Papers by Rayko Halitschke
Plant Physiology, Apr 1, 2003
Plants respond to herbivore attack with a dramatic functional reorganization that involves the ac... more Plants respond to herbivore attack with a dramatic functional reorganization that involves the activation of direct and indirect defenses and tolerance, which in turn make large demands on primary metabolism. Here we provide the first characterization of the transcriptional reorganization that occurs after insect attack in a model plant-herbivore system: Nicotiana attenuata Torr. ex Wats.-Manduca sexta. We used mRNA differential display to characterize one-twentieth of the insect-responsive transcriptome of N. attenuata and verified differential expression for 27 cDNAs. Northern analyses were used to study the effects of folivory and exposure to airborne methyl jasmonate and for kinetic analyses throughout a 16-hlight/8-h-dark cycle. Sequence similarity searches allowed putative functions to be assigned to 15 transcripts. Genes were related to photosynthesis, electron transport, cytoskeleton, carbon and nitrogen metabolism, signaling, and a group responding to stress, wounding, or invasion of pathogens. Overall, transcripts involved in photosynthesis were strongly down-regulated, whereas those responding to stress, wounding, and pathogens and involved in shifting carbon and nitrogen to defense were strongly up-regulated. The majority of transcripts responded similarly to airborne methyl jasmonate and folivory, and had tissue-and diurnal-specific patterns of expression. Transcripts encoding Thr deaminase (TD) and a putative retrotransposon were absent in control plants, but were strongly induced after herbivory. Full-length sequences were obtained for TD and the pathogen-inducible ␣-dioxygenase, PIOX. Effects of abiotic and biotic stimuli were investigated for transcripts encoding TD, importin ␣, PIOX, and a GAL83-like kinase cofactor.
Molecular Breeding, 2014
Acylsugars are secondary metabolites with proven insect resistance properties that are produced b... more Acylsugars are secondary metabolites with proven insect resistance properties that are produced by many Solanaceous species including Solanum pennellii, a wild relative of tomato. The acylsugar chemotypes of S. pennellii varies greatly within its natural range and might be the product of plant/insect coevolution. The S. pennellii accession LA716 was used to transfer increased levels of acylsugar production into the cultivated tomato, resulting in the acylsugar tomato breeding line CU071026. S. pennellii accession LA716 produces high levels of acylsugars with chemotypes that differ greatly from those produced by CU071026 or the trace acylsugars of cultivated tomato. Understanding the genetic regulation of acylsugar chemistry will aid efforts to breed acylsugar production into cultivated tomato, allowing for alteration of both acylsugar base moieties and fatty acid profiles. This study uses a BC1F1 population produced from the cross of S. pennellii LA716 and CU071026 with CU071026 as the recurrent parent to identify QTL that change the fatty acid profile of acylsugars. Multiple QTL and epistatic interactions between QTL were detected including three QTL on chromosomes 2, 5, and 7, which significantly alter the percentage of extended iso-odd branched fatty acids and straight chain fatty acids on the acylsugars. We also report the introgression of one of these QTL, FA 2, into CU071026, resulting in a new tomato line with significantly increased i11:0 as a percentage of the fatty acids in its acylsugars. Candidate genes for these QTL and the impacts of altering acylsugar fatty acid are discussed.
