Nicole M. van Dam

Rewiring of the Jasmonate Signaling Pathway in Arabidopsis during Insect Herbivory

Plant defenses against insect herbivores and necrotrophic pathogens are differentially regulated by different branches of the jasmonic acid (JA) signaling pathway. In Arabidopsis, the basic helix-loop-helix leucine zipper transcription factor (TF) MYC2 and the APETALA2/ETHYLENE RESPONSE FACTOR (AP2/ERF) domain TF ORA59 antagonistically control these distinct branches of the JA pathway. Feeding by larvae of the specialist insect herbivore Pieris rapae activated MYC2 transcription and stimulated expression of the MYC2-branch marker gene VSP2, while it suppressed transcription of ORA59 and the ERF-branch marker gene PDF1.2. Mutant jin1 and jar1-1 plants, which are impaired in the MYC2-branch of the JA pathway, displayed a strongly enhanced expression of both ORA59 and PDF1.2 upon herbivory, indicating that in wild-type plants the MYC2-branch is prioritized over the ERF-branch during insect feeding. Weight gain of P. rapae larvae in a no-choice setup was not significantly affected, but in a two-choice setup the larvae consistently preferred jin1 and jar1-1 plants, in which the ERF-branch was activated, over wild-type Col-0 plants, in which the MYC2-branch was induced. In MYC2- and ORA59-impaired jin1-1/RNAi-ORA59 plants this preference was lost, while in ORA59-overexpressing 35S:ORA59 plants it was gained, suggesting that the herbivores were stimulated to feed from plants that expressed the ERF-branch rather than that they were deterred by plants that expressed the MYC2-branch. The feeding preference of the P. rapae larvae could not be linked to changes in glucosinolate levels. Interestingly, application of larval oral secretion into wounded leaf tissue stimulated the ERF-branch of the JA pathway, suggesting that compounds in the oral secretion have the potential to manipulate the plant response toward the caterpillar-preferred ERF-regulated branch of the JA response. Our results suggest that by activating the MYC2-branch of the JA pathway, plants prevent stimulation of the ERF-branch by the herbivore, thereby becoming less attractive to the attacker.

The impact of the absence of aliphatic glucosinolates on insect herbivory in Arabidopsis

Aliphatic glucosinolates are compounds which occur in high concentrations in Arabidopsis thaliana and other Brassicaceae species. They are important for the resistance of the plant to pest insects. Previously, the biosynthesis of these compounds was shown to be regulated by transcription factors MYB28 and MYB29. We now show that MYB28 and MYB29 are partially redundant, but in the absence of both, the synthesis of all aliphatic glucosinolates is blocked. Untargeted and targeted biochemical analyses of leaf metabolites showed that differences between single and double knock-out mutants and wild type plants were restricted to glucosinolates. Biosynthesis of long-chain aliphatic glucosinolates was blocked by the myb28 mutation, while short-chain aliphatic glucosinolates were reduced by about 50% in both the myb28 and the myb29 single mutants. Most remarkably, all aliphatic glucosinolates were completely absent in the double mutant. Expression of glucosinolate biosynthetic genes was slightly but significantly reduced by the single myb mutations, while the double mutation resulted in a drastic decrease in expression of these genes. Since the myb28myb29 double mutant is the first Arabidopsis genotype without any aliphatic glucosinolates, we used it to establish the relevance of aliphatic glucosinolate biosynthesis to herbivory by larvae of the lepidopteran insect Mamestra brassicae. Plant damage correlated inversely to the levels of aliphatic glucosinolates observed in those plants: Larval weight gain was 2.6 fold higher on the double myb28myb29 mutant completely lacking aliphatic glucosinolates and 1.8 higher on the single mutants with intermediate levels of aliphatic glucosinolates compared to wild type plants.

Interactions between aboveground and belowground induced responses against phytophages

