Genome analysis and hyphal movement characterization of the hitchhiker endohyphal Enterobacter sp. from Rhizoctonia solani.

Bacterial-fungal interactions are pervasive in the rhizosphere. While an increasing number of endohyphal bacteria have been identified, little is known about their ecology and impact on the associated fungal hosts and the surrounding environment. In this study, we characterized the genome of an Enterobacter sp. Crenshaw (En-Cren), which was isolated from the generalist fungal pathogen Rhizoctonia solani, and examined the genetic potential of the bacterium with regard to the phenotypic traits associated with the fungus. Overall, the En-Cren genome size was typical for members of the genus and was capable of free-living growth. The genome was 4.6 MB in size, and no plasmids were detected. Several prophage regions and genomic islands were identified that harbor unique genes in comparison with phylogenetically closely related Enterobacter spp. Type VI secretion system and cyanate assimilation genes were identified from the bacterium, while some common heavy metal resistance genes were absent. En-Cren contains the key genes for indole-3-acetic acid (IAA) and phenylacetic acid (PAA) biosynthesis, and produces IAA and PAA in vitro, which may impact the ecology or pathogenicity of the fungal pathogen in vivo. En-Cren was observed to move along hyphae of R. solani and on other basidiomycetes and ascomycetes in culture. The bacterial flagellum is essential for hyphal movement, while other pathways and genes may also be involved.IMPORTANCEThe genome characterization and comparative genomics analysis of Enterobacter sp. Crenshaw provided the foundation and resources for a better understanding of the ecology and evolution of this endohyphal bacteria in the rhizosphere. The ability to produce indole-3-acetic acid and phenylacetic acid may provide new angles to study the impact of phytohormones during the plant-pathogen interactions. The hitchhiking behavior of the bacterium on a diverse group of fungi, while inhibiting the growth of some others, revealed new areas of bacterial-fungal signaling and interaction, which have yet to be explored.

Maize terpene synthase 1 impacts insect behavior via the production of monoterpene volatiles ß-myrcene and linalool.

Plant-derived volatiles are important mediators of plant-insect interactions as they can provide cues for host location and quality, or act as direct or indirect defense molecules. The volatiles produced by Zea mays (maize) include a range of terpenes, likely produced by several of the terpene synthases (TPS) present in maize. Determining the roles of specific terpene volatiles and individual TPSs in maize-insect interactions is challenging due to the promiscuous nature of TPSs in vitro and their potential for functional redundancy. In this study, we used metabolite GWAS of a sweetcorn diversity panel infested with Spodoptera frugiperda (fall armyworm) to identify genetic correlations between TPSs and individual volatiles. This analysis revealed a correlation between maize terpene synthase 1 (ZmTPS1) and emission of the monoterpene volatiles linalool and ß-myrcene. Electroantennogram assays showed gravid S. frugiperda could detect both linalool and ß-myrcene. Quantification of headspace volatiles in a maize tps1 loss-of-function mutant confirmed that ZmTPS1 is an important contributor to linalool and ß-myrcene emission in maize. Furthermore, pairwise choice assays between tps1 mutant and wild-type plants showed that ZmTPS1, and by extension its volatile products, aid host location in the chewing insect S. frugiperda, yet repel the sap-sucking pest, Rhopalosiphum maidis (corn leaf aphid). On the other hand, ZmTPS1 had no impact on indirect defense via the recruitment of the parasitoid Cotesia marginiventris. ZmTPS1 is therefore an important mediator of the interactions between maize and its insect pests.

The impact of climate change on maize chemical defenses.

Climate change is increasingly affecting agriculture, both at the levels of crops themselves, and by altering the distribution and damage caused by insect or microbial pests. As global food security depends on the reliable production of major crops such as maize (Zea mays), it is vital that appropriate steps are taken to mitigate these negative impacts. To do this a clear understanding of what the impacts are and how they occur is needed. This review focuses on the impact of climate change on the production and effectiveness of maize chemical defenses, including volatile organic compounds, terpenoid phytoalexins, benzoxazinoids, phenolics, and flavonoids. Drought, flooding, heat stress, and elevated concentrations of atmospheric carbon dioxide, all impact the production of maize chemical defenses, in a compound and tissue-specific manner. Furthermore, changes in stomatal conductance and altered soil conditions caused by climate change can impact environmental dispersal and effectiveness certain chemicals. This can alter both defensive barrier formation and multitrophic interactions. The production of defense chemicals is controlled by stress signaling networks. The use of similar networks to co-ordinate the response to abiotic and biotic stress can lead to complex integration of these networks in response to the combinatorial stresses that are likely to occur in a changing climate. The impact of multiple stressors on maize chemical defenses can therefore be different from the sum of the responses to individual stressors and challenging to predict. Much work remains to effectively leverage these protective chemicals in climate-resilient maize.

