A Tea Hydroperoxide Lyase Gene,CsiHPL1, Regulates Tomato Defense Response Against Prodenia Litura (Fabricius) and Alternaria Alternata f. sp. Lycopersici by Modulating Green Leaf Volatiles (GLVs) Release and Jasmonic Acid (JA) Gene Expression

Hydroperoxide lyases (HPLs) play important roles in modulating plant defense by regulating the release of green leaf volatiles (GLVs) and the jasmonic acid (JA) pathway. CsiHPL1—a chloroplast-localized tea gene that encodes HPL—was previously cloned and predicted to be a regulator of plant defense responses. CsiHPL1 was expressed constitutively in transgenic tomato (Solanum lycopersicum) plants to define its function in plant defense. CsiHPL1 overexpression caused tomato to release more constitutive and wound-induced GLVs [including (Z)-hexenal and (Z)-3-hexen-1-ol]. CsiHPL1 transgenic lines also exhibited lower levels of resistance to the larva of the tomato-chewing herbivore Prodenia litura (Fabricius) but enhanced resistance to the necrotrophic fungus Alternaria alternata f. sp. lycopersici (AAL). Furthermore, transgenic lines exhibited decreased expression levels of JA-related genes (SlAOS and SlPI-II) induced by P. litura and AAL infection. We thus concluded that constitutive expression of CsiHPL1 can regulate tomato resistance to P. litura and AAL by modulating GLV release and JA gene expression. Application of these results will be helpful in controlling plant defenses against herbivore attack and fungal disease.     

Roles of (Z)-3-hexenol in plant-insect interactions

Green leaf C6-volatiles are among the most important herbivore-induced plant volatiles (HIPVs). They play important roles in mediating the behavior of herbivores and their natural enemies, and in triggering the plant-plant communication to prevent further attacks. Recently, wound-induced ubiquitous (Z)-3-hexenol, a C6-alcohol synthesized in the lipoxygenase/HPL pathway, was proved to be the most important info chemical for the herbivore repellence/attraction and natural enemy attraction in tritrophic interactions, as well as for the induction of gene expression in neighboring unattacked plants. In spite of the conflict functions of (Z)-3-hexenol in direct and indirect plant defenses, its positive roles in the indirect defense and the priming effect are consistent. Therefore, this compound can be used to develop novel insect pest control strategies.Key words: green leaf volatiles, (Z)-3-hexenol, direct defense, indirect defense, primingTo date, nearly 2000 volatile compounds have been identified in plant species from over 90 families.¹ These compounds are released from plant organs above or below the ground, and some are induced by biotic activities. Herbivore feeding stimulates the plants to release green leaf volatiles (GLVs), terpenoids, nitrogen-containing nitriles and oximes, methyl salicylate, etc. Production of these volatiles by plants involves at least three biosynthetic pathways: the fatty acid/lipoxygenase pathway for green leaf volatiles, the isoprenoid pathway for terpenoids, and the shikimic acid pathway for methyl salicylate.² Herbivore-damaged plants emit some of the most common GLVs and terpenoids that play important roles in mediating the behaviors of herbivores and their natural enemies, as well as in triggering the plant-plant communication.¹ Recently, functional studies on green leaf C6-volatiles have received wide attention and made exciting progresses. Especially, accumulating evidences on the C6-volatile (Z)-3-hexenol support its role in mediating indirect defense responses of plant.     

Feeding on tea GH19 chitinase enhances tea defense responses induced by regurgitant derived from Ectropis grisescens