Theoretical and Applied Genetics, 2009
The potato R locus is required for the production of red pelargonidin-based anthocyanin pigments ... more The potato R locus is required for the production of red pelargonidin-based anthocyanin pigments in potato (Solanum tuberosum L.). Red color also requires tissue-speciWc regulatory genes, such as D (for expression in tuber skin) and F (expression in Xowers). A related locus, P, is required for production of blue/purple anthocyanins; P is epistatic to R. We have previously reported that the dihydroXavonol 4-reductase gene (dfr) co-segregates with R. To test directly whether R corresponds to dfr, we placed the allele of dfr associated with red color under the control of the CaMV 35S promoter and introduced it into the potato cultivar Prince Hairy (genotype dddd rrrr P-), which has white tubers and pale blue Xowers. Transgenic Prince Hairy tubers remained white, but Xower color changed to purple. Three independent transgenic lines, as well as a vectortransformed line, were then crossed with the red-skinned variety Chieftain (genotype D-R-pppp), to establish populations that segregated for D, R, P, and the dfr transgene or empty vector. Markers were used to genotype progeny at D and R. Progeny carrying the empty vector in the genetic background D-rrrr produced white or purple tubers, while progeny with the same genotype and the dfr transgene produced red or purple tubers. HPLC and LC-MS/MS analyses of anthocyanins present in Chieftain and in a red-skinned progeny clone with the dfr transgene in a D-rrrr background revealed no qualitative diVerences. Thus, dfr can fully complement R, both in terms of tuber color and anthocyanin composition.
The Plant Journal, 2006
When attacked by herbivores, plants release volatile organic compounds (VOCs) that attract natura... more When attacked by herbivores, plants release volatile organic compounds (VOCs) that attract natural enemies of the herbivores and function as indirect defenses. Whether or not neighboring plants 'eavesdrop' on these VOCs remains controversial because most studies use unrealistic experimental conditions and VOC exposures. In order to manipulate exposures of wild-type (WT) Nicotiana attenuata'receiver' plants, we elicited transformed 'emitter' plants, whose production of herbivore-induced C6 green leaf volatiles (GLVs) or terpenoid volatiles was genetically silenced, and placed them up-wind of WT 'receiver' plants in open-flow experimental chambers. We compared the transcriptional and secondary metabolite defense responses of WT receiver plants exposed to VOCs from these transgenic emitter plants with those of plants exposed to VOCs from WT emitter plants. No differences in the constitutive accumulation of defense metabolites and the signal molecule jasmonic acid (JA) were found. Additional elicitation of receiver plants revealed that exposure to WT, GLV-deficient and terpenoid-deficient volatile blends did not prime induced defenses, JA accumulation, or the expression of lipoxygenase 3 (NaLOX3), a gene involved in JA biosynthesis. However, exposure to wound- and herbivore-induced VOCs significantly altered the transcriptional patterns in receiver plants. We identified GLV-dependent genes by complementing the GLV-deficient volatile blend with a mixture of synthetic GLVs. Blends deficient in GLVs or cis-alpha-bergamotene regulated numerous genes in receiver plants that did not respond to the complete VOC blends of WT emitters, indicating a suppressive effect of GLVs and terpenoids. Whether these transcriptional responses translate into changes in plant fitness in nature remains to be determined.
The Plant Journal, 2007
Arabidopsis and tomato plants mutated in the F-box protein COI1 mediating jasmonate (JA) response... more Arabidopsis and tomato plants mutated in the F-box protein COI1 mediating jasmonate (JA) responses are more susceptible to herbivores in laboratory trials, but the exact mechanisms of COI1-mediated resistance are not known. We silenced COI1 by transformation with an inverted repeat construct (ir-coi1) in Nicotiana attenuata, a plant the direct and indirect defenses of which against various herbivores have been well studied. ir-coi1 plants are male sterile and impaired in JA-elicited direct [nicotine, caffeoylputrescine and trypsin proteinase inhibitor (TPI) activity] and indirect (cis-alpha-bergamotene emission) defense responses; responses not elicited by JA treatment (ethylene production and flower TPI activity) were unaffected. Larvae of Manduca sexta, a common herbivore of N. attenuata, gained three times more mass feeding on ir-coi1 than on wild-type (WT) plants in glasshouse experiments. By regularly moving caterpillars to unattacked leaves of the same plant, we demonstrate that larvae on WT plants can grow and consume leaves as fast as those on ir-coi1 plants, a result that underscores the role of COI1 in mediating locally induced resistance in attacked leaves, and the importance of herbivore movement in avoiding the induced defenses of a plant. When transplanted into native habitats in the Great Basin Desert, ir-coi1 plants suffer greatly from damage by the local herbivore community, which includes herbivores not commonly found on N. attenuata WT plants. Choice assays with field-grown plants confirmed the increased attractiveness of ir-coi1 plants for both common and unusual herbivores. We conclude that NaCOI1 is essential for induced resistance in N. attenuata, and that ir-coi1 plants highlight the benefits of herbivore movement for avoiding induced defenses.