The evolution of strictly monofunctional naphthoquinol C-methyltransferases is vital in cyanobacteria and plastids.

Prenylated quinones are membrane-associated metabolites that serve as vital electron carriers for respiration and photosynthesis. The UbiE (EC 2.1.1.201)/MenG (EC 2.1.1.163) C-methyltransferases catalyze pivotal ring methylations in the biosynthetic pathways of many of these quinones. In a puzzling evolutionary pattern, prokaryotic and eukaryotic UbiE/MenG homologs segregate into 2 clades. Clade 1 members occur universally in prokaryotes and eukaryotes, excluding cyanobacteria, and include mitochondrial COQ5 enzymes required for ubiquinone biosynthesis; Clade 2 members are specific to cyanobacteria and plastids. Functional complementation of an Escherichia coli ubiE/menG mutant indicated that Clade 1 members display activity with both demethylbenzoquinols and demethylnaphthoquinols, independently of the quinone profile of their original taxa, while Clade 2 members have evolved strict substrate specificity for demethylnaphthoquinols. Expression of the gene-encoding bifunctional Arabidopsis (Arabidopsis thaliana) COQ5 in the cyanobacterium Synechocystis or its retargeting to Arabidopsis plastids resulted in synthesis of a methylated variant of plastoquinone-9 that does not occur in nature. Accumulation of methylplastoquinone-9 was acutely cytotoxic, leading to the emergence of suppressor mutations in Synechocystis and seedling lethality in Arabidopsis. These data demonstrate that in cyanobacteria and plastids, co-occurrence of phylloquinone and plastoquinone-9 has driven the evolution of monofunctional demethylnaphthoquinol methyltransferases and explains why plants cannot capture the intrinsic bifunctionality of UbiE/MenG to simultaneously synthesize their respiratory and photosynthetic quinones.

Maize Terpene Synthase 8 (ZmTPS8) Contributes to a Complex Blend of Fungal-Elicited Antibiotics.

In maize (Zea mays), fungal-elicited immune responses include the accumulation of terpene synthase (TPS) and cytochrome P450 monooxygenases (CYP) enzymes resulting in complex antibiotic arrays of sesquiterpenoids and diterpenoids, including α/ß-selinene derivatives, zealexins, kauralexins and dolabralexins. To uncover additional antibiotic families, we conducted metabolic profiling of elicited stem tissues in mapping populations, which included B73 × M162W recombinant inbred lines and the Goodman diversity panel. Five candidate sesquiterpenoids associated with a chromosome 1 locus spanning the location of ZmTPS27 and ZmTPS8. Heterologous enzyme co-expression studies of ZmTPS27 in Nicotiana benthamiana resulted in geraniol production while ZmTPS8 yielded α-copaene, δ-cadinene and sesquiterpene alcohols consistent with epi-cubebol, cubebol, copan-3-ol and copaborneol matching the association mapping efforts. ZmTPS8 is an established multiproduct α-copaene synthase; however, ZmTPS8-derived sesquiterpene alcohols are rarely encountered in maize tissues. A genome wide association study further linked an unknown sesquiterpene acid to ZmTPS8 and combined ZmTPS8-ZmCYP71Z19 heterologous enzyme co-expression studies yielded the same product. To consider defensive roles for ZmTPS8, in vitro bioassays with cubebol demonstrated significant antifungal activity against both Fusarium graminearum and Aspergillus parasiticus. As a genetically variable biochemical trait, ZmTPS8 contributes to the cocktail of terpenoid antibiotics present following complex interactions between wounding and fungal elicitation.

Metabolic link between auxin production and specialized metabolites in Sorghum bicolor.

