JA-isoleucine Research Articles

Physiological and metabolomic analyses reveal the resistance response mechanism to tea aphid infestation in new shoots of tea plants (Camellia sinensis)

The tea plant (Camellia sinensis (L.) O. Kuntze) is an important woody economic crop and often attacked by various insect pests such as tea aphids (Toxoptera aurantii Boyer de Fonscolombe). To reveal tea plant responses to tea aphid infestation, tea aphid-resistant cultivar ‘Taicha13’ and -susceptible one ‘Jingfeng’ were first screened, and then widely targeted metabolomics profiling, biochemical parameter and RT-qPCR analyses were performed in three samples, TCN (tea aphid non-infested ‘Taicha13’), JFN (tea aphid non-infested ‘Jingfeng’) and JFA (tea aphid-infested ‘Jingfeng’). The results showed amnio acid (plus proline) and protein content, and amnio acid: sucrose ratio were significantly less abundant in TCN relative to JFN, suggesting TCN is low nutritional quality for tea aphids. Metabolite profiling revealed that flavonoids and JA-isoleucine (JA-Ile) were more enriched in TCN and JFA compared to JFN, proposing effective roles of flavonoids and JA-Ile in tea plant defense against tea aphid infestation. RT-qPCR results demonstrated that five JA biosynthesis genes (CsLOX2, CsLOX3, CsOPR, CsAOC and CsJAR) in exception of CsAOS, and six flavonoid biosynthesis genes (CsPAL, CsC4H, CsCHS1, CsCHS2, CsFL and CsDFR) shared similarly higher abundance in TCN than in JFN, and exhibited elevated expression levels in JFA relative to JFN, further suggesting flavonoids and JA confer tea plant resistance against tea aphid feeding. Overall, our findings uncovered that tea aphid-infestation activated defensive responses and highlighted involvement of nutritional quality, flavonoids and JA-Ile in tea plant resistance against tea aphid infestation.

Silencing an ATP-Dependent Caseinolytic Protease Proteolytic Subunit Gene Enhances the Resistance of Rice to Nilaparvata lugens.

The ATP-dependent caseinolytic protease (Clp) system has been reported to play an important role in plant growth, development, and defense against pathogens. However, whether the Clp system is involved in plant defense against herbivores remains largely unclear. We explore the role of the Clp system in rice defenses against brown planthopper (BPH) Nilaparvata lugens by combining chemical analysis, transcriptome, and molecular analyses, as well as insect bioassays. We found the expression of a rice Clp proteolytic subunit gene, OsClpP6, was suppressed by infestation of BPH gravid females and mechanical wounding. Silencing OsClpP6 enhanced the level of BPH-induced jasmonic acid (JA), JA-isoleucine (JA-Ile), and ABA, which in turn promoted the production of BPH-elicited rice volatiles and increased the resistance of rice to BPH. Field trials showed that silencing OsClpP6 decreased the population densities of BPH and WBPH. We also observed that silencing OsClpP6 decreased chlorophyll content in rice leaves at early developmental stages and impaired rice root growth and seed setting rate. These findings demonstrate that an OsClpP6-mediated Clp system in rice was involved in plant growth-defense trade-offs by affecting the biosynthesis of defense-related signaling molecules in chloroplasts. Moreover, rice plants, after recognizing BPH infestation, can enhance rice resistance to BPH by decreasing the Clp system activity. The work might provide a new way to breed rice varieties that are resistant to herbivores.

Halomonas ventosaeJPT10 promotes salt tolerance in foxtail millet (Setaria italica) by affecting the levels of multiple antioxidants and phytohormones.