Abstract Since their discovery about thirty years ago, induced plant responses have mainly been studied in interactions of plants with aboveground (AG) pathogens, herbivores and their natural enemies. Many induced responses, however, are known to be systemic and thus it is likely that responses induced by AG phytophages affect belowground (BG) phytophages feeding on the same plant, and vice versa . The awareness that interactions between AG and BG phytophages may be an important aspect in the evolution of induced responses came only recently and little research has been done to date. In this review we first summarise ecological studies that show how AG phytophages may affect BG phytophages, and vice versa . Then we focus on mechanisms governing interactions between AG and BG induced responses, such as cross-talk between signals. We chose the genus Nicotiana and the family Brassicaceae as two examples of plant groups that have been well studied for their induced responses both AG and BG – but not in concert – and explore how interactions between AG and BG induced compounds may link multitrophic interactions associated with these plants. We propose that future research on AG and BG interactions should focus on: 1). Identification of compounds and signalling pathways involved in AG and BG induced responses and analysis of their interaction mechanisms, 2). Evaluation of how induced responses affect interactions between BG and AG phytophages and their natural enemies, 3). Evaluation of the effects of AG and BG phytophages -in combination with their natural enemies- on plant fitness to identify keystone interactions that are driving the natural selection for induced responses in plants. Seit ihrer Entdeckung vor ca. dreisig Jahren werden induzierte pflanzliche Antworten der Pflanzen zumeist mit solchen Pathogenen, Herbivoren und deren naturlichen Feinden untersucht, die an oberirdischen Pflanzenteilen zu finden sind. Viele induzierte Antworten der Pflanzen konnen aber systemisch sein. Daher ist es wahrscheinlich, dass pflanzliche Antworten, die durch oberirdische Organismen induziert werden, auch solche Phytophagen beeinflussen, die unterirdisch an der Pflanze fressen, und umgekehrt. Das Bewustsein darum, dass Interaktionen zwischen ober- und unterirdischen Phytophagen ein wichtiger Aspekt in der Evolution von induzierten pflanzlichen Abwehrreaktionen sein konnen, kam erst in neuerer Zeit auf. Deshalb gibt es bisher wenig Forschung auf diesem Gebiet. In diesem Uberblick werden zunachst die Studien zusammengefasst, die den Einfluss oberirdischer Phytophager auf die unterirdischen Phytophagen zeigen und umgekehrt. Weiterhin wird auf die Mechanismen fokussiert, die Interaktionen zwischen ober- und unterirdisch fressenden Phytophagen steuern, wie z.B. Wechselwirkungen zwischen Signalen. Die Gattung Nicotiana und die Familie Brassicaceae werden als Modellpflanzen ausgewahlt, an denen die induzierte Abwehr gegen entweder oberirdische oder unterirdische Phytophage – aber nicht gegen beide gleichzeitig – bereits gut untersucht wurde. Es wird analysiert, wie Interaktionen zwischen ober- und unterirdisch induzierten Verbindungen mit multitrophischen Interaktionen dieser Pflanzen in Zusammenhang stehen. Es werden Vorschlage unterbreitet, worauf zukunftige Forschung an ober- und unterirdisch induzierten Interaktionen fokussieren sollte: (1) Identifizierung der Verbindungen und der Signalwege, die bei ober- und unterirdischer Induktion von Bedeutung sind und Analyse der Interaktionsmechanismen, (2) Untersuchung der Frage, wie induzierte Reaktionen der Pflanze sich auf Interaktionen zwischen ober- und unterirdischen Phytophagen und deren naturlichen Feinden auswirken, (3) Analyse der Auswirkungen der Effekte von ober- und unterirdisch fressenden Phytophagen unter Berucksichtigung auch ihrer naturlichen Feinde auf die pflanzliche Fitness, um solche Schlusselinteraktionen zu identifizieren, die entscheidend sind fur die Selektion der induzierten pflanzlichen Reaktion auf Angriffe durch Phytophage.

GENETIC VARIATION IN DEFENSE CHEMISTRY IN WILD CABBAGES AFFECTS HERBIVORES AND THEIR ENDOPARASITOIDS

Populations of wild Brassica oleracea L. grow naturally along the Atlantic coastlines of the United Kingdom and France. Over a very small spatial scale (i.e., <15 km) these populations differ in the expression of the defensive compounds, glucosinolates (GS). Thus far, very few studies have examined interactions between genetically distinct populations of a wild plant species and associated consumers in a multitrophic framework. Here, we compared the development of a specialist (Pieris rapae) and a generalist (Mamestra brassicae) insect herbivore and their endoparasitoids (Cotesia rubecula and Microplitis mediator, respectively) on three wild populations and one cultivar of B. oleracea under controlled greenhouse conditions. Herbivore performance was differentially affected by the plant population on which they were reared. Plant population influenced only development time and pupal mass in P. rapae, whereas plant population also had a dramatic effect on survival of M. brassicae. Prolonged development time in P. rapae corresponded with high levels of the indole GS, neoglucobrassicin, whereas reduced survival in M. brassicae coincided with high levels of the aliphatic GS, gluconapin and sinigrin. The difference between the two species can be explained by the fact that the specialist P. rapae is adapted to feed on plants containing GS and has evolved an effective detoxification system against aliphatic GS. The different B. oleracea populations also affected development of the endoparasitoids. Differences in food-plant quality for the hosts were reflected in adult size in C. rubecula and survival in M. mediator, and further showed that parasitoid performance is also affected by herbivore diet.