Aldoximes are amino acid derivatives that serve as intermediates for numerous specialized metabolites including cyanogenic glycosides, glucosinolates, and auxins. Aldoxime formation is mainly catalyzed by cytochrome P450 monooxygenases of the 79 family (CYP79s) that can have broad or narrow substrate specificity. Except for SbCYP79A1, aldoxime biosynthetic enzymes in the cereal sorghum (Sorghum bicolor) have not been characterized. This study identified nine CYP79-encoding genes in the genome of sorghum. A phylogenetic analysis of CYP79 showed that SbCYP79A61 formed a subclade with maize ZmCYP79A61, previously characterized to be involved in aldoxime biosynthesis. Functional characterization of this sorghum enzyme using transient expression in Nicotiana benthamiana and stable overexpression in Arabidopsis thaliana revealed that SbCYP79A61 catalyzes the production of phenylacetaldoxime (PAOx) from phenylalanine but, unlike the maize enzyme, displays no detectable activity against tryptophan. Additionally, targeted metabolite analysis after stable isotope feeding assays revealed that PAOx can serve as a precursor of phenylacetic acid (PAA) in sorghum and identified benzyl cyanide as an intermediate of PAOx-derived PAA biosynthesis in both sorghum and maize. Taken together, our results demonstrate that SbCYP79A61 produces PAOx in sorghum and may serve in the biosynthesis of other nitrogen-containing phenylalanine-derived metabolites involved in mediating biotic and abiotic stresses.

Production of the Green Leaf Volatile (Z)-3-Hexenal by a Zea mays Hydroperoxide Lyase.

Plant-produced volatile compounds play important roles in plant signaling and in the communication of plants with other organisms. Many plants emit green leaf volatiles (GLVs) in response to damage or attack, which serve to warn neighboring plants or attract predatory or parasitic insects to help defend against insect pests. GLVs include aldehydes, esters, and alcohols of 6-carbon compounds that are released rapidly following wounding. One GLV produced by maize (Zea mays) is the volatile (Z)-3-hexenal; this volatile is produced from the cleavage of (9Z,11E,15Z)-octadecatrienoic acid by hydroperoxide lyases (HPLs) of the cytochrome P450 CYP74B family. The specific HPL in maize involved in (Z)-3-hexenal production had not been determined. In this study, we used phylogenetics with known HPLs from other species to identify a candidate HPL from maize (ZmHPL). To test the ability of the putative HPL to produce (Z)-3-hexenal, we constitutively expressed the gene in Arabidopsis thaliana ecotype Columbia-0 that contains a natural loss-of-function mutant in AtHPL and examined the transgenic plants for restored (Z)-3-hexenal production. Volatile analysis of leaves from these transgenic plants showed that they did produce (Z)-3-hexenal, confirming that ZmHPL can produce (Z)-3-hexenal in vivo. Furthermore, we used gene expression analysis to show that expression of ZmHPL is induced in maize in response to both wounding and the insect pests Spodoptera frugiperda and Spodoptera exigua. Our study demonstrates that ZmHPL can produce GLVs and highlights its likely role in (Z)-3-hexenal production in response to mechanical damage and herbivory in maize.

Dedicated farnesyl diphosphate synthases circumvent isoprenoid-derived growth-defense tradeoffs in Zea mays.

Zea mays (maize) makes phytoalexins such as sesquiterpenoid zealexins, to combat invading pathogens. Zealexins are produced from farnesyl diphosphate in microgram per gram fresh weight quantities. As farnesyl diphosphate is also a precursor for many compounds essential for plant growth, the question arises as to how Z. mays produces high levels of zealexins without negatively affecting vital plant systems. To examine if specific pools of farnesyl diphosphate are made for zealexin synthesis we made CRISPR/Cas9 knockouts of each of the three farnesyl diphosphate synthases (FPS) in Z. mays and examined the resultant impacts on different farnesyl diphosphate-derived metabolites. We found that FPS3 (GRMZM2G098569) produced most of the farnesyl diphosphate for zealexins, while FPS1 (GRMZM2G168681) made most of the farnesyl diphosphate for the vital respiratory co-factor ubiquinone. Indeed, fps1 mutants had strong developmental phenotypes such as reduced stature and development of chlorosis. The replication and evolution of the fps gene family in Z. mays enabled it to produce dedicated FPSs for developmentally related ubiquinone production (FPS1) or defense-related zealexin production (FPS3). This partitioning of farnesyl diphosphate production between growth and defense could contribute to the ability of Z. mays to produce high levels of phytoalexins without negatively impacting its growth.