Plant growth-promoting bacterias (PGPBs) can increase crop output under normal and abiotic conditions. However, the mechanisms underlying the plant salt tolerance-promoting role of PGPBs still remain largely unknown. In this study, we demonstrated that Halomonas ventosae JPT10 promoted the salt tolerance of both dicots and monocots. Physiological analysis revealed that JPT10 reduced reactive oxygen species accumulation by improving the antioxidant capability of foxtail millet seedlings. The metabolomic analysis of JPT10-inoculated foxtail millet seedlings led to the identification of 438 diversely accumulated metabolites, including flavonoids, phenolic acids, lignans, coumarins, sugar, alkaloids, organic acids, and lipids, under salt stress. Exogenous apigenin and chlorogenic acid increased the salt tolerance of foxtail millet seedlings. Simultaneously, JPT10 led to greater amounts of abscisic acid (ABA), indole-3-acetic acid (IAA), salicylic acid (SA), and their derivatives but lower levels of 12-oxo-phytodienoic acid (OPDA), jasmonate (JA), and JA-isoleucine (JA-Ile) under salt stress. Exogenous JA, methyl-JA, and OPDA intensified, whereas ibuprofen or phenitone, two inhibitors of JA and OPDA biosynthesis, partially reversed, the growth inhibition of foxtail millet seedlings caused by salt stress. Our results shed light on the response of foxtail millet seedlings to H. ventosae under salt stress and provide potential compounds to increase salt tolerance in foxtail millet and other crops.

Sodium selenite increases drought tolerance by promoting jasmonic acid biosynthesis in cucumber

Jasmonic acid (JA) is an important and widely distributed plant hormone. However, the molecular and physiological mechanism of JA in improving drought tolerance in response to sodium selenite is limited. This work was performed to investigate the effects of exogenous sodium selenite application in promoting drought tolerance of cucumber. The drought tolerance of cucumber seedlings is enhanced under the application of selenite, positively influencing shoot fresh weight and chlorophyll relative content and altering the chloroplast ultrastructure. The contents of JA and JA-isoleucine (JA-ILE) increased significantly in response to selenite application under drought conditions. Furthermore, the expression of JA biosynthesis and regulatory genes, namely, LOX (Lipoxygenase), AOC (alleneoxide cyclase), AOS (alleneoxide synthase), and MYC2 (the basic helix-loop-helix (bHLH) protein) was upregulated to greater levels when selenite was added in combination with drought treatment. This study provides methods to mitigate drought stress and valuable theoretical support for further understanding the plant response to drought signals.

9,10-KODA, an α-ketol produced by the tonoplast-localized 9-lipoxygenase ZmLOX5, plays a signaling role in maize defense against insect herbivory

13-Lipoxygenases (LOXs) initiate the synthesis of jasmonic acid (JA), the best-understood oxylipin hormone in herbivory defense. However, the roles of 9-LOX-derived oxylipins in insect resistance remain unclear. Here, we report a novel anti-herbivory mechanism mediated by a tonoplast-localized 9-LOX, ZmLOX5, and its linolenic acid-derived product, 9-hydroxy-10-oxo-12(Z),15(Z)-octadecadienoic acid (9,10-KODA). Transposon-insertional disruption of ZmLOX5 resulted in the loss of resistance to insect herbivory. lox5 knockout mutants displayed greatly reduced wound-induced accumulation of multiple oxylipins and defense metabolites, including benzoxazinoids, abscisic acid (ABA), and JA-isoleucine (JA-Ile). However, exogenous JA-Ile failed to rescue insect defense in lox5 mutants, while applications of 1 μM 9,10-KODA or the JA precursor, 12-oxo-phytodienoic acid (12-OPDA), restored wild-type resistance levels. Metabolite profiling revealed that exogenous 9,10-KODA primed the plants for increased production of ABA and 12-OPDA, but not JA-Ile. While none of the 9-oxylipins were able to rescue JA-Ile induction, the lox5 mutant accumulated lower wound-induced levels of Ca2+, suggesting this as a potential explanation for lower wound-induced JA. Seedlings pretreated with 9,10-KODA exhibited rapid or more robust wound-induced defense gene expression. In addition, an artificial diet supplemented with 9,10-KODA arrested fall armyworm larvae growth. Finally, analysis of single and double lox5 and lox10 mutants showed that ZmLOX5 also contributed to insect defense by modulating ZmLOX10-mediated green leaf volatile signaling. Collectively, our study uncovered a previously unknown anti-herbivore defense and hormone-like signaling activity for a major 9-oxylipin α-ketol.

