Progress 07/01/04 to 12/31/07
Outputs
OUTPUTS: The goals of this project are to determine whether plants can discriminate among different pest species when initiating defense responses, how responses are coordinated and supplied with resources, and whether any of this matters to plant success. The last year of this project was funded by a regular NSF grant and the following studies were pursued. 1. We continued to analyze data from two large microarray studies. This involved annotation from the primary literature of thousands of genes, bioinformatic analysis of their upstream promoter regions, comparisons of hormone-responsive genes with other studies to identify participation of signaling pathways, and qRT-PCR analysis of transcription factors differentially expressed in response to insects. 2. A study of caterpillar feeding preferences on Arabidopsis leaf-age cohorts was completed and formed the basis of a Masters thesis and undergraduate Honors Thesis. A specialist species was found to feed preferentially on young
leaves with greater defense capacity while a generalist fed primarily on low-defense older leaves. Because these two caterpillar species alter the abundance of differentially-defended leaves and elicit different defense responses, they influence each other's food availability and feeding behavior, which has community ecology implications. Profiling glucosinolates in individual leaves of different ages in response to feeding by each species confirmed that both insect species are capable of inducing glucosinolates, but because of their preferences for different leaf ages they can appear to differ in their ability to induce based a whole plant profile of glucosinolates. 3. Work continued on source-sink control of defense responses in poplar and arabidopsis. Our previous work on poplar had demonstrated that induction of carbon-based defenses (tannins) in young leaves in response to insect feeding (or its surrogate) depended on phloem transport of carbon from older leaves. This year we
demonstrated that delivery of nitrogen to damaged leaves was not similarly enhanced, and we are in the process of measuring levels of nitrogen based defenses. 4. Studies of the response of plants to green leaf volatiles produced in response to insect feeding werecontinued in poplar and Arabidopsis. In poplar, we documented a new phenomenon of 'self-priming' of carbon based defenses by volatiles released in response to gypsy moth feeding. In Arabidopsis, we documented priming of glucosinolate defenses by major components of green leaf volatiles, resulting in faster glucosinolate induction by subsequent insect attack.Invited presentations on these results have been given at national and international meetings, including a Gordon Research Conference.
PARTICIPANTS: 1) Arabidopsis Microarray Project: Collaborators are H. M. Appel, J. C. Schultz, and R. R. Raina (Syracuse University), S. Raina (Syracuse University), S. B. MaqBool (Syracuse University), G. Jagadeeswaran (Syracuse University), B. R. Acharya (Syracuse University), J. Ehlting (University of British Columbia), Joerg Bohlman (University of British Columbia), H. Fescmyer (PSU). 2) Arabidopsis Leaf Age Project: Collaborators are H. M. Appel and J. C. Schultz. 3) Sink Strength Project: Collaborators are H. M. Appel, J. C. Schultz, T. A. Arnold (Dickinson University). 4) GLV Project: Collaborators are H. M. Appel, J. C. Schultz, J. H. Tumlinson, J.E. Carlson, C.M. De Moraes, M.C. Mescher, and Research Associates C. J. Frost, J. Engelberth and I. Seidl-Adams.
TARGET AUDIENCES: Scientists investigating plant stress, plant-pest interactions, and natural plant defenses.
Impacts
Impacts of these studies are mainly long-term. All of the phenomena studied contribute to plant resistance to pests, and most can be manipulated (enhanced) via breeding and/or biotechnology. The glucosinoate and green leaf volatile work provides specific targets for enhancing plant resistance. The poplar work has implications for carbon storage and sequestration in relation to global change. Our better understanding from the microarray work of how plants respond to multiple stresses will help the design of effective strategies to manage biotic stress under field conditions. Because we do our experiments with greater statistical rigor and under more realistic conditions than do many laboratories working with "model systems" (e.g. Arabidopsis), our results are having the effect of questioning many generalizations, with a presumed longer term impact of improving experimental design and producing more useful experimental results.
Publications
- Jagadeeswaran G., S. Raina, B. R. Acharya, S. B. Maqbool, S. L. Mosher, H. M. Appel, J. C. Schultz, D. F. Klessig, and R. Raina. 2007. Arabidopsis GH3-LIKE DEFENSE GENE 1 is required for accumulation of salicylic acid, activation of defense responses and resistance to Pseudomonas syringae. Plant J 51:234-46.
- Acharya B. R., S. Raina, S. B. Maqbool, G. Jagadeeswaran, S. L. Mosher, H. M. Appel, J. C. Schultz, D. F. Klessig, and R. Raina. 2007. Overexpression of CRK13, an Arabidopsis cysteine-rich receptor-like kinase, results in enhanced resistance to Pseudomonas syringae. Plant J. 50:488-99.
- Frost, C. J., H. M. Appel, J. E. Carlson, C. M. De Moraes, M. C. Mescher, and J. C. Schultz. 2007. Within-plant signalling via volatiles overcomes vascular constraints on systemic signalling and primes responses against herbivores, Ecol. Letters 10:490-498.