Xanthomonas hortorum pv. gardneri TAL effector AvrHah1 is necessary and sufficient for increased persistence of Salmonella enterica on tomato leaves.

Salmonella enterica is ubiquitous in the plant environment, persisting in the face of UV stress, plant defense responses, desiccation, and nutrient limitation. These fluctuating conditions of the leaf surface result in S. enterica population decline. Biomultipliers, such as the phytopathogenic bacterium Xanthomonas hortorum pv. gardneri (Xhg), alter the phyllosphere to the benefit of S. enterica. Specific Xhg-dependent changes to this niche that promote S. enterica persistence remain unclear, and this work focuses on identifying factors that lead to increased S. enterica survival on leaves. Here, we show that the Xhg transcription activator-like effector AvrHah1 is both necessary and sufficient for increased survival of S. enterica on tomato leaves. An Xhg avrHah1 mutant fails to influence S. enterica survival while addition of avrHah1 to X. vesicatoria provides a gain of function. Our results indicate that although Xhg stimulates a robust immune response from the plant, AvrHah1 is not required for these effects. In addition, we demonstrate that cellular leakage that occurs during disease is independent of AvrHah1. Investigation of the interaction between S. enterica, Xhg, and the plant host provides information regarding how an inhospitable environment changes during infection and can be transformed into a habitable niche.

Kaempferol as a precursor for ubiquinone (coenzyme Q) biosynthesis: An atypical node between specialized metabolism and primary metabolism.

Ubiquinone (coenzyme Q) is a vital respiratory cofactor and liposoluble antioxidant. Studies have shown that plants derive approximately a quarter of 4-hydroxybenzoate, which serves as the direct ring precursor of ubiquinone, from the catabolism of kaempferol. Biochemical and genetic evidence suggests that the release of 4-hydroxybenzoate from kaempferol is catalyzed by heme-dependent peroxidases and that 3-O-glycosylations of kaempferol act as a negative regulator of this process. These findings not only represent an atypical instance of primary metabolite being derived from specialized metabolism but also raise the question as to whether ubiquinone contributes to the ROS scavenging and signaling functions already established for flavonols.

A dedicated flavin-dependent monooxygenase catalyzes the hydroxylation of demethoxyubiquinone into ubiquinone (coenzyme Q) in Arabidopsis.

Ubiquinone (Coenzyme Q) is a vital respiratory cofactor and liposoluble antioxidant. In plants, it is not known how the C-6 hydroxylation of demethoxyubiquinone, the penultimate step in ubiquinone biosynthesis, is catalyzed. The combination of cross-species gene network modeling along with mining of embryo-defective mutant databases of Arabidopsis thaliana identified the embryo lethal locus EMB2421 (At1g24340) as a top candidate for the missing plant demethoxyubiquinone hydroxylase. In marked contrast with prototypical eukaryotic demethoxyubiquinone hydroxylases, the catalytic mechanism of which depends on a carboxylate-bridged di-iron domain, At1g24340 is homologous to FAD-dependent oxidoreductases that instead use NAD(P)H as an electron donor. Complementation assays in Saccharomyces cerevisiae and Escherichia coli demonstrated that At1g24340 encodes a functional demethoxyubiquinone hydroxylase and that the enzyme displays strict specificity for the C-6 position of the benzoquinone ring. Laser-scanning confocal microscopy also showed that GFP-tagged At1g24340 is targeted to mitochondria. Silencing of At1g24340 resulted in 40 to 74% decrease in ubiquinone content and de novo ubiquinone biosynthesis. Consistent with the role of At1g24340 as a benzenoid ring modification enzyme, this metabolic blockage could not be bypassed by supplementation with 4-hydroxybenzoate, the immediate precursor of ubiquinone's ring. Unlike in yeast, in Arabidopsis overexpression of demethoxyubiquinone hydroxylase did not boost ubiquinone content. Phylogenetic reconstructions indicated that plant demethoxyubiquinone hydroxylase is most closely related to prokaryotic monooxygenases that act on halogenated aromatics and likely descends from an event of horizontal gene transfer between a green alga and a bacterium.