Multiple isoforms of chitinases participate in plant defense against outside invaders. However, the functions of hydrolase family 19 (GH19) chitinases on pest control remain largely unknown. Here we reported the isolation and functional analysis of a gene CsChi19, which encodes a GH19 endochitinase protein of 332 amino acid residues from tea plant (Camellia sinensis). CsChi19 expression levels were upregulated in response to mechanical wounding, infestation by two important pests: the tea geometrid Ectropis grisescens and the tea green leafhopper Empoasca (Matsumurasca) onukii, a fungal pathogen Colletotrichum fructicola, and treatment with two phytohormones: jasmonic acid (JA) and salicylic acid. CsChi19 was heterologously expressed in Escherichia coli, and its catalytic function was further elucidated. The protein could hydrolyze colloidal chitin, and the optimum temperature and pH for its activity was 40°C and pH 5.0. CsChi19 were found to be toxic to tea pests when they were fed on artificial diets containing this protein. Interestingly, the regurgitant derived from E. grisescens fed with artificial diets containing CsChi19 protein induced stronger expression of CsMPK3, more JA burst, more accumulation of defense-related secondary metabolites, and more emission of volatiles than the regurgitant derived from E. grisescens fed only with artificial diets. Our results provide first evidence that CsChi19 is involved in mediating a novel defense mechanism of tea plant through altering the composition of the regurgitant.     

The rice hydroperoxide lyase OsHPL3 functions in defense responses by modulating the oxylipin pathway

As important signal molecules, jasmonates (JAs) and green leaf volatiles (GLVs) play diverse roles in plant defense responses against insect pests and pathogens. However, how plants employ their specific defense responses by modulating the levels of JA and GLVs remains unclear. Here, we describe identification of a role for the rice HPL3 gene, which encodes a hydroperoxide lyase (HPL), OsHPL3/CYP74B2, in mediating plant‐specific defense responses. The loss‐of‐function mutant hpl3‐1 produced disease‐resembling lesions spreading through the whole leaves. A biochemical assay revealed that OsHPL3 possesses intrinsic HPL activity, hydrolyzing hydroperoxylinolenic acid to produce GLVs. The hpl3‐1 plants exhibited enhanced induction of JA, trypsin proteinase inhibitors and other volatiles, but decreased levels of GLVs including (Z)‐3‐hexen‐1‐ol. OsHPL3 positively modulates resistance to the rice brown planthopper [BPH, Nilaparvata lugens (Stål)] but negatively modulates resistance to the rice striped stem borer [SSB, Chilo suppressalis (Walker)]. Moreover, hpl3‐1 plants were more attractive to a BPH egg parasitoid, Anagrus nilaparvatae, than the wild‐type, most likely as a result of increased release of BPH‐induced volatiles. Interestingly, hpl3‐1 plants also showed increased resistance to bacterial blight (Xanthomonas oryzae pv. oryzae). Collectively, these results indicate that OsHPL3, by affecting the levels of JA, GLVs and other volatiles, modulates rice‐specific defense responses against different invaders.     

Antisense LOX expression increases herbivore performance by decreasing defense responses and inhibiting growth-related transcriptional reorganization in Nicotiana attenuata

Inhibition of jasmonic acid (JA) signaling has been shown to decrease herbivore resistance, but the responsible mechanisms are largely unknown because insect resistance is poorly understood in most model plant systems. We characterize three members of the lipoxygenase (LOX) gene family in the native tobacco plant Nicotiana attenuata and manipulate, by antisense expression, a specific, wound- and herbivory-induced isoform (LOX3) involved in JA biosynthesis. In three independent lines, antisense expression reduced wound-induced JA accumulation but not the release of green leaf volatiles (GLVs). The impaired JA signaling reduced two herbivore-induced direct defenses, nicotine and trypsin protease inhibitors (TPI), as well as the potent indirect defense, the release of volatile terpenes that attract generalist predators to feeding herbivores. All these defenses could be fully restored by methyl-JA (MeJA) treatment, with the exception of the increase in TPI activity, which was partially restored, suggesting the involvement of additional signals. The impaired ability to produce chemical defenses resulted in lower resistance to Manduca sexta attack, which could also be restored by MeJA treatment. Expression analysis using a cDNA microarray, specifically designed to analyze M. sexta-induced gene expression in N. attenuata, revealed a pivotal role for LOX3-produced oxylipins in upregulating defense genes (protease inhibitor, PI; xyloglucan endotransglucosylase/hydrolase, XTH; threonine deaminase, TD; hydroperoxide lyase, HPL), suppressing both downregulated growth genes (RUBISCO and photosystem II, PSII) and upregulated oxylipin genes (alpha-dioxygenase, alpha-DOX). By genetically manipulating signaling in a plant with a well-characterized ecology, we demonstrate that the complex phenotypic changes that mediate herbivore resistance are controlled by a specific part of the oxylipin cascade.     

