TY - CHAP ID - 2541 TI - Plant–Insect Bioassay for Testing Arabidopsis Resistance to the Generalist Herbivore Spodoptera littoralis T2 - Jasmonate in Plant Biology PB - Methods Mol Biol PY - 2020 SP - 69-78 AU - Mielke, S. AU - Gasperini, D. VL - 2085 UR - SN - 978-1-0716-0142-6 DO - 10.1007/978-1-0716-0142-6_5 AB - Jasmonates are essential engineers of plant defense responses against many pests, including herbivorous insects. Herbivory induces the production of jasmonic acid (JA) and its bioactive conjugate jasmonoyl-l-isoleucine (JA-Ile), which then triggers a large transcriptional reprogramming to promote plant acclimation. The contribution of the JA pathway, including its components and regulators, to defense responses against insect herbivory can be evaluated by conducting bioassays with a wide range of host plants and insect pests. Here, we describe a detailed and reproducible protocol for testing feeding behavior of the generalist herbivore Spodoptera littoralis on the model plant Arabidopsis thaliana and hence infer the contribution of JA-mediated plant defense responses to a chewing insect. A2 - Champion, A. & Laplaze, L., eds. C1 - Molecular Signal Processing ER - TY - INPR ID - 2425 TI - Inhibition of jasmonate-mediated plant defences by the fungal metabolite higginsianin B JO - bioRxiv PY - 2019 SP - AU - Dallery, J.-F. AU - Zimmer, M. AU - Halder, V. AU - Suliman, M. AU - Pigné, S. AU - Le Goff, G. AU - Gianniou, D. D. AU - Trougakos, I. P. AU - Ouazzani, J. AU - Gasperini, D. AU - O’Connell, R. J. VL - UR - https://dx.doi.org/10.1101/651562 DO - 10.1101/651562 AB - Infection of Arabidopsis thaliana by the ascomycete fungus Colletotrichum higginsianum is characterised by an early symptomless biotrophic phase followed by a destructive necrotrophic phase. The fungal genome contains 77 secondary metabolism-related biosynthetic gene clusters (BGCs), and their expression during the infection process is tightly regulated. Deleting CclA, a chromatin regulator involved in repression of some BGCs through H3K4 trimethylation, allowed overproduction of 3 families of terpenoids and isolation of 12 different molecules. These natural products were tested in combination with methyl jasmonate (MeJA), an elicitor of jasmonate responses, for their capacity to alter defence gene induction in Arabidopsis. Higginsianin B inhibited MeJA-triggered expression of the defence reporter VSP1p:GUS, suggesting it may block bioactive JA-Ile synthesis or signalling in planta. Using the JA-Ile sensor Jas9-VENUS, we found that higginsianin B, but not three other structurally-related molecules, suppressed JA-Ile signalling by preventing degradation of JAZ proteins, the repressors of JA responses. Higginsianin B likely blocks the 26S proteasome-dependent degradation of JAZ proteins because it inhibited chymotrypsin- and caspase-like protease activities. The inhibition of target degradation by higginsianin B also extended to auxin signalling, as higginsianin B treatment reduced IAA-dependent expression of DR5p:GUS. Overall, our data indicate that specific fungal secondary metabolites can act similarly to protein effectors to subvert plant immune and developmental responses. A2 - C1 - Molecular Signal Processing ER - TY - JOUR ID - 2325 TI - Shoot-to-root translocation of the jasmonate precursor 12-oxo-phytodienoic acid (OPDA) coordinates plant growth responses following tissue damage JO - bioRxiv PY - 2019 SP - AU - Schulze, A. AU - Zimmer, M. AU - Mielke, S. AU - Stellmach, H. AU - Melnyk, C. W. AU - Hause, B. AU - Gasperini, D. VL - UR - https://doi.org/10.1101/517193 DO - 10.1101/517193 AB - Multicellular organisms rely upon the movement of signaling molecules across cells, tissues and organs to communicate among distal sites. In plants, herbivorous insects, necrotrophic pathogens and mechanical wounding stimulate the activation of the jasmonate (JA) pathway, which in turn triggers the transcriptional changes necessary to protect plants against those challenges, often at the expense of growth. Although previous evidence indicated that JA species can translocate