Comprehensive Phytohormone Profiling of Kohlrabi during In Vitro Growth and Regeneration: The Interplay with Cytokinin and Sucrose

The establishment of an efficient protocol for in vitro growth and regeneration of kohlrabi (Brassica oleracea var. gongylodes) allowed us to closely examine the phytohormone profiles of kohlrabi seedlings at four growth stages (T1–T4), additionally including the effects of cytokinins (CKs)—trans-zeatin (transZ) and thidiazuron (TDZ)—and high sucrose concentrations (6% and 9%). Resulting phytohormone profiles showed complex time-course patterns. At the T2 stage of control kohlrabi plantlets (with two emerged true leaves), levels of endogenous CK free bases and gibberellin GA20 increased, while increases in jasmonic acid (JA), JA-isoleucine (JA-Ile), indole-3-acetic acid (IAA) and indole-3-acetamide (IAM) peaked later, at T3. At the same time, the content of most of the analyzed IAA metabolites decreased. Supplementing growth media with CK induced de novo formation of shoots, while both CK and sucrose treatments caused important changes in most of the phytohormone groups at each developmental stage, compared to control. Principal component analysis (PCA) showed that sucrose treatment, especially at 9%, had a stronger effect on the content of endogenous hormones than CK treatments. Correlation analysis showed that the dynamic balance between the levels of certain bioactive phytohormone forms and some of their metabolites could be lost or reversed at particular growth stages and under certain CK or sucrose treatments, with correlation values changing between strongly positive and strongly negative. Our results indicate that the kohlrabi phytohormonome is a highly dynamic system that changes greatly along the developmental time scale and also during de novo shoot formation, depending on exogenous factors such as the presence of growth regulators and different sucrose concentrations in the growth media, and that it interacts intensively with these factors to facilitate certain responses.

The UDP-glycosyltransferase MtUGT84A1 regulates anthocyanin accumulation and plant growth via JA signaling in Medicago truncatula

Anthocyanins confer plants beneficial qualities and play protective roles under extreme light conditions. Here, we characterized a novel UDP flavonoid 3- O -glucosyltransferase ( UFGT ) gene MtUGT84A1 and demonstrated its involvement in anthocyanin accumulation and feedback inhibition of JA signaling in M. truncatula. The loss-of-function mutants mtugt84a1–1 and mtugt84a1–2 exhibited emerald hypocotyls and dwarfism. In vitro and in vivo assays revealed that MtUGT84A1 glycosylated peonidin and delphinidin at the 3-OH position. Constitutive expression of MtUGT84A1 rescued the phenotypic abnormalities of mtugt84a1 and significantly increased the anthocyanin content in the MtUGT84A1 -overexpression plants. Moreover, in mtugt84a1 , the contents of JA and JA isoleucine (JA-Ile) were doubled. The transcription level of MtMYC2 was significantly increased in mtugt84a1 , while decreased in the MtUGT84A1- overexpression plants, indicating MtUGT84A1 inhibits JA pathway. Under high-intensity light or MeJA treatment, MtUGT84A1 was elevated causing drastic accumulation of anthocyanins in the wildtype, but the pigments in mtugt84a1 did not alter clearly. Therefore, MtUGT84A1 not only functions as a structural gene for the final step of anthocyanin biosynthesis, but also regulates JA signaling by feedback inhibition. The findings unveil the critical roles of MtUGT84A1 -modulated glycosylation of anthocyanidins and shed light on the feedback regulatory loop of MtUGT84A1 in JA-mediated biosynthesis of natural products. • MtUGT84A1 encodes UDP flavonoid 3- O -glucosyltransferase. • MtUGT84A1 glucosylates peonidin and delphinidin at the 3-OH position. • MtUGT84A1 regulates JA signaling by feedback inhibition. • MtUGT84A1 modulates both anthocyanin accumulation and shoot growth of M. truncatula.

The HD-Zip transcription factor SlHB15A regulates abscission by modulating jasmonoyl-isoleucine biosynthesis.