- Engelberth J, I. Seidl-Adams I, J. C. Schultz, and J. H. Tumlinson. 2007. Insect elicitors and exposure to green leafy volatiles differentially upregulate major octadecanoids and transcripts of 12-oxo phytodienoic acid reductases in Zea mays. Mol Plant-Microbe Interact. 20:707-716.
- LoVerde, B. 2007. Effect of leaf age on glucosinolate induction of Arabidopsis by caterpillars. Honors Thesis. The Pennsylvania State University, University Park, PA. 75 pp.
- McCartney, N. 2007. Analysis of Pieris rapae and Spodoptera exigua feeding site preferences on Arabidopsis thaliana Columbia ecotype in relation to within-plant chemical and structrual heterogeneity. M.S. Thesis. The Pennsylvania State University, University Park, PA. 130 pp.
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Progress 01/01/06 to 12/31/06
Outputs
The goals of this project are to determine whether plants can discriminate among different pest species when initiating defense responses, how responses are coordinated and supplied with resources, and whether any of this matters to plant success. The first year and a half of this project was funded by an NSF Arabidopsis 2010 grant; current support comes from a regular NSF grant. Several studies were underway and others completed during the past year. A brief summary: 1. An investigation of the role(s) of Arabidopsis thaliana receptor-like kinsases (RLKs) in identifying attacking pests was completed. Chemical and data analyses are still underway. Few of the small number of RLKs we were able to investigate appear to play a role in insect recognition, but some are involved in pathogen recognition. One paper relating to responses to pathogens has been accepted for publication. 2. A long study of the role of auxin, the bacterial toxin coronatine, and other plant hormones
on bacterial invasion of A. thaliana plants was completed. Auxin accumulates during infection by Pseudomonas syringae; its source was shown to be de novo synthesis, but no clear impact of this accumulation on pathogen success could be shown. The impacts of the bacterial toxin coronatine were shown to be independent of a signaling pathway ('oxylipin pathway') thought to be required for its activity. This work plus some important methods development comprises a successfully-defended PhD thesis (degree conferred 12/06), and papers are in preparation for publication. 3. We continued to analyze data from two large microarray studies. The data have been divided into those pertaining to functional genes, transcription factors, and signal-pathways. Invited presentations on these results have been given at national and international meetings, including a Gordon Research Conference. We hope to submit publications before the end of 2006. 4. A study of caterpillar feeding preferences on
Arabidopsis leaf-age cohorts was completed and is being written up as part of a Master's thesis. A specialist species was found to feed preferentially on young leaves with greater defense capacity while a generalist fed primarily on low-defense older leaves. Because these two caterpillar species alter the abundance of differentially-defended leaves and elicit different defense responses, they influence each other's food availability and feeding behavior, which has community ecology implications. 5. A long study of gall elicitation on grape leaves by phylloxera was concluded and written up as a PhD thesis (successfully defended, degree to be conferred 12/06). Results indicate that the insect elicits gall development by activating genes in the plant's floral development program ectopically. This is the first molecular evidence conerning gall development ever obtained. Publications are in preparation. 6. Work began as part of a newly-funded project continuing our studies of source-sink
control of defense responses in poplar. We are awaiting stable isotope data from a contract lab from these experiments and expect a publication to emerge early in 2007.
Impacts
Impacts of these studies are mainly long-term. All of the phenomena studied contribute to plant resistance to pests, and most can be manipulated (enhanced) via breeding and/or biotechnology. The poplar work has implications for carbon storage and sequestration in relation to global change. Because we do our experiments with greater statistical rigor and under more realistic conditions than do many laboratories working with "model systems" (e.g. Arabidopsis), our results are having the effect of questioning many generalizations.
Publications
- Fine, P. V. A., Miller, Z. J., Mesones, I., Irazuzta, S., Appel, H. M., Stevens, M. H., Saaksjarvi, I., Schultz, J. C., and Coley, P. D. 2006. The growth-defense tradeoff and habitat specialization by plants in Amazonian forests. Ecology 87: S150-S162 Suppl. S.