Wound-induced green leaf volatiles cause the release of acetylated derivatives and a terpenoid in maize

Green leaf volatiles (GLVs), generally occurring C6 alcohols, aldehydes and acetates from plants, play an important role in plant-plant communication. These compounds induce intact plants to produce jasmonic acid, and induce defense-related gene expression and the release of volatile compounds. Here, we address wound-induced GLVs cause the release of acetylated derivatives and a terpenoid, (E)-4,8-dimethylnona-1,3,7-triene (DMNT) in intact maize, which may be a type of plant-plant interaction mediated by airborne GLVs. Upon exposure of intact maize seedlings to wound-induced GLVs, (Z)-3-hexenyl acetate was consistently the most abundant compound released. Exogenous application of individual alcohols and aldehydes mostly resulted in the release of corresponding acetate esters. C6-alcohols with a double bond between the second and third, or the third and fourth carbon atoms, C5- or C6-aldehydes, and (Z)-3-hexenyl acetate triggered the release of DMNT. When (Z)-3-hexenyl acetate and hexyl acetate were used to treat maize seedlings, they were recovered from the plants. These data demonstrated that: (1) apart from direct adsorption and re-release of acetate esters, absorption and conversion of exogenous alcohols and aldehydes into acetate esters occurred, and (2) DMNT was induced by a range of aldehydes and unsaturated alcohols.     

Induced defenses of Veronica spicata: Variability in herbivore-induced volatile organic compounds

Plants release volatile organic compounds (VOCs) that have many eco-physiological functions. Induction of plant VOCs is known to occur upon herbivory. Herbivore-induced VOCs are involved in the attraction of predators and parasitoids, a phenomenon known as an indirect defense of plants. We measured the VOC profiles of the wild species Veronica spicata with and without larval feeding and oviposition by the specialist butterfly Melitaea cinxia. V. spicata showed great plasticity when deploying indirect defences. The induction of several ubiquitous terpenoids and green leaf volatiles (GLVs) was associated with larval feeding, whereas the increase of two ketones, 6-methyl-5-hepten-2-one and t-geranylacetone and the suppression of GLVs were associated with oviposition by the butterfly.     

Enhanced transcriptome responses in herbivore-infested tea plants by the green leaf volatile (Z)-3-hexenol

Journal of Plant Research - Green leaf volatiles (GLVs) play a vital role in enhancing herbivore-associated defense responses, but the mechanism by which they precisely regulate such responses is...     

Variability and stability of tea weevil‐induced volatile emissions from tea plants with different weevil densities,photoperiod and infestation duration

Abstract After herbivore attack, many plants emit herbivore‐induced plant volatiles (HIPVs). HIPVs can attract carnivores and/or repel herbivores, thereby mediating tritrophic plant–herbivore–carnivore interactions. HIPVs act as chemical information between organisms; hence, their variability and stability are vital. In the present study, variations in the volatile emissions, from the tea plant Camellia sinensis (O. Ktze) damaged by the tea weevil Myllocerinus aurolineatus (Voss) (Coleoptera: Curculionidae), with weevil densities, photoperiod and infestation duration, were investigated. The volatiles induced by high‐density weevils were more abundant in composition and amount than those induced by low‐density weevils, whether at noon, night or after weevil removal. The induced volatile emissions were similar on the second and third day after infestation, and the emissions of the major induced compounds displayed diurnal cycles. Linalool, (E,E)‐α‐farnesene, and benzyl nitrile were emitted mainly at noon, whereas 1,3,8‐p‐menthatriene and (E)‐β‐ocimene were maximally emitted at night. Given the different emission dynamics, significant differences were found between noon‐ and night‐induced volatiles. In summary, tea plants damaged by different weevil densities emitted a relatively stable signal at a particular time. This stability could be attributed to the similarities under the two densities of the main induced volatile compounds, their relative ratios and the emission dynamics of the induced volatiles.     