from damaged into distal sites, the identity of the mobile compound(s), the tissues through which they translocate and the consequences of their relocation remain unknown. Here, we demonstrated that endogenous JA species generated after shoot injury translocate to unharmed roots via the phloem vascular tissue in Arabidopsis thaliana. By wounding wild-type shoots of chimeric plants and by quantifying the relocating compounds from their JA-deficient roots, we uncovered that the JA-Ile precursor 12-oxo-phytodienoic acid (OPDA) is a mobile JA species. Our data also showed that OPDA is a primary mobile compound relocating to roots where, upon conversion to the bioactive hormone, it induces JA-mediated gene expression and root growth inhibition. Collectively, our findings reveal the existence of long-distance transport of endogenous OPDA which serves as a communication molecule to coordinate shoot-to-root responses, and highlight the importance of a controlled distribution of JA species among organs during plant stress acclimation. A2 - C1 - Molecular Signal Processing; Cell and Metabolic Biology ER - TY - JOUR ID - 2427 TI - Wound-Induced Shoot-to-Root Relocation of JA-Ile Precursors Coordinates Arabidopsis Growth JO - Mol Plant PY - 2019 SP - 1383-1394 AU - Schulze, A. AU - Zimmer, M. AU - Mielke, S. AU - Stellmach, H. AU - Melnyk, C. W. AU - Hause, B. AU - Gasperini, D. VL - 12 UR - https://dx.doi.org/10.1016/j.molp.2019.05.013 DO - 10.1016/j.molp.2019.05.013 AB - Multicellular organisms rely on the movement of signaling molecules across cells, tissues, and organs to communicate among distal sites. In plants, localized leaf damage activates jasmonic acid (JA)-dependent transcriptional reprogramming in both harmed and unharmed tissues. Although it has been indicated that JA species can translocate from damaged into distal sites, the identity of the mobile compound(s), the tissues through which they translocate, and the effect of their relocation remain unknown. Here, we found that following shoot wounding, the relocation of endogenous jasmonates through the phloem is essential to initiate JA signaling and stunt growth in unharmed roots of Arabidopsis thaliana. By employing grafting experiments and hormone profiling, we uncovered that the hormone precursor cis-12-oxo-phytodienoic acid (OPDA) and its derivatives, but not the bioactive JA-Ile conjugate, translocate from wounded shoots into undamaged roots. Upon root relocation, the mobile precursors cooperatively regulated JA responses through their conversion into JA-Ile and JA signaling activation. Collectively, our findings demonstrate the existence of long-distance translocation of endogenous OPDA and its derivatives, which serve as mobile molecules to coordinate shoot-to-root responses, and highlight the importance of a controlled redistribution of hormone precursors among organs during plant stress acclimation. A2 - C1 - Molecular Signal Processing; Cell and Metabolic Biology ER - TY - JOUR ID - 2441 TI - Interplay between Plant Cell Walls and Jasmonate Production JO - Plant Cell Physiol PY - 2019 SP - 2629-2637 AU - Mielke, S. AU - Gasperini, D. VL - 60 UR - https://dx.doi.org/10.1093/pcp/pcz119 DO - 10.1093/pcp/pcz119 AB - Plant cell walls are sophisticated carbohydrate-rich structures representing the immediate contact surface with the extracellular environment, often serving as the first barrier against biotic and abiotic stresses. Notably, a variety of perturbations in plant cell walls result in upregulated jasmonate (JA) production, a phytohormone with essential roles in defense and growth responses. Hence, cell wall-derived signals can initiate intracellular JA-mediated responses and the elucidation of the underlying signaling pathways could provide novel insights into cell wall maintenance and remodeling, as well as advance our understanding on how is JA biosynthesis initiated. This Mini Review will describe current knowledge about cell wall-derived damage signals and their effects on JA biosynthesis, as well as provide future perspectives. A2 - C1 - Molecular Signal Processing