345599.