Performance of generalist and specialist herbivores and their endoparasitoids differs on cultivated and wild Brassica populations.

Through artificial selection, domesticated plants often contain modified levels of primary and secondary metabolites compared to their wild progenitors. It is hypothesized that the changed chemistry of cultivated plants will affect the performance of insects associated with these plants. In this paper, the development of several specialist and generalist herbivores and their endoparasitoids were compared when reared on a wild and cultivated population of cabbage, Brassica oleracea, and a recently established feral Brassica species. Irrespective of insect species or the degree of dietary specialization, herbivores and parasitoids developed most poorly on the wild population. For the specialists, plant population influenced only development time and adult body mass, whereas for the generalists, plant populations also affected egg-to-adult survival. Two parasitoid species, a generalist (Diadegma fenestrale) and a specialist (D. semiclausum), were reared from the same host (Plutella xylostella). Performance of D. semiclausum was closely linked to that of its host, whereas the correlation between survival of D. fenestrale and host performance was less clear. Plants in the Brassicaceae characteristically produce defense-related glucosinolates (GS). Levels of GS in leaves of undamaged plants were significantly higher in plants from the wild population than from the domesticated populations. Moreover, total GS concentrations increased significantly in wild plants after herbivory, but not in domesticated or feral plants. The results of this study reveal that a cabbage cultivar and plants from a wild cabbage population exhibit significant differences in quality in terms of their effects on the growth and development of insect herbivores and their natural enemies. Although cultivated plants have proved to be model systems in agroecology, we argue that some caution should be applied to evolutionary explanations derived from studies on domesticated plants, unless some knowledge exists on the history of the system under investigation.

The “Raison D'être” of pyrrolizidine alkaloids inCynoglossum officinale: Deterrent effects against generalist herbivores

In this study we tested whether pyrrolizidine alkaloids (PAs) ofCynoglossum officinale serve as antifeedants against herbivores. Total PA N-oxide extracts of the leaves significantly deterred feeding by generalist herbivores. Specialist herbivores did not discriminate between food with high and low PA levels. Three PAs fromC. officinale, heliosupine, echinatine, and 3′-acetylechinatine, equally deterred feeding by the polyphagous larvae ofSpodoptera exigua. Although the plants mainly contain PAs in their N-oxide form, reduced PAs deterred feeding byS. exigua more efficiently than PA N-oxides. On rosette plants, the monophagous weevilMogulones cruciger significantly consumed more of the youngest leaves, which had the highest PA level and the highest nitrogen percentage. Larvae ofEthmia bipunctella, which are oligophagous within the Boraginaceae, did not discriminate between leaves. All generalist herbivores tested significantly avoided the youngest leaves with the highest PA levels. In the field, the oldest leaves also were relatively more damaged by herbivores than the youngest leaves. It is hypothesized that the skewed distribution of PAs over the leaves of rosette plants reflects optimal defense distribution within the plant.

Root herbivory reduces growth and survival of the shoot feeding specialist Pieris rapae on Brassica nigra

Plants may respond to herbivore attacks by changing their chemical profile. Such induced responses occur both locally and systemically throughout the plant. In this paper we studied how Brassica nigra (L.) Koch (Brassicaceae) plants respond to two different root feeders, the endoparasitic nematode Pratylenchus penetrans Cobb (Tylenchida: Pratylenchidae) and the larvae of the cabbage root fly Delia radicum L. (Diptera: Anthomyiidae). We tested whether the activities of the root feeders affected the survival and development of the shoot feeding crucifer specialist Pieris rapae (L.) (Lepidoptera: Pieridae) via systemically induced changes in the shoots. Overall, P. rapae larvae grew slower and produced fewer pupae on plants that were infested with root feeders, especially on plants infested with P. penetrans. This effect could not be attributed to lower water or protein levels in these plants, as the percentage of water in the controls and root infested shoots was similar, and protein content was even higher in root infested plants. Both glucosinolate as well as phenolic levels were affected by root feeding. Initially, glucosinolate levels were the lowest in root infested plants, but on P. penetrans infested plants they increased more rapidly after P. rapae started feeding than in controls or D. radicum infested plants. Plants with D. radicum feeding on their roots had the highest phenolic levels at all harvest dates. Our results indicate that root feeding can significantly alter the nutritional quality of shoots by changes in secondary metabolite levels and hence the performance of a specialist shoot feeder.