The Plant Journal, 2014
The fitness consequences of mating system variation (e.g. inbreeding) have been studied for at le... more The fitness consequences of mating system variation (e.g. inbreeding) have been studied for at least 200 years, yet the ecological consequences of this variation remain poorly understood. Most plants are capable of inbreeding, and also exhibit a remarkable suite of adaptive phenotypic responses to ecological stresses such as herbivory. We tested the consequences of experimental inbreeding on phenotypic plasticity in resistance and growth (tolerance) traits in Solanum carolinense (Solanaceae). Inbreeding reduced the ability of plants to up-regulate resistance traits following damage. Moreover, inbreeding disrupted growth trait responses to damage, indicating the presence of deleterious mutations at loci regulating growth under stress. Production of the phytohormones abscisic and indole acetic acid, and wounding-induced up-regulation of the defence signalling phytohormone jasmonic acid were all significantly reduced under inbreeding, indicating a phytohormonal basis for inbreeding effects on growth and defence trait regulation. We conclude that the plasticity of induced responses is negatively affected by inbreeding, with implications for fragmented populations facing mate limitation and stress as a consequence of environmental change.
The Plant Journal, 2003
Inhibition of jasmonic acid (JA) signaling has been shown to decrease herbivore resistance, but t... more 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.
PLoS Biology, 2004
Plants produce metabolites that directly decrease herbivore performance, and as a consequence, he... more Plants produce metabolites that directly decrease herbivore performance, and as a consequence, herbivores are selected for resistance to these metabolites. To determine whether these metabolites actually function as defenses requires measuring the performance of plants that are altered only in the production of a certain metabolite. To date, the defensive value of most plant resistance traits has not been demonstrated in nature. We transformed native tobacco (Nicotiana attenuata) with a consensus fragment of its two putrescine N-methyl transferase (pmt) genes in either antisense or inverted-repeat (IRpmt) orientations. Only the latter reduced (by greater than 95%) constitutive and inducible nicotine. With D 4-nicotinic acid (NA), we demonstrate that silencing pmt inhibits nicotine production, while the excess NA dimerizes to form anatabine. Larvae of the nicotine-adapted herbivore Manduca sexta (tobacco hornworm) grew faster and, like the beetle Diabrotica undecimpunctata, preferred IRpmt plants in choice tests. When planted in their native habitat, IRpmt plants were attacked more frequently and, compared to wild-type plants, lost 3fold more leaf area from a variety of native herbivores, of which the beet armyworm, Spodoptera exigua, and Trimerotropis spp. grasshoppers caused the most damage. These results provide strong evidence that nicotine functions as an efficient defense in nature and highlights the value of transgenic techniques for ecological research.
Planta, 2013
Plant protease inhibitors (PIs) are among the most well-studied and widely distributed resistance... more Plant protease inhibitors (PIs) are among the most well-studied and widely distributed resistance traits that plants use against their herbivore attackers. There are different types of plant PIs which putatively function against the different types of proteases expressed in insect guts. Serine protease inhibitors (SPIs) and cysteine protease inhibitors (CPIs) are hypothesized to differentially function against the predominant gut proteases in lepidopteran and coleopteran herbivores, respectively. Here, we test the hypothesis that tall goldenrod, Solidago altissima, can specifically respond to damage by different herbivores and differentially induce SPIs and CPIs in response to damage by lepidopteran and coleopteran herbivores. Moreover, we ask if the concerted induction of different types of PIs accounts for variation in induced resistance to herbivory. We altered and optimized a rapid and effective existing methodology to quantitatively analyze both SPI and CPI activity simultaneously from a single tissue sample and to use the same plant extracts directly for characterization of inhibitory effects on insect gut protease activity. We found that both SPIs and CPIs are induced in S. altissima in response to damage, regardless of the damaging herbivore species. However, only SPIs were effective against Spodoptera exigua gut proteases. Our data suggest that plant PI responses are not necessarily specific to the identity of the attacking organism but that different components of generally induced defense traits can specifically affect different herbivore species. While providing an efficient and broadly applicable methodology to analyze multiple PIs extracted from the same tissue, this study furthers our understanding of specificity in induced plant resistance.