A maize leucine-rich repeat receptor-like protein kinase mediates responses to fungal attack.

MAIN CONCLUSION: A maize receptor kinase controls defense response to fungal pathogens by regulating jasmonic acid and antimicrobial phytoalexin production. Plants use a range of pattern recognition receptors to detect and respond to biotic threats. Some of these receptors contain leucine-rich repeat (LRR) domains that recognize microbial proteins or peptides. Maize (Zea mays) has 226 LRR-receptor like kinases, making it challenging to identify those important for pathogen recognition. In this study, co-expression analysis with genes for jasmonic acid and phytoalexin biosynthesis was used to identify a fungal induced-receptor like protein kinase (FI-RLPK) likely involved in the response to fungal pathogens. Loss-of-function mutants in fi-rlpk displayed enhanced susceptibility to the necrotrophic fungal pathogen Cochliobolus heterostrophus and reduced accumulation of jasmonic acid and the anti-microbial phytoalexins -kauralexins and zealexins- in infected tissues. In contrast, fi-rlpk mutants displayed increased resistance to stem inoculation with the hemibiotrophic fungal pathogen Fusarium graminearum. These data indicate that FI-RLPK is important for fungal recognition and activation of defenses, and that F. graminearum may be able to exploit FI-RLPK function to increase its virulence.

Metabolite analysis of Arabidopsis CYP79A2 overexpression lines reveals turnover of benzyl glucosinolate and an additive effect of different aldoximes on phenylpropanoid repression.

Indole-3-acetaldoxime (IAOx) and phenylacetaldoxime (PAOx) are precursors for the growth hormones indole-3-acetic acid (IAA) and phenylacetic acid (PAA) and the defense compounds glucosinolates in Brassicales. Our recent work has shown that Arabidopsis transgenic lines overexpressing AtCYP79A2, a PAOx-production enzyme, accumulate the PAOx-derived compounds benzyl glucosinolate and PAA. Here we report that they also accumulate the benzyl glucosinolate hydrolysis products benzyl isothiocyanate and benzyl cyanide, which indicates that the turnover of benzyl glucosinolate can occur in intact tissues. Myrosinases or ß-glucosidases are known to catalyze glucosinolate breakdown. However, transcriptomics analysis detected no substantial increase in expression of known myrosinases or putative ß-glucosidases in AtCYP79A2 overexpressing lines. It was previously shown that accumulation of aldoximes or their derivatives represses the phenylpropanoid pathway. For instance, ref2 mutant having a defect in one of the aldoxime catabolic enzymes decreases phenylpropanoid production. Considering that AtCYP79A2 is not expressed in most organs under optimal growth condition, ref2 accumulates aliphatic aldoximes but not PAOx. Interestingly, overexpression of AtCYP79A2 in ref2 resulted in a further decrease in sinapoylmalate content compared to ref2. This indicates that accumulation of PAOx has an additive effect on phenylpropanoid pathway suppression mediated by other aldoximes.

Zea mays Volatiles that Influence Oviposition and Feeding Behaviors of Spodoptera frugiperda.

Fall armyworm (Spodoptera frugiperda) is a major global pest of many crops, including maize (Zea mays). This insect is known to use host plant-derived volatile organic compounds to locate suitable hosts during both its adult and larval stages, yet the function of individual compounds remains mostly enigmatic. In this study, we use a combination of volatile profiling, electrophysiological assays, pair-wise choice behavioral assays, and chemical supplementation treatments to identify and assess specific compounds from maize that influence S. frugiperda host location. Our findings reveal that methyl salicylate and (E)-alpha-bergamotene are oviposition attractants for adult moths but do not impact larval behavior. While geranyl acetate can act as an oviposition attractant or repellent depending on the host volatile context and (E)-4,8-dimethyl-1,3,7-nonatriene (DMNT) is an oviposition deterrent. These compounds can also be attractive to the larvae when applied to specific maize inbreds. These data show that S. frugiperda uses different plant volatile cues for host location in its adult and larval stage and that the background volatile context that specific volatiles are perceived in, alters their impact as behavioral cues.