3,3-Dimethyl-1-butanol, a parakairomone component to Aleurodicus dispersus (Hemiptera: Aleyrodidae)

C-6-based green leaf volatiles (GLVs) are signal molecules to herbivorous insects and play an important role in plant–herbivore interactions. How isomerization of GLVs affects insect’s olfactory response has been rarely tested. In laboratory and field experiments, we examined the effect of hexanol isomers on olfactory orientation of the spiraling whitefly, Aleruodicus dispersus Russell, a highly polyphagous pest. In a Y-tube oflactometer, we found that (±)-2-hexanol, 3-methyl-3-pentanol and 3,3-dimethyl-1-butanol significantly attracted female A. dispersus. The trap captures of 3,3-dimethyl-1-butanol were significantly more than that of (±)-2-hexanol and 3-methyl-3-pentanol, and its optimum concentration was 1 μ1/ml. We suggest that the anthropogenic compound 3,3-dimethyl-1-butanol can be exploited as a parakairomone (synthetic analogues of kairomone) to monitor and control adult A. dispersus.     

Isolation and characterization of Histone1 gene and its promoter from tea plant (Camellia sinensis)

One 1.2 kbp long sequence was cloned by using PCR with primers that were designed from cDNA sequence of CsH1 gene (Genbank: EU716314) from tea plant (Camellia sinensis). According to the 1.2 kbp sequence, a 0.6 kbp sequence was isolated from tea plant genomic DNA using DNA Walking Method. Sequence analysis revealed that the 1.2 kbp sequence is a CsH1 gene consisting of 1 exon and 2 introns, the border of exton and intron sequences conforming to the GT–AG rule, and the 0.6 kbp sequence was found to be the promoter of CsH1 gene which contains basic promoter elements, TATA-box and CAAT-box. Abscisic acid responsiveness cis-acting element, elictor-responsive element, GA response element, light response cis-acting element and TC-rich repeats were also represented. To further study the activity of this promoter, the sequence was used to drive a GUS fusion gene in Agrobacterium-mediated transformation of tea plant somatic embryos, leaf discs and calli of tobacco (Nicotiana tabacum L.) where a high level of GUS expression was both observed in the tobacco calli and tea plant somatic embryos. These results suggest that the CsH1 gene promoter isolated is capable of conferring nuclear gene expression.     

Three-step pathway engineering results in more incidence rate and higher emission of nerolidol and improved attraction of Diadegma semiclausum

The concentration and ratio of terpenoids in the headspace volatile blend of plants have a fundamental role in the communication of plants and insects. The sesquiterpene (E)-nerolidol is one of the important volatiles with effect on beneficial carnivores for biologic pest management in the field. To optimize de novo biosynthesis and reliable and uniform emission of (E)-nerolidol, we engineered different steps of the (E)-nerolidol biosynthesis pathway in Arabidopsis thaliana.Introduction of a mitochondrial nerolidol synthase gene mediates de novo emission of (E)-nerolidol and linalool. Co-expression of the mitochondrial FPS1 and cytosolic HMGR1 increased the number of emitting transgenic plants (incidence rate) and the emission rate of both volatiles. No association between the emission rate of transgenic volatiles and their growth inhibitory effect could be established. (E)-Nerolidol was to a large extent metabolized to non-volatile conjugates.     

Green leaf volatiles protect maize (Zea mays) seedlings against damage from cold stress

Although considerable evidence has accumulated on the defensive activity of plant volatile organic compounds against pathogens and insect herbivores, less is known about the significance of volatile organic compounds emitted by plants under abiotic stress. Here, we report that green leaf volatiles (GLVs), which were previously shown to prime plant defences against insect herbivore attack, also protect plants against cold stress (4 °C). We show that the expression levels of several cold stress‐related genes are significantly up‐regulated in maize (Zea mays) seedlings treated with physiological concentrations of the GLV, (Z)‐3‐hexen‐1‐yl acetate (Z‐3‐HAC), and that seedlings primed with Z‐3‐HAC exhibit increased growth and reduced damage after cold stress relative to unprimed seedlings. Together, these data demonstrate the protective and priming effect of GLVs against cold stress and suggest an activity of GLVs beyond the activation of typical plant defence responses against herbivores and pathogens.     