Plant organ abscission, a process that is important for development and reproductive success, is inhibited by the phytohormone auxin and promoted by another phytohormone, jasmonic acid (JA). However, the molecular mechanisms underlying the antagonistic effects of auxin and JA in organ abscission are unknown. We identified a tomato (Solanum lycopersicum) class III homeodomain-leucine zipper transcription factor, HOMEOBOX15A (SlHB15A), which was highly expressed in the flower pedicel abscission zone and induced by auxin. Knocking out SlHB15A using clustered regularly interspaced short palindromic repeats-associated protein 9 technology significantly accelerated abscission. In contrast, overexpression of microRNA166-resistant SlHB15A (mSlHB15A) delayed abscission. RNA sequencing and reverse transcription-quantitative PCR analyses showed that knocking out SlHB15A altered the expression of genes related to JA biosynthesis and signaling. Furthermore, functional analysis indicated that SlHB15A regulates abscission by depressing JA-isoleucine (JA-Ile) levels through inhabiting the expression of JASMONATE-RESISTANT1 (SlJAR1), a gene involved in JA-Ile biosynthesis, which could induce abscission-dependent and abscission-independent ethylene signaling. SlHB15A bound directly to the SlJAR1 promoter to silence SlJAR1, thus delaying abscission. We also found that flower removal enhanced JA-Ile content and that application of JA-Ile severely impaired the inhibitory effects of auxin on abscission. These results indicated that SlHB15A mediates the antagonistic effect of auxin and JA-Ile during tomato pedicel abscission, while auxin inhibits abscission through the SlHB15A-SlJAR1 module.

SlWRKY45 interacts with jasmonate-ZIM domain proteins to negatively regulate defense against the root-knot nematode Meloidogyne incognita in tomato.

Parasitic root-knot nematodes (RKNs) cause a severe reduction in crop yield and seriously threaten agricultural production. The phytohormones jasmonates (JAs) are important signals regulating resistance to multiple biotic and abiotic stresses. However, the molecular mechanism for JAs-regulated defense against RKNs in tomato remains largely unclear. In this study, we found that the transcription factor SlWRKY45 interacted with most JA-ZIM domain family proteins (JAZs), key repressors of the JA signaling. After infection by the RKN Meloidogyne incognita, the slwrky45 mutants exhibited lower gall numbers and egg numbers per gram of roots than wild type, whereas overexpression of SlWRKY45 attenuated resistance to Meloidogyne incognita. Under M. incognita infection, the contents of jasmonic acid (JA) and JA-isoleucine (JA-Ile) in roots were repressed by SlWRKY45-overexpression. Furthermore, SlWRKY45 bound to and inhibited the promoter of the JA biosynthesis gene ALLENE OXIDE CYCLASE (AOC), and repressed its expression. Overall, our findings revealed that the SlJAZ-interaction protein SlWRKY45 attenuated RKN-regulated JA biosynthesis and repressed defense against the RKN M. incognita in tomato.

Impact of maize hormonal interactions on the performance of Spodoptera frugiperda in plants infected with Clavibacter michiganensis subsp. nebraskensis

In nature, plants interact with multiple organisms, below and aboveground. Although interactions of plants with single biotic stressors are well characterized, knowledge of how the immune system responds to multiple biotic stressors is lacking. It is known that the two most important pathways involved in the plant immune defenses are jasmonic acid (JA) and salicylic acid (SA). The crosstalk between these plant hormonal signaling pathways seems to fine-tune the plant responses to different stressors. In this study, we characterized maize (Zea mays) hormonal interactions under the attack of two economically important pests: the bacterial pathogen that causes Goss’s bacterial wilt and leaf blight (Clavibacter michiganensis subsp. nebraskensis), and the chewing insect, fall armyworm (Spodoptera frugiperda). Our results indicate that the Goss’s wilt-resistant maize plants when pretreated with bacteria became more vulnerable to the subsequent attack by the fall armyworm larvae. Furthermore, plant hormone analysis demonstrated that the antagonistic interactions of JA-Isoleucine (JA-Ile), biologically active form of JA, and SA in maize plants contributed to enhanced plant susceptibility to herbivory. Collectively, our results suggest that plant hormonal interactions may play a major role in maize defense against multiple biotic stressors. Furthermore, this study will improve our understanding and ability to predict plant-induced hormone responses in an economically important crop.