Aldoximes are precursors of auxins in Arabidopsis and maize.

Two natural auxins, phenylacetic acid (PAA) and indole-3-acetic acid (IAA), play crucial roles in plant growth and development. One route of IAA biosynthesis uses the glucosinolate intermediate indole-3-acetaldoxime (IAOx) as a precursor, which is thought to occur only in glucosinolate-producing plants in Brassicales. A recent study showed that overproducing phenylacetaldoxime (PAOx) in Arabidopsis increases PAA production. However, it remains unknown whether this increased PAA resulted from hydrolysis of PAOx-derived benzyl glucosinolate or, like IAOx-derived IAA, is directly converted from PAOx. If glucosinolate hydrolysis is not required, aldoxime-derived auxin biosynthesis may occur beyond Brassicales. To better understand aldoxime-derived auxin biosynthesis, we conducted an isotope-labelled aldoxime feeding assay using an Arabidopsis glucosinolate-deficient mutant sur1 and maize, and transcriptomics analysis. Our study demonstrated that the conversion of PAOx to PAA does not require glucosinolates in Arabidopsis. Furthermore, maize produces PAA and IAA from PAOx and IAOx, respectively, indicating that aldoxime-derived auxin biosynthesis also occurs in maize. Considering that aldoxime production occurs widely in the plant kingdom, aldoxime-derived auxin biosynthesis is likely to be more widespread than originally believed. A genome-wide transcriptomics study using PAOx-overproduction plants identified complex metabolic networks among IAA, PAA, phenylpropanoid and tryptophan metabolism.

3-O-glycosylation of kaempferol restricts the supply of the benzenoid precursor of ubiquinone (Coenzyme Q) in Arabidopsis thaliana.

Ubiquinone (Coenzyme Q) is a vital respiratory cofactor and antioxidant in eukaryotes. The recent discovery that kaempferol serves as a precursor for ubiquinone's benzenoid moiety both challenges the conventional view of flavonoids as specialized metabolites, and offers new prospects for engineering ubiquinone in plants. Here, we present evidence that Arabidopsis thaliana mutants lacking kaempferol 3-O-rhamnosyltransferase (ugt78d1) and kaempferol 3-O-glucosyltransferase (ugt78d2) activities display increased de novo biosynthesis of ubiquinone and increased ubiquinone content. These data are congruent with the proposed model that unprotected C-3 hydroxyl of kaempferol triggers the oxidative release of its B-ring as 4-hydroxybenzoate, which in turn is incorporated into ubiquinone. Ubiquinone content in the ugt78d1/ugt78d2 double knockout represented 160% of wild-type level, matching that achieved via exogenous feeding of 4-hydroxybenzoate to wild-type plants. This suggests that 4-hydroxybenzoate is no longer limiting ubiquinone biosynthesis in the ugt78d1/ugt78d2 plants. Evidence is also shown that the glucosylation of 4-hydroxybenzoate as well as the conversion of the immediate precursor of kaempferol, dihydrokaempferol, into dihydroquercetin do not compete with ubiquinone biosynthesis in A. thaliana.

Metabolomics by UHPLC-HRMS reveals the impact of heat stress on pathogen-elicited immunity in maize.

INTRODUCTION: Studies investigating crop resistance to abiotic and biotic stress have largely focused on plant responses to singular forms of stress and individual biochemical pathways that only partially represent stress responses. Thus, combined abiotic and biotic stress treatments and the global assessment of their elicited metabolic expression remains largely unexplored. In this study, we employed targeted and untargeted metabolomics to investigate the molecular responses of maize (Zea mays) to abiotic, biotic, and combinatorial stress. OBJECTIVE: We compared the inducible metabolomes of heat-stressed (abiotic) and C. heterostrophus-infected (biotic) maize and examined the effects of heat stress on the ability of maize to defend itself against C. heterostrophus. METHODS: Ultra-high-performance liquid chromatography-high-resolution mass spectrometry was performed on plants grown under control conditions (28 °C), heat stress (38 °C), Cochliobolus heterostrophus infection, or combinatorial stress [heat (38 °C) + C. heterostrophus infection]. RESULTS: Multivariate analyses revealed differential metabolite expression between heat stress, C. heterostrophus infection, and their respective controls. In combinatorial experiments, treatment with heat stress prior to fungal inoculation negatively impacted maize disease resistance against C. heterostrophus, and distinct metabolome separation between combinatorial stressed plants and the non-heat-stressed infected controls was observed. Targeted analysis revealed inducible primary and secondary metabolite responses to abiotic/biotic stress, and combinatorial experiments indicated that deficiency in the hydroxycinnamic acid, p-coumaric acid, may contribute to the heat-induced susceptibility of maize to C. heterostrophus. CONCLUSION: These findings demonstrate that abiotic stress can predispose crops to more severe disease symptoms, underlining the increasing need to investigate defense chemistry in plants under combinatorial stress.