ER - TY - JOUR ID - 2019 TI - Agronomic assessment of the wheat semi-dwarfing gene Rht8 in contrasting nitrogen treatments and water regimes JO - Field Crops Res PY - 2016 SP - 150-160 AU - Kowalski, A. M. AU - Gooding, M. AU - Ferrante, A. AU - Slafer, G. A. AU - Orford, S. AU - Gasperini, D. AU - Griffiths, S. VL - 191 UR - https://dx.doi.org/10.1016/j.fcr.2016.02.026 DO - 10.1016/j.fcr.2016.02.026 AB - Reduced height 8 (Rht8) is the main alternative to the GA-insensitive Rht alleles in hot and dry environments where it reduces plant height without yield penalty. The potential of Rht8 in northern-European wheat breeding remains unclear, since the close linkage with the photoperiod-insensitive allele Ppd-D1a is unfavourable in the relatively cool summers. In the present study, two near-isogenic lines (NILs) contrasting for the Rht8/tall allele from Mara in a UK-adapted and photoperiod-sensitive wheat variety were evaluated in trials with varying nitrogen fertiliser (N) treatments and water regimes across sites in the UK and Spain.The Rht8 introgression was associated with a robust height reduction of 11% regardless of N treatment and water regime and the Rht8 NIL was more resistant to root-lodging at agronomically-relevant N levels than the tall NIL. In the UK with reduced solar radiation over the growing season than the site in Spain, the Rht8 NIL showed a 10% yield penalty at standard agronomic N levels due to concomitant reduction in grain number and spike number whereas grain weight and harvest index were not significantly different to the tall NIL. The yield penalty associated with the Rht8 introgression was overcome at low N and in irrigated conditions in the UK, and in the high-temperature site in Spain. Decreased spike length and constant spikelet number in the Rht8 NIL resulted in spike compaction of 15%, independent of N and water regime. The genetic interval of Rht8 overlaps with the compactum gene on 2DS, raising the possibility of the same causative gene. Further genetic dissection of these loci is required. A2 - C1 - Molecular Signal Processing ER - TY - JOUR ID - 1961 TI - Cotyledon Wounding of Arabidopsis Seedlings JO - Bio Protoc PY - 2016 SP - e1712 AU - Gasperini, D. AU - Acosta, I. F. AU - Farmer, E. E. VL - 6 UR - https://dx.doi.org/10.21769/BioProtoc.1712 DO - 10.21769/BioProtoc.1712 AB - Damage to plant organs through both biotic and abiotic injury is very common in nature. Arabidopsis thaliana 5-day-old (5-do) seedlings represent an excellent system in which to study plant responses to mechanical wounding, both at the site of the damage and in distal unharmed tissues. Seedlings of wild type, transgenic or mutant lines subjected to single or repetitive cotyledon wounding can be used to quantify morphological alterations (e.g., root length, Gasperini et al., 2015), analyze the dynamics of reporter genes in vivo (Larrieu et al., 2015; Gasperini et al., 2015), follow transcriptional changes by quantitative RT-PCR (Acosta et al., 2013; Gasperini et al., 2015) or examine additional aspects of the wound response with a plethora of downstream procedures. Here we illustrate how to rapidly and reliably wound cotyledons of young seedlings, and show the behavior of two promoters driving the expression of β-glucuronidase (GUS) in entire seedlings and in the primary root meristem, following single or repetitive cotyledon wounding respectively. We describe two procedures that can be easily adapted to specific experimental needs. A2 - C1 - Molecular Signal Processing ER - TY - JOUR ID - 1963 TI - Multilayered Organization of Jasmonate Signalling in the Regulation of Root Growth JO - PLOS Genet PY - 2015 SP - e1005300 AU - Gasperini, D. AU - Chételat, A. AU - Acosta, I. F. AU - Goossens, J. AU - Pauwels, L. AU - Goossens, A. AU - Dreos, R. AU - Alfonso, E. AU - Farmer, E. E. VL - 11 UR - https://dx.doi.org/10.1371/journal.pgen.1005300 DO - 10.1371/journal.pgen.1005300 AB - Physical damage can strongly affect plant growth, reducing the biomass of developing organs situated at a distance from wounds. These effects, previously