Ectopic expression of LhMYC2 increases susceptibility to Botrytis cinerea in Arabidopsis thaliana

Gray mold disease, mainly caused by Botrytis elliptica (Berk.) Cooke and Botrytis cinerea Pers., leads to severe losses in lily cut flower and bulb production. MYC2 is a critical regulator of the activation of jasmonate-mediated defense responses in plants; however, information about the lily MYC2 gene is limited. Therefore, functional characterization of MYC2 in lily, especially its role in plant immune responses, should be performed. Here, significant differences between the Botrytis-resistant Lilium hybrid ‘Sorbonne’ and the Botrytis-susceptible ‘Tresor’ were found following B. cinerea inoculation, as indicated by jasmonic acid (JA) and JA-isoleucine (JA-Ile) accumulation and related gene expression. More JA and JA-Ile were detected in ‘Sorbonne’ than in ‘Tresor’ following fungal inoculation, and higher transcript levels of JA biosynthesis genes (LhAOS, LhAOC, and LhOPR3) and a signaling gene (LhCOI1) were detected in ‘Sorbonne’ than in ‘Tresor’. In contrast, expression of the critical signaling regulator LhMYC2 was higher in ‘Tresor’ than in ‘Sorbonne’. LhMYC2 was then isolated from ‘Sorbonne’ and found to be similar to several plant MYC2 homologs that have pivotal roles in JA signaling. The expression of LhMYC2 increased significantly in response to JA and salicylic acid (SA) in ‘Sorbonne’. Ectopic expression of LhMYC2 in Arabidopsis resulted in greater susceptibility to B. cinerea than that observed in wild-type plants. This susceptibility was coupled with the transcriptional changes in SA- and JA-responsive genes. Overall, our findings indicate that LhMYC2 plays a negative role in Arabidopsis resistance to B. cinerea.

Jasmonic Acid-Isoleucine (JA-Ile) Is Involved in the Host-Plant Resistance Mechanism Against the Soybean Aphid (Hemiptera: Aphididae).

Host-plant resistance (HPR) is an important tool for pest management, affording both economic and environmental benefits. The mechanisms of aphid resistance in soybean are not well understood, but likely involve the induction of the jasmonic acid (JA) pathway, and possibly other phytohormone signals involved in plant defense responses. Despite the efficacy of aphid resistance in soybean, virulent aphids have overcome this resistance through mostly unknown mechanisms. Here, we have used metabolomic tools to define the role of plant phytohormones, especially the JA pathway, in regulating interactions between aphid-resistant soybean and virulent aphids. We hypothesized that virulent aphids avoid or suppress the JA pathway to overcome aphid resistance. Our results suggested that aphid-resistant soybean increased accumulation of JA-isoleucine (JA-Ile) only when infested with avirulent aphids; virulent aphids did not cause induction of JA-Ile. Further, applying JA-Ile to aphid-resistant soybean reduced subsequent virulent aphid populations. The concentrations of other phytohormones remained unchanged due to aphid feeding, highlighting the importance of JA-Ile in this interaction. These results increase our knowledge of soybean resistance mechanisms against soybean aphids and contribute to our understanding of aphid virulence mechanisms, which will in turn promote the durability of HPR.

Reshaping of the Arabidopsis thaliana Proteome Landscape and Co-regulation of Proteins in Development and Immunity

Proteome remodeling is a fundamental adaptive response, and proteins in complexes and functionally related proteins are often co-expressed. Using a deep sampling strategy we define core proteomes of Arabidopsis thaliana tissues with around 10 000 proteins per tissue, and absolutely quantify (copy numbers per cell) nearly 16 000 proteins throughout the plant lifecycle. A proteome-wide survey of global post-translational modification revealed amino acid exchanges pointing to potential conservation of translational infidelity in eukaryotes. Correlation analysis of protein abundance uncovered potentially new tissue- and age-specific roles of entire signaling modules regulating transcription in photosynthesis, seed development, and senescence and abscission. Among others, the data suggest a potential function of RD26 and other NAC transcription factors in seed development related to desiccation tolerance as well as a possible function of cysteine-rich receptor-like kinases (CRKs) as ROS sensors in senescence. All of the components of ribosome biogenesis factor (RBF) complexes were found to be co-expressed in a tissue- and age-specific manner, indicating functional promiscuity in the assembly of these less-studied protein complexes in Arabidopsis.Furthermore, we characterized detailed proteome remodeling in basal immunity by treating Arabidopsis seeldings with flg22. Through simultaneously monitoring phytohormone and transcript changes upon flg22 treatment, we obtained strong evidence of suppression of jasmonate (JA) and JA-isoleucine (JA-Ile) levels by deconjugation and hydroxylation by IAA-ALA RESISTANT3 (IAR3) and JASMONATE-INDUCED OXYGENASE 2 (JOX2), respectively, under the control of JASMONATE INSENSITIVE 1 (MYC2), suggesting an unrecognized role of a new JA regulatory switch in pattern-triggered immunity. Taken together, the datasets generated in this study present extensive coverage of the Arabidopsis proteome in various biological scenarios, providing a rich resource available to the whole plant science community.