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Progress 01/01/05 to 12/31/05
Outputs
Goals of this 17month-old project are to 1) Characterize the ability of plants to recognize and respond specifically to pests and other environmental stimuli. 2) Elucidate underlying signaling systems that coordinate and produce appropriate biochemical responses. 3) Determine the ecological significance of coordinated responses. Systems are poplar and the brassicaceous weed, Arabidopsis thaliana; we report the following progress: 1. We published the first study of Arabidopsis responses to insects and showed dependence of responses on signaling pathways. Defense metabolite profiles were obtained, but unexpected discovery of many unknowns plus collaborator Jones' departure from PSU have delayed publication. A whole-genome microarray study of Arabidopsis responses to 4 insects produced an unprecedented data set still being interpreted, but showing for the first time that the plant responds specifically to different insect species. This work has been reported by
invitation at several international venues. 2. Unfunded studies showing that plant volatiles elicit glucosinolate defense responses in Arabidopsis were extended to demonstrate a priming effect, previously shown only for corn. 3. Unfunded thesis work on the role of the growth hormone auxin in plant defense responses to bacteria continued, to be completed as a PhD thesis during 2006. 4. Ongoing research characterizing receptor-like kinases in Arabidopsis produced several transgenic lines expressing the reporter construct PR1-GFP. Seeds from the initial transformation were plated on media containing hygromycin; resulting plants were analyzed to find the strongest PR1 expression in response to salicylic acid. Lines producing chimeric RLKs containing the BRI1 or FLS2 receptor domain fused to the 348/351/FLS2 kinase domains were produced. Preliminary analysis indicated that these RLKs are modular and their domains can be swapped to create sensors with novel functionalities. To assess this
we prepared cDNA microarrays containing kinase domains representing 223 LRR-RLKs. cDNAs were amplified using gene-specific primers and printed on glass slides in quadruplicates at different positions in the glass slides. For hybridization, total RNA was isolated from various plant tissues and from plants grown in various stress conditions listed below. For each treatment, at least two replicate experiments were done using different sets of plants. These experiments represent a total of more than 200 microarray hybridizations. Seeds from chimeric RLK plants are being analyzed to identify homozygous lines. Mutant plants with receptors knocked out have been planted for screening with elicitors. This subcontracted work (to Syracuse University) comprised the vast majority of research expenditures during this period. 5. Unfunded thesis work on the molecular basis of phylloxera gall development on grape leaves continued. Quantitative realtime analyses have been developed for 15 flower- and
fruit-development genes, and show that leaf gall development incorporates elements of fruit development programs.
Impacts
Impacts of this fundamental research project are likely to be longer-term. Discovering genetic and other underlying mechanisms of plant resistance to pests will eventually permit breeding or engineering for enhanced performance. Our findings indicate that this must be done in realistic ecological contexts, and we are trying to move the research community in this direction. The goal of our receptor studies is to develop the ability to use plants as environmental monitors and pest scouts for application in precision agriculture. The work with phylloxera will eventually suggest genes that could be involved in resistance to this important pest.
Publications
- Allison, S. D. and Schultz, J. C. 2005. Biochemical responses of chestnut oak to a galling cynipid. J. Chemical Ecol. 31: 151-166.
- Mewis, I., Appel, H. M., Hom, A., Raina, R. and Schultz, J. C. 2005. Major signaling pathways modulate Arabidopsis thaliana (L.) glucosinolate accumulation and response to both phloem feeding and chewing insects. Plant Physiology 138: 1149-1162.
- Walls, R., Appel, H., Cipollini, M., and Schultz, J. 2005. Fertility, root reserves and the cost of inducible defenses in the perennial plant Solanum carolinense. J. Chemical Ecol 31: 2263-2288.
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Progress 01/01/04 to 12/31/04
Outputs
This project has been in place for 5 months. Its goals are to 1) Characterize the ability of plants to recognize and respond specifically and concurrently to pests and other environmental stimuli. 2) Elucidate underlying signaling systems that coordinate and produce appropriate biochemical responses. 3) Determine the ecological significance of coordinated responses. Study systems are poplar and the brassicaceous weed, Arabidopsis thaliana. During the past 6 months we have carried out experiments aimed at determining patterns of gene expression and defense production by both plant species in response to a variety of insect and bacterial pests. Samples from these experiments are being analyzed at present, using microarray, real time PCR, and liquid chromatography-mass spectrometry approaches. We are presently examining statistical results from a whole-genome microarray analysis of Arabidopsis responses to 2 aphid and 2 Lepidopteran species as well as wounding carried
out over the past 12 months. These reveal significant differences in which and how many genes are up- or down-regulated by these 4 insects and indicate that specialist insects elicit less up-regulation and more down-regulation than do generalists. Studies with the pathogenic bacterium Pseudomonas syringae suggest that elicited increases in auxin accumulation may subvert plant defense responses. We have found that green leafy volatiles emitted by wounded plants elicit glucosinolate defense responses in Arabidopsis plants, and that insect feeding elicits emission of isothiocyanate volatiles from Arabidopsis. A continuing research program characterizing receptor-like kinases in Arabidopsis has produced a series of transgenic lines with chimeric receptors capable of indicating exposure to various stimuli via growth- or green fluorescent protein reporter constructs. We are beginning to be able to combine metabolomics and functional genomics to identify mechanisms underlying plant
resistance and susceptibility to pests. The collaboration with A. Dan Jones in Chemistry is both groundbreaking and essential to success in these efforts.
Impacts
Impacts of this fundamental research project are likely to be longer-term. Discovering genetic and other underlying mechanisms of plant resistance to pests will eventually permit breeding or engineering for enhanced performance. The goal of our receptor studies is to develop the ability to use plants as environmental monitors and pest scouts for application in precision agriculture. While these are popular ambitions, one contribution we are making to this field is to make it evident to lab-bound investigators how important it is to carry out chemical and genetic studies under ecologically-realistic conditions. We hope this will hasten applicability of research findings.
Publications
- No publications reported this period
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