studied in leaves, require the activation of jasmonate (JA) signalling. Using a novel assay involving repetitive cotyledon wounding in Arabidopsis seedlings, we uncovered a function of JA in suppressing cell division and elongation in roots. Regulatory JA signalling components were then manipulated to delineate their relative impacts on root growth. The new transcription factor mutant myc2-322B was isolated. In vitro transcription assays and whole-plant approaches revealed that myc2-322B is a dosage-dependent gain-of-function mutant that can amplify JA growth responses. Moreover, myc2-322B displayed extreme hypersensitivity to JA that totally suppressed root elongation. The mutation weakly reduced root growth in undamaged plants but, when the upstream negative regulator NINJA was genetically removed, myc2-322B powerfully repressed root growth through its effects on cell division and cell elongation. Furthermore, in a JA-deficient mutant background, ninja1 myc2-322B still repressed root elongation, indicating that it is possible to generate JA-responses in the absence of JA. We show that NINJA forms a broadly expressed regulatory layer that is required to inhibit JA signalling in the apex of roots grown under basal conditions. By contrast, MYC2, MYC3 and MYC4 displayed cell layer-specific localisations and MYC3 and MYC4 were expressed in mutually exclusive regions. In nature, growing roots are likely subjected to constant mechanical stress during soil penetration that could lead to JA production and subsequent detrimental effects on growth. Our data reveal how distinct negative regulatory layers, including both NINJA-dependent and -independent mechanisms, restrain JA responses to allow normal root growth. Mechanistic insights from this work underline the importance of mapping JA signalling components to specific cell types in order to understand and potentially engineer the growth reduction that follows physical damage. A2 - C1 - Molecular Signal Processing ER - TY - JOUR ID - 1962 TI - Axial and Radial Oxylipin Transport JO - Plant Physiol PY - 2015 SP - 2244-2254 AU - Gasperini, D. AU - Chauvin, A. AU - Acosta, I. F. AU - Kurenda, A. AU - Stolz, S. AU - Chételat, A. AU - Wolfender, J.-L. AU - Farmer, E. E. VL - 169 UR - https://dx.doi.org/10.1104/pp.15.01104 DO - 10.1104/pp.15.01104 AB - Jasmonates are oxygenated lipids (oxylipins) that control defense gene expression in response to cell damage in plants. How mobile are these potent mediators within tissues? Exploiting a series of 13-lipoxygenase (13-lox) mutants in Arabidopsis (Arabidopsis thaliana) that displays impaired jasmonic acid (JA) synthesis in specific cell types and using JA-inducible reporters, we mapped the extent of the transport of endogenous jasmonates across the plant vegetative growth phase. In seedlings, we found that jasmonate (or JA precursors) could translocate axially from wounded shoots to unwounded roots in a LOX2-dependent manner. Grafting experiments with the wild type and JA-deficient mutants confirmed shoot-to-root oxylipin transport. Next, we used rosettes to investigate radial cell-to-cell transport of jasmonates. After finding that the LOX6 protein localized to xylem contact cells was not wound inducible, we used the lox234 triple mutant to genetically isolate LOX6 as the only JA precursor-producing LOX in the plant. When a leaf of this mutant was wounded, the JA reporter gene was expressed in distal leaves. Leaf sectioning showed that JA reporter expression extended from contact cells throughout the vascular bundle and into extravascular cells, revealing a radial movement of jasmonates. Our results add a crucial element to a growing picture of how the distal wound response is regulated in rosettes, showing that both axial (shoot-to-root) and radial (cell-to-cell) transport of oxylipins plays a major role in the wound response. The strategies developed herein provide unique tools with which to identify intercellular jasmonate transport routes. A2 - C1 - Molecular Signal Processing ER - TY - JOUR ID - 1964 TI - The squeeze cell hypothesis for the activation