Coronatine is more potent than jasmonates in regulating Arabidopsis circadian clock

Recent studies establish a crucial role of the circadian clock in regulating plant defense against pathogens. Whether pathogens modulate host circadian clock as a potential strategy to suppress host innate immunity is not well understood. Coronatine is a toxin produced by the bacterial pathogen Pseudomonas syringae that is known to counteract Arabidopsis defense through mimicking defense signaling molecules, jasmonates (JAs). We report here that COR preferentially suppresses expression of clock-related genes in high throughput gene expression studies, compared with the plant-derived JA molecule methyl jasmonate (MJ). COR treatment dampens the amplitude and lengthens the period of all four reporters tested while MJ and another JA agonist JA-isoleucine (JA-Ile) only affect some reporters. COR, MJ, and JA-Ile act through the canonical JA receptor COI1 in clock regulation. These data support a stronger role of the pathogen-derived molecule COR than plant-derived JA molecules in regulating Arabidopsis clock. Further study shall reveal mechanisms underlying COR regulation of host circadian clock.

Jasmonates: biosynthesis, perception and signal transduction.

Jasmonates (JAs) are physiologically important molecules involved in a wide range of plant responses from growth, flowering, senescence to defence against abiotic and biotic stress. They are rapidly synthesised from α-linolenic acid (ALA; C18:3 ∆9,12,15) by a process of oxidation, cyclisation and acyl chain shortening involving co-operation between the chloroplast and peroxisome. The active form of JA is the isoleucine conjugate, JA-isoleucine (JA-Ile), which is synthesised in the cytoplasm. Other active metabolites of JA include the airborne signalling molecules, methyl JA (Me-JA) and cis-jasmone (CJ), which act as inter-plant signalling molecules activating defensive genes encoding proteins and secondary compounds such as anthocyanins and alkaloids. One of the key defensive metabolites in many plants is a protease inhibitor that inactivates the protein digestive capabilities of insects, thereby, reducing their growth. The receptor for JA-Ile is a ubiquitin ligase termed as SCFCoi1 that targets the repressor protein JA Zim domain (JAZ) for degradation in the 26S proteasome. Removal of JAZ allows other transcription factors (TFs) to activate the JA response. The levels of JA-Ile are controlled through catabolism by hydroxylating enzymes of the cytochrome P450 (CYP) family. The JAZ proteins act as metabolic hubs and play key roles in cross-talk with other phytohormone signalling pathways in co-ordinating genome-wide responses. Specific subsets of JAZ proteins are involved in regulating different response outcomes such as growth inhibition versus biotic stress responses. Understanding the molecular circuits that control plant responses to pests and pathogens is a necessary pre-requisite to engineering plants with enhanced resilience to biotic challenges for improved agricultural yields.

Jasmonate is required for the response to osmotic stress in rice

Plants have the ability to alleviate the harmful effects caused by abiotic and biotic stress. Phytohormones play a very important role in the acclimation to these stresses. To study the role of jasmonate in the acclimation to osmotic stress, an ALLENE OXIDE CYCLASE (AOC) mutant of rice (cpm2), disrupted in the biosynthesis of jasmonic acid (JA), and its wild type (WT) background were employed to investigate their responses to osmotic stress caused by treatment with polyethylene glycol (PEG) 6000. WT showed tolerance to osmotic stress, correlated with a fast transient increase of JA and JA-isoleucine (JA-Ile) in the shoots prior to an increase in abscisic acid (ABA), followed by a second increase in jasmonates when exposing to osmotic stress during 24 h. In roots, the pattern of hormonal increase was similar, but the response appeared to be faster, and remained transient, also with respect to low levels of jasmonates upon continuing osmotic stress. The mutant, which was containing extremely low levels of jasmonates, was hypersensitive to the stress. However, ABA accumulated in both, shoots and roots of cpm2, to similar (but not equal) levels as those seen in the WT, demonstrating that the biosynthesis or catabolism of ABA in response to osmotic stress is at least partially independent of JA, but can be modulated by JA. Our results suggest that jasmonates operate in parallel, presumably synergistically, to ABA, and are indispensable for osmotic stress tolerance in rice.