Oecologia, 2006
Plants release volatile organic compounds (VOCs) in response to wounding and herbivore attack, so... more Plants release volatile organic compounds (VOCs) in response to wounding and herbivore attack, some of which trigger responses in neighboring unattacked plants in the laboratory under conditions that are not likely to occur in the real world. Whether plants 'eavesdrop' on the volatile emissions of their neighbors in nature is not known. The best documented field study of between-species signaling via above-ground VOCs involves increases in fitness parameters of native tobacco (Nicotiana attenuata) transplanted adjacent to clipped sagebrush (Artemesia tridentata tridentata). Clipped sagebrush releases many biologically active VOCs, including methyl jasmonate (MeJA), methacrolein and a series of terpenoid and green leaf VOCs, of which MeJA, while active under laboratory conditions, is not released in sufficient quantities to directly elicit induced resistance in the field. Here we demonstrate, with laboratory and field-based experiments, that priming (rather than direct elicitation) of native N. attenuata's induced chemical defenses by a sagebrush-released VOC bouquet can account for earlier findings. With microarrays enriched in N. attenuata herbivore-regulated genes, we found transcriptional responses in tobacco growing adjacent to clipped sagebrush foliage, but failed to detect the direct elicitation of defensive chemicals or proteins. However, we observed an accelerated production of trypsin proteinase inhibitors when Manduca sexta caterpillars fed on plants previously exposed to clipped sagebrush. This readying of a defense response, termed priming, results in lower total herbivore damage to plants exposed to clipped sagebrush and in a higher mortality rate of young Manduca caterpillars. Our study demonstrates priming of plant defense responses as a mechanism of plant-plant signaling in nature, and provides an example for the analysis of between-plant signaling under ecologically realistic conditions. Although we describe priming as a potential mechanism for signaling between plants in nature, we critically discuss the ecological relevance of the particular interaction.
New Phytologist, 2011
• Herbivory is thought to be detrimental to plant fitness and commonly results in a metabolic shi... more • Herbivory is thought to be detrimental to plant fitness and commonly results in a metabolic shift in the plant: photosynthetic processes are typically downregulated, while resource allocation to defenses is increased in herbivore-attacked plants, resulting in fitness costs of induced plant responses. • Wild tobacco, Nicotiana attenuata, attacked by Tupiocoris notatus mirid bugs becomes resistant against more damaging herbivores through mirid-induced direct and indirect defenses. However, mirid-induced resistance and tissue loss do not result in a reduction of plant fitness. These findings suggest induced metabolic responses allowing the plant to compensate for the lost tissue and resources allocated to defenses. • While feeding by Manduca sexta larvae results in a strong down-regulation of photosynthesis, we demonstrate a specific induction of elevated photosynthetic activity in N. attenuata leaves by elicitors in mirid salivary secretions. The elevated CO 2 assimilation rate is sufficient to compensate for the loss of photosynthetically active tissue and balances the net photosynthesis of infested leaves. • We discuss the observed increase in the plant's primary metabolic activity as a mechanism that allows plants to alleviate negative fitness effects of mirid attack and mediates the vaccination effects that result in a net benefit in environments with multiple herbivores.