The 13-lipoxygenase MSD2 and the ω-3 fatty acid desaturase MSD3 impact Spodoptera frugiperda resistance in Sorghum.

MAIN CONCLUSION: Linolenic acid produced by the ω-3 fatty acid desaturase MSD3 in sorghum is used for insect-induced jasmonic acid production and is important for resistance against Spodoptera frugiperda. Jasmonic acid (JA) is a phytohormone that regulates both plant development and stress responses. In sorghum (Sorghum bicolor), the ω-3 fatty acid desaturase Multiseeded3 (MSD3) and the 13-lipoxygenase Multiseeded2 (MSD2) are important for producing JA to regulate panicle development and spikelet fertility, but their function in plant defense remains unknown. In this study, we examined whether these genes are important for the production of JA in response to herbivory by the insect pest Spodoptera frugiperda. Compared to wild-type controls, the msd3 mutant accumulated less JA in leaves of both infested and uninfested plants, revealing that MSD3 is involved in stress-induced JA production. In contrast, herbivore-induced JA production in the msd2 mutant was indistinguishable from wild type, indicating that MSD2 does not function in herbivore-induced JA production. An increase of S. frugiperda growth was observed on both the msd3 and msd2 mutants, hinting at roles for both JA and additional oxylipins in sorghum's defense responses.

Setaria viridis as a model for translational genetic studies of jasmonic acid-related insect defenses in Zea mays.

Little is known regarding insect defense pathways in Setaria viridis (setaria), a model system for panicoid grasses, including Zea mays (maize). It is thus of interest to compare insect herbivory responses of setaria and maize. Here we use metabolic, phylogenetic, and gene expression analyses to measure a subset of jasmonic acid (JA)-related defense responses to leaf-chewing caterpillars. Phylogenetic comparisons of known defense-related maize genes were used to identify putative orthologs in setaria, and candidates were tested by quantitative PCR to determine transcriptional responses to insect challenge. Our findings show that while much of the core JA-related metabolic and genetic responses appear conserved between setaria and maize, production of downstream secondary metabolites such as benzoxazinoids and herbivore-induced plant volatiles are dissimilar. This diversity of chemical defenses and gene families involved in secondary metabolism among grasses presents new opportunities for cross species engineering. The high degree of genetic similarity and ease of orthologous gene identification between setaria and maize make setaria an excellent species for translational genetic studies, but the species specificity of downstream insect defense chemistry makes some pathways unamenable to cross-species comparisons.

Fighting on two fronts: Elevated insect resistance in flooded maize.

To grow and thrive plants must be able to adapt to both adverse environmental conditions and attack by a variety of pests. Elucidating the sophisticated mechanisms plants have developed to achieve this has been the focus of many studies. What is less well understood is how plants respond when faced with multiple stressors simultaneously. In this study, we assess the response of Zea mays (maize) to the combinatorial stress of flooding and infestation with the insect pest Spodoptera frugiperda (fall armyworm). This combined stress leads to elevated production of the defence hormone salicylic acid, which does not occur in the individual stresses, and the resultant salicylic acid-dependent increase in S. frugiperda resistance. Remodelling of phenylpropanoid pathways also occurs in response to this combinatorial stress leading to increased production of the anti-insect C-glycosyl flavones (maysins) and the herbivore-induced volatile phenolics, benzyl acetate, and phenethyl acetate. Furthermore, changes in cellular redox status also occur, as indicated by reductions in peroxidase and polyphenol oxidase activity. These data suggest that metabolite changes important for flooding tolerance and anti-insect defence may act both additively and synergistically to provide extra protection to the plant.