Relative contribution of LOX10, green leaf volatiles and JA to wound-induced local and systemic oxylipin and hormone signature in Zea mays (maize)

Green leaf volatiles (GLVs) and jasmonates (JAs) are the best-characterized groups of fatty acid-derived oxylipin signals that regulate wound-associated defenses. Beyond these two major groups of defense signals, plants produce an array of oxylipins in response to wounding, which possess potent signaling and/or insecticidal activities. In this study, we assessed the relative contribution of JAs and GLVs to wound-induced systemic signaling and the associated regulation of oxylipins in local and systemic tissues of maize (Zea mays). For this, we utilized GLV- and JA-deficient mutants, lox10 single and opr7opr8 double mutants, respectively, and profiled oxylipins in untreated leaves and roots, and in locally wounded and systemic leaves. In contrast to the studies in dicots, no systemic induction of JAs was observed in maize. Instead, a JA precursor, 12-OPDA, as well as ketols and C12/13 oxo-acids derived from 13-lipoxygenases (LOXs), were preferentially induced in both locally wounded and systemic unwounded leaves. Several 9-LOX-derived oxylipins (9-oxylipins) including hydroxides and ketones were also significantly induced locally. JA and JA-isoleucine (JA-Ile) were rapidly induced within 0.5 h, and were followed by a second increase in local tissue 4 h after wounding. GLV-deficient lox10 mutants displayed reduced levels of most 13-oxylipins, and elevated levels of several 9-oxylipins and the a-dioxygenase (DOX) product, 2-HOD. lox10 mutants were completely devoid of C6 volatiles and their C12 counterparts, and greatly decreased in C5 volatiles and their C13 oxo-acid counterparts. Thus, in addition to being the sole LOX isoform providing substrate for GLV synthesis, LOX10 is a major 13-LOX that provides substrate to several LOX branches that produce an array of 13-oxylipin products, including C5 volatiles. Interestingly, the rapid JA and JA-Ile increase at 0.5–2 h post-wounding was only moderately affected by the LOX10 mutation, while significantly reduced levels were observed at 4 h post-wounding. Combined with the previous findings that GLVs activate JA biosynthesis, these results suggest that both LOX10-derived substrates and/or GLVs are involved in the large second phase of JA synthesis proximal to the wound. Analyses of opr7opr8 mutants revealed that wound-induced oxylipin responses were positively regulated by JA signaling. The local and systemic accumulation of SA was not altered in the two mutants. Collectively, our results identified a subset of oxylipins strongly induced in wounded and systemic leaves, but their impact on insect defenses remain elusive. The lack of systemic induction of JAs points to substantial difference between systemic wound responses in studied dicots and maize. Our results show that GLV-deficiency and reduced JA in lox10 mutants had a greater impact on wound-induced local and systemic tissue oxylipin responses compared to the solely JA-deficient opr7opr8 double mutants. This suggests that GLVs or other LOX10-derived products heavily contribute to overall basal and wound-induced oxylipin responses. The specific roles of the GLV- and/or JA-dependent oxylipins in wound responses and defense remain to be further investigated by a combination of multiple orders of oxylipin-deficient mutants.

Accumulation of Catechin and Proanthocyanidins in Black Poplar Stems After Infection by Plectosphaerella populi: Hormonal Regulation, Biosynthesis and Antifungal Activity.