of jasmonate synthesis in response to wounding JO - New Phytol PY - 2014 SP - 282-288 AU - Farmer, E. E. AU - Gasperini, D. AU - Acosta, I. F. VL - 204 UR - https://dx.doi.org/10.1111/nph.12897 DO - 10.1111/nph.12897 AB - Jasmonates are lipid mediators that control defence gene expression in response to wounding and other environmental stresses. These small molecules can accumulate at distances up to several cm from sites of damage and this is likely to involve cell‐to‐cell jasmonate transport. Also, and independently of jasmonate synthesis, transport and perception, different long‐distance wound signals that stimulate distal jasmonate synthesis are propagated at apparent speeds of several cm min–1 to tissues distal to wounds in a mechanism that involves clade 3 GLUTAMATE RECEPTOR‐LIKE (GLR) genes. A search for jasmonate synthesis enzymes that might decode these signals revealed LOX6, a lipoxygenase that is necessary for much of the rapid accumulation of jasmonic acid at sites distal to wounds. Intriguingly, the LOX6 promoter is expressed in a distinct niche of cells that are adjacent to mature xylem vessels, a location that would make these contact cells sensitive to the release of xylem water column tension upon wounding. We propose a model in which rapid axial changes in xylem hydrostatic pressure caused by wounding travel through the vasculature and lead to slower, radially dispersed pressure changes that act in a clade 3 GLR‐dependent mechanism to promote distal jasmonate synthesis. A2 - C1 - Molecular Signal Processing ER - TY - JOUR ID - 1965 TI - Role of NINJA in root jasmonate signaling JO - Proc Natl Acad Sci USA PY - 2013 SP - 15473-15478 AU - Acosta, I. F. AU - Gasperini, D. AU - Chételat, A. AU - Stolz, S. AU - Santuari, L. AU - Farmer, E. E. VL - 110 UR - https://dx.doi.org/10.1073/pnas.1307910110 DO - 10.1073/pnas.1307910110 AB - Wound responses in plants have to be coordinated between organs so that locally reduced growth in a wounded tissue is balanced by appropriate growth elsewhere in the body. We used a JASMONATE ZIM DOMAIN 10 (JAZ10) reporter to screen for mutants affected in the organ-specific activation of jasmonate (JA) signaling in Arabidopsis thaliana seedlings. Wounding one cotyledon activated the reporter in both aerial and root tissues, and this was either disrupted or restricted to certain organs in mutant alleles of core components of the JA pathway including COI1, OPR3, and JAR1. In contrast, three other mutants showed constitutive activation of the reporter in the roots and hypocotyls of unwounded seedlings. All three lines harbored mutations in Novel Interactor of JAZ (NINJA), which encodes part of a repressor complex that negatively regulates JA signaling. These ninja mutants displayed shorter roots mimicking JA-mediated growth inhibition, and this was due to reduced cell elongation. Remarkably, this phenotype and the constitutive JAZ10 expression were still observed in backgrounds lacking the ability to synthesize JA or the key transcriptional activator MYC2. Therefore, JA-like responses can be recapitulated in specific tissues without changing a plant’s ability to make or perceive JA, and MYC2 either has no role or is not the only derepressed transcription factor in ninja mutants. Our results show that the role of NINJA in the root is to repress JA signaling and allow normal cell elongation. Furthermore, the regulation of the JA pathway differs between roots and aerial tissues at all levels, from JA biosynthesis to transcriptional activation. A2 - C1 - Molecular Signal Processing ER - TY - JOUR ID - 1966 TI - Genetic and physiological analysis of Rht8 in bread wheat: an alternative source of semi-dwarfism with a reduced sensitivity to brassinosteroids JO - J Exp Bot PY - 2012 SP - 4419-4436 AU - Gasperini, D. AU - Greenland, A. AU - Hedden, P. AU - Dreos, R. AU - Harwood, W. AU - Griffiths, S. VL - 63 UR - https://dx.doi.org/10.1093/jxb/ers138 DO - 10.1093/jxb/ers138 AB - Over the next decade, wheat grain production must increase to meet the demand of a fast growing human population. One strategy to