Molecular Ecology, 2004
Lepidopteran larvae oral secretions and regurgitant (R), which contain a plethora of potential el... more Lepidopteran larvae oral secretions and regurgitant (R), which contain a plethora of potential elicitors, are known to dramatically change a plant's wound response. We demonstrate, with a detailed microarray and secondary metabolite analysis, that the two most abundant fatty acid-amino acid conjugates (FACs) in the R of the specialist herbivore Manduca sexta (Lepidoptera, Sphingidae) can account for all measured direct (trypsin proteinase inhibitor: TPI) and indirect (cis-α α α α-bergamotene) defences, the endogenous jasmonic acid burst that elicits them, and 86% of the induced transcriptional changes (89% up and 83% down) in its native host Nicotiana attenuata and hence are necessary and sufficient for the Manducaspecific modulation of the wound response. FACs were not found in eggs, but detected in larvae of all instars after their first meal. FACs were found in all regions of the alimentary canal and in the frass, but did not occur in salivary or mandibular glands, extracts of which were not active in any assay. Individual larvae differed substantially in their FAC composition and two FAC chemotypes were discernible: N-linolenoyl-L-glutamine biased R and N-linolenoyl-L-glutamate biased R. We created synthetic blends of FACs to mimic these chemical phenotypes and determined whether plants respond differently to the different R chemotypes. Micorarray and TPI analysis revealed that plants do not differentiate. N. attenuata plants use FACs from feeding caterpillars to tailor their wound responses but do not use the variability in FAC ratios to recognize attack from an individual caterpillar.
Journal of Plant Growth Regulation, 2004
Herbivore attack elicits defense responses in host plants by a complex chain of events that start... more Herbivore attack elicits defense responses in host plants by a complex chain of events that starts with the introduction of herbivore-specific elicitors into the wounds at the feeding or oviposition site, their recognition by the plant, and activation of several signaling cascades that trigger defense responses that finally increase resistance. Oxylipin signaling plays a central role in the activation of these herbiw)re-induced responses. Wounding activates some but not all of these defense responses, but herbiw~re attack frequently amplifies the oxylipin responses well beyond that elicited by wounding alone, suggesting recognition of herbivore attack. In addition to their signaling role within the plant, oxylipins can also directly influence the performance of herbivores or attract natural enemies to feeding herbiw)res. Here we review the literature on the regulation and function of herbiw)re-specific oxylipin signaling and the direct effects of oxylipins on herbiw)re performance.
Electrochemistry Communications, 2010
Here, we show that quorum sensing (QS) modulates the current generation of the anode-respiring ba... more Here, we show that quorum sensing (QS) modulates the current generation of the anode-respiring bacterium Pseudomonas aeruginosa because it controls the production of phenazines, which mediate the electron transfer to the anode. The current generation by a wildtype (WT) strain P. aeruginosa PA14 and the GacS/GacA protein-regulatory mutant retS was investigated under different environmental conditions. The retS mutant generated significantly higher current (45-fold) than the WT under anaerobic conditions. Anaerobic current generation by the WT was 28-fold higher with extraneously supplied lactones (a QS-signaling molecule). Compared to anaerobic conditions, the WT with some oxygen (microaerobic conditions) exhibited enhanced phenazine production (39-fold) and current levels (48-fold). Ironrich medium and microaerobic conditions had a negative impact on current generation by retS. All these results were directly linked to QS activity in P. aeruginosa, thus, demonstrating the importance of this bacterial communication system for current generation in BESs. We also show that BESs represent a new tool for real-time investigation of phenazine-related QS activity.
Ecology Letters, 2007
The attraction of natural enemies of herbivores by volatile organic compounds as an induced indir... more The attraction of natural enemies of herbivores by volatile organic compounds as an induced indirect defence has been studied in several plant systems. The evidence for their defensive function originates mainly from laboratory studies with trained parasitoids and predators; the defensive function of these emissions for plants in natural settings has been rarely demonstrated. In native populations and laboratory Y-tube choice experiments with transgenic Nicotiana attenuata plants unable to release particular volatiles, we demonstrate that predatory bugs use terpenoids and green leaf volatiles (GLVs) to locate their prey on herbivore-attacked plants. By attracting predators with volatile signals, this native plant reduces its herbivore loaddemonstrating the defensive function of herbivore-induced volatile emissions. However, plants producing GLVs are also damaged more by flea beetles. The implications of these conflicting ecological effects for the evolution of induced volatile emissions and for the development of sustainable agricultural practices are discussed.