Flavan-3-ols including the monomeric catechin and the polymeric proanthocyanidins (PAs) are abundant phenolic metabolites in poplar (Populus spp.) previously described to protect leaves against pathogen infection. However, it is not known whether stems are also defended in this way. Here we investigated flavan-3-ol accumulation, activity, and the regulation of formation in black poplar (P. nigra) stems after infection by a newly described fungal stem pathogen, Plectosphaerella populi, which forms canker-like lesions in stems. We showed that flavan-3-ol contents increased in P. populi-infected black poplar stems over the course of infection compared to non-infected controls. Transcripts of leucoanthocyanidin reductase (LAR) and anthocyanidin reductase (ANR) genes involved in the last steps of flavan-3-ol biosynthesis were also upregulated upon fungal infection indicating de novo biosynthesis. Amending culture medium with catechin and PAs reduced the mycelial growth of P. populi, suggesting that these metabolites act as anti-pathogen defenses in poplar in vivo. Among the hormones, salicylic acid (SA) was higher in P. populi-infected tissues compared to the non-infected controls over the course of infection studied, while jasmonic acid (JA) and JA-isoleucine (JA-Ile) levels were higher than controls only at the early stages of infection. Interestingly, cytokinins (CKs) were also upregulated in P. populi-infected stems. Poplar saplings treated with CK showed decreased levels of flavan-3-ols and SA in stems suggesting a negative association between CK and flavan-3-ol accumulation. Taken together, the sustained upregulation of SA in correlation with catechin and PA accumulation suggests that this is the dominant hormone inducing the formation of antifungal flavan-3-ols during P. populi infection of poplar stems.

COP9 Signalosome CSN4 and CSN5 Subunits Are Involved in Jasmonate-Dependent Defense Against Root-Knot Nematode in Tomato.

COP9 signalosome (CSN) is an evolutionarily conserved regulatory component of the ubiquitin/proteasome system that plays crucial roles in plant growth and stress tolerance; however, the mechanism of COP9-mediated resistance to root-knot nematodes (RKNs, e.g. Meloidogyne incognita) is not fully understood in plants. In the present study, we found that RKN infection in the roots rapidly increases the transcript levels of CSN subunits 4 and 5 (CSN4 and CSN5) and their protein accumulation in tomato (Solanum lycopersicum) plants. Suppression of CSN4 or CSN5 expression resulted in significantly increased number of egg masses and aggravated RKN-induced lipid peroxidation of cellular membrane but inhibited RKN-induced accumulation of CSN4 or CSN5 protein in tomato roots. Importantly, the RKN-induced accumulation of jasmonic acid (JA) and JA-isoleucine (JA-Ile), as well as the transcript levels of JA-related biosynthetic and signaling genes were compromised by CSN4 or CSN5 gene silencing. Moreover, protein–protein interaction assays demonstrated that CSN4 and CSN5B interact with the jasmonate ZIM domain 2 (JAZ2), which is the signaling component of the JA pathway. Silencing of CSN4 or CSN5 also compromises RKN-induced JAZ2 expression. Together, our findings indicate that CSN4 and CSN5 play critical roles in JA-dependent basal defense against RKN.

Abscisic and Jasmonic Acids Contribute to Soybean Tolerance to the Soybean Aphid (Aphis glycines Matsumura)

Plant resistance can provide effective, economical, and sustainable pest control. Tolerance to the soybean aphid has been identified and confirmed in the soybean KS4202. Although its resistance mechanisms are not fully understood, evidence suggests that enhanced detoxification of reactive oxygen species (ROS) is an active system under high aphid infestation. We further explored tolerance by evaluating the differences in constitutive and aphid-induced defenses in KS4202 through the expression of selected defense-related transcripts and the levels of the phytohormones abscisic acid (ABA), jasmonic acid (JA), JA-isoleucine (JA-Ile), cis-(+)-12-oxo-phytodienoic acid (OPDA), and salicylic acid (SA) over several time points. Higher constitutive levels of ABA and JA, and basal expression of ABA- and JA-related transcripts were found in the tolerant genotype. Conversely, aphid-induced defenses in KS4202 were expressed as an upregulation of peroxidases under prolonged aphid infestation (>7 days). Our results point at the importance of phytohormones in constitutive defense in KS4202 tolerance to the soybean aphid. Understanding the underlying mechanisms of tolerance will assist breeding for soybean with these traits, and perhaps help extend the durability of Rag (Resistance to Aphis glycines)-mediated resistance genes.