Traumatin- and Dinortraumatin-containing Galactolipids in Arabidopsis: THEIR FORMATION IN TISSUE-DISRUPTED LEAVES AS COUNTERPARTS OF GREEN LEAF VOLATILES*

Green leaf volatiles (GLVs) consisting of six-carbon aldehydes, alcohols, and their esters, are biosynthesized through the action of fatty acid hydroperoxide lyase (HPL), which uses fatty acid hydroperoxides as substrates. GLVs form immediately after disruption of plant leaf tissues by herbivore attacks and mechanical wounding and play a role in defense against attackers that attempt to invade through the wounds. The fates and the physiological significance of the counterparts of the HPL reaction, the 12/10-carbon oxoacids that are formed from 18/16-carbon fatty acid 13-/11-hydroperoxides, respectively, are largely unknown. In this study, we detected monogalactosyl diacylglycerols (MGDGs) containing the 12/10-carbon HPL products in disrupted leaf tissues of Arabidopsis, cabbage, tobacco, tomato, and common bean. They were identified as an MGDG containing 12-oxo-9-hydroxy-(E)-10-dodecenoic acid and 10-oxo-7-hydroxy-(E)-8-decenoic acid and an MGDG containing two 12-oxo-9-hydroxy-(E)-10-dodecenoic acids as their acyl groups. Analyses of Arabidopsis mutants lacking HPL indicated that these MGDGs were formed enzymatically through an active HPL reaction. Thus, our results suggested that in disrupted leaf tissues, MGDG-hydroperoxides were cleaved by HPL to form volatile six-carbon aldehydes and non-volatile 12/10-carbon aldehyde-containing galactolipids. Based on these results, we propose a novel oxylipin pathway that does not require the lipase reaction to form GLVs.     

Production of phenolics and the emission of volatile organic compounds by perennial ryegrass (Lolium perenne L.)/Neotyphodium lolii association as a response to infection by Fusarium poae

Grasses very often form symbiotic associations with Neotyphodium/Epichloë endophytic fungi. These endophytes often allow the host grass to be protected from different pathogens. However, there is little known about the mechanisms of such endophyte influence on the host. Thus, the purpose of this research was to examine the effect of the N. lolii endophyte on the total production of phenolic compounds, VOCs emission and the resistance of three perennial ryegrass genotypes infected by pathogenic Fusarium poae. Analyses of total phenolics content were performed in control (not inoculated) and inoculated plants after 1, 2, 3, 4, 5, and 6 days (DAI) and for VOCs after 0, 3, 6 and 12 DAI. The presence of endophytes significantly reduced the disease index in two of the three genotypes relative to that in E−. Plants infected by N. lolii exhibited higher production of phenolics relative to the E− plants. The highest amounts of phenolics were observed on the second and sixth DAI. Genotype Nl22 showed the strongest effect of the endophyte on the production of phenolics, which increased by over 61%. Both the endophyte infected and non-infected plants emitted most abundantly two GLVs ((Z)-3-hexenal, (Z)-3-hexen-1-yl acetate), three terpenes (linalool, (Z)-ocimene, β-caryophyllene) and three shikimic acid pathway derivatives (benzyl acetate, indole, and methyl salicylate). The endophyte presence and the intervals of VOCs detection were a highly significant source of variation for all emitted volatiles (P < 0.001). The genotype of the perennial ryegrass significantly affected only the emission of methyl salicylate (P < 0.05) and β-caryophyllene (P < 0.05). Most of the VOCs ((Z)-3-hexen-1-yl acetate, (Z)-3-hexenal, linalool and methyl salicylate) reached their highest levels of emission on the sixth DAI, when averaged over genotypes and endophyte status. The results highlight the role of Neotyphodium spp. in the mediation of quadro-trophic interactions among plants, symbiotic endophytes, invertebrate herbivores and plant pathogenic fungi. Our results also confirm the fact that symbiotic plants can activate a defense reaction faster than non-symbiotic plants after a pathogen attack. Thus, N. lolii can be involved in the defense of perennial ryegrass against pathogens and potentially could be central to the host plants’ protection.