Root and shoot glucosinolates: a comparison of their diversity, function and interactions in natural and managed ecosystems

The role of glucosinolates in aboveground plant–insect and plant–pathogen interactions has been studied widely in both natural and managed ecosystems. Fewer studies have considered interactions between root glucosinolates and soil organisms. Similarly, data comparing local and systemic changes in glucosinolate levels after root- and shoot-induction are scarce. An analysis of 74 studies on constitutive root and shoot glucosinolates of 29 plant species showed that overall, roots have higher concentrations and a greater diversity of glucosinolates than shoots. Roots have significantly higher levels of the aromatic 2-phenylethyl glucosinolate, possibly related to the greater effectiveness and toxicity of its hydrolysis products in soil. In shoots, the most dominant indole glucosinolate is indol-3-ylglucosinolate, whereas roots are dominated by its methoxyderivatives. Indole glucosinolates were the most responsive after jasmonate or salicylate induction, but increases after jasmonate induction were most pronounced in the shoot. In general, root glucosinolate levels did not change as strongly as shoot levels. We postulate that roots may rely more on high constitutive levels of glucosinolates, due to the higher and constant pathogen pressure in soil communities. The differences in root and shoot glucosinolate patterns are further discussed in relation to the molecular regulation of glucosinolate biosynthesis, the within-tissue distribution of glucosinolates in the roots, and the use of glucosinolate-containing crops for biofumigation. Comparative studies of tissue-specific biosynthesis and regulation in relation to the biological interactions in aboveground and belowground environments are needed to advance investigations of the evolution and further utilization of glucosinolates in natural and managed ecosystems.

Chemical diversity in Brassica oleracea affects biodiversity of insect herbivores.

Intraspecific variation in plants plays a major role in the composition and diversity of the associated insect community. Resistance traits of plants are likely candidates mediating community composition. However, it is debated whether total concentrations of chemical compounds or specific compounds determine herbivore resistance, and how chemical diversity among plant genotypes in turn affects the composition of the associated herbivore community. To study the role of specific chemical compounds in affecting the herbivore community, we used cultivated Brassica oleracea. The cultivars differ qualitatively in glucosinolate profile, i.e., foliar composition of different glucosinolate compounds, and only a little in total concentration of glucosinolates, the secondary metabolites specific for the Brassicaceae family. In field and laboratory experiments, we tested whether individual compounds explained differences in herbivore community composition, and whether herbivores with a similar degree of host plant specialization responded in a similar way to variation in glucosinolate profiles. In the field B. oleracea cultivars differed widely in species richness and composition of the herbivore community, as well as in the density of insects they harbored. Plants with high concentrations of the short side chain alkenyl glucosinolate, glucoiberin, harbored low herbivore diversity. Higher biodiversity was found when plants had glucosinolate profiles containing high concentrations of glucosinolates with elongated side chains, which are biosynthetically linked to glucoiberin. Although glucosinolates are known to have differential effects on generalist and specialist herbivores, all herbivore species exhibited similar responses to the intraspecific variation in foliar glucosinolate profiles of the B. oleracea cultivars. This observation is supported by the correspondence between oviposition preferences of the specialist herbivore Pieris rapae and the generalist Mamestra brassicae in the field and the laboratory, using the same cultivars, and may be due to the relatively low concentrations of glucosinolates in cultivars. Our results show that variation in the concentration of short side-chain glucosinolates affects the composition of the herbivore community associated with brassicaceous plants.

Metabolomics in the Rhizosphere: Tapping into Belowground Chemical Communication

The rhizosphere is densely populated with a variety of organisms. Interactions between roots and rhizosphere community members are mostly achieved via chemical communication. Root exudates contain an array of primary and secondary plant metabolites that can attract, deter, or kill belowground insect herbivores, nematodes, and microbes, and inhibit competing plants. Metabolomics of root exudates can potentially help us to better understand this chemical dialogue. The main limitations are the proper sampling of the exudate, the sensitivity of the metabolomics platforms, and the multivariate data analysis to identify causal relations. Novel technologies may help to generate a spatially explicit metabolome of the root and its exudates at a scale that is relevant for the rhizosphere community.