Ecology, 2010
Plants employ hormone-mediated signaling pathways to defend against pathogens and insects. We tes... more Plants employ hormone-mediated signaling pathways to defend against pathogens and insects. We tested predictions about the relative effect of jasmonate and salicylate pathways and how they mediate interactions between pathogens and herbivores. We employed two pathogens of tomato, Pseudomonas syringae (Pst) and tobacco mosaic virus (TMV), that are known to elicit distinct components of the two pathways, and we address the consequences of their induction for resistance in wild-type and salicylate-deficient transgenic plants in field experiments. We report that Pst infection induced jasmonic acid and proteinase inhibitors (PIs), and reduced the growth of Spodoptera exigua caterpillars on wild-type and salicylate-deficient plants. Pst and TMV both induced salicylic acid in wild-type but not salicylate-deficient plants. Although TMV did not affect jasmonic acid or PIs, infection increased caterpillar growth on wild-type plants, but not on salicylate-deficient plants. Aphid population growth was higher on salicylate-deficient compared to wild-type plants, and lower on salicylate-induced plants compared to controls. Natural aphid colonization was reduced on TMV-infected wild types, but not on salicylate-deficient plants. In sum, jasmonate-mediated resistance is induced by some pathogens, independent of salicylate, and salicylate-mediated induction by other pathogens results in induced susceptibility to a chewer and resistance to an aphid. We conclude with a predictive model for the expression of defense pathways and their consequences.
Ecology, 2013
Early-season herbivory can cause plants to induce resistance to subsequent herbivores, and this u... more Early-season herbivory can cause plants to induce resistance to subsequent herbivores, and this ubiquitous plant defense strategy has been shown to be adaptive when subsequent vegetative-feeding herbivores impact plant fitness. However, a growing number of studies show that leaf herbivory can also induce defenses in plant reproductive tissues, which may deter mutualists such as pollinators and seed dispersers, or antagonists such as florivores and seed predators. Due to their direct interaction with plant reproductive tissues, deterrence of these mutualists or antagonists may have large negative or positive effects on plant fitness. Despite numerous predictions, we know little regarding the fitness consequences of leaf-toreproductive tissue induction in nature. Here, using observations of natural populations and manipulative field experiments, we show that leaf herbivory by the invasive Japanese beetle (Popillia japonica) induces an accumulation of jasmonic acid and complex phenolics in reproductive tissues of the common evening primrose (Oenothera biennis). Consequently, seed predation by three native Lepidoptera is reduced by 77% on beetle-induced plants, while leaf herbivory itself has little impact on lifetime reproductive output. These results show that O. biennis benefits from being consumed by an invasive folivore, and that leaf-to-reproductive tissue induction can be an adaptive trait. Induced plant defenses can therefore be beneficial in a broader community context than previously realized.
Current Opinion in Plant Biology, 2001
Current Opinion in Plant Biology, 2002
Plants release volatiles after herbivore attack in a highly regulated fashion. These compounds at... more Plants release volatiles after herbivore attack in a highly regulated fashion. These compounds attract natural enemies and function as indirect defenses. Whether neighboring plants 'eavesdrop' on these volatile signals and tailor their defenses accordingly remains controversial. Recent laboratory studies have identified transcriptional changes that occur in plants in response to certain volatiles. These changes occur under conditions that enhance the probability of signal perception and response. Field studies have demonstrated repeatable increases in the herbivore resistance of plants growing downwind of damaged plants.
Chemoecology, 2010
Abstract Solidago altissima is a dominant perennial of old field succession in North America with... more Abstract Solidago altissima is a dominant perennial of old field succession in North America with the ability to form dense, almost monospecific stands; the plant is also considered an aggressive invasive throughout Europe and Japan. S. altissima's success is facilitated by large ...
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Papers by Rayko Halitschke