meet this challenge is to raise wheat productivity by optimizing plant stature. The Reduced height 8 (Rht8) semi-dwarfing gene is one of the few, together with the Green Revolution genes, to reduce stature of wheat (Triticum aestivum L.), and improve lodging resistance, without compromising grain yield. Rht8 is widely used in dry environments such as Mediterranean countries where it increases plant adaptability. With recent climate change, its use could become increasingly important even in more northern latitudes. In the present study, the characterization of Rht8 was furthered. Morphological analyses show that the semi-dwarf phenotype of Rht8 lines is due to shorter internodal segments along the wheat culm, achieved through reduced cell elongation. Physiological experiments show that the reduced cell elongation is not due to defective gibberellin biosynthesis or signalling, but possibly to a reduced sensitivity to brassinosteroids. Using a fine-resolution mapping approach and screening 3104 F2 individuals of a newly developed mapping population, the Rht8 genetic interval was reduced from 20.5 cM to 1.29 cM. Comparative genomics with model genomes confined the Rht8 syntenic intervals to 3.3 Mb of the short arm of rice chromosome 4, and to 2 Mb of Brachypodium distachyon chromosome 5. The very high resolution potential of the plant material generated is crucial for the eventual cloning of Rht8. A2 - C1 - Molecular Signal Processing ER - TY - JOUR ID - 1967 TI - S-RNase-like Sequences in Styles of Coffea (Rubiaceae). Evidence for S-RNase Based Gametophytic Self-Incompatibility? JO - Trop Plant Biol PY - 2011 SP - 237-249 AU - Asquini, E. AU - Gerdol, M. AU - Gasperini, D. AU - Igic, B. AU - Graziosi, G. AU - Pallavicini, A. VL - 4 UR - https://dx.doi.org/10.1007/s12042-011-9085-2 DO - 10.1007/s12042-011-9085-2 AB - Although RNase-based self-incompatibility (SI) is suspected to operate in a wide group of plant families, it has been characterized as the molecular genetic basis of SI in only three distantly related families, Solanaceae, Plantaginaceae, and Rosaceae, all described over a decade ago. Previous studies found that gametophytic SI, controlled by a multi-allelic S-locus, operates in the coffee family (Rubiaceae). The molecular genetic basis of this mechanism remains unknown, despite the immense importance of coffee as an agricultural commodity. Here, we isolated ten sequences with features of T2-S-type RNases from two Coffea species. While three of the sequences were identified in both species and clearly do not appear to be S-locus products, our data suggest that six sequences may be S-alleles in the self-incompatible C. canephora, and one may be a relict in the self-compatible C. arabica. We demonstrate that these sequences show style-specific expression, display polymorphism in C. canephora, and cluster with S-locus products in a phylogenetic analysis that includes other plant families with RNase-based SI. Although our results are not definitive, in part because the available plant materials were limited and data patterns relatively complex, our results strongly hint that RNase-based SI mechanism operates in the Rubiaceae family. A2 - C1 - Molecular Signal Processing ER - TY - JOUR ID - 1960 TI - Differential responses of Coffea arabica L. leaves and roots to chemically induced systemic acquired resistance JO - Genome PY - 2006 SP - 1594-1605 AU - De Nardi, B. AU - Dreos, R. AU - Del Terra, L. AU - Martellossi, C. AU - Asquini, E. AU - Tornincasa, P. AU - Gasperini, D. AU - Pacchioni, B. AU - Rathinavelu, R. AU - Pallavicini, A. AU - Graziosi, G. VL - 49 UR - https://dx.doi.org/10.1139/g06-125 DO - 10.1139/g06-125 AB - Coffea arabica is susceptible to several pests and diseases, some of which affect the leaves and roots. Systemic acquired resistance (SAR) is the main defence mechanism activated in plants in response to pathogen attack. Here, we report the effects of benzo(1,2,3)thiadiazole-7-carbothioic acid-s-methyl ester (BTH), a SAR chemical inducer, on the expression profile of C. arabica. Two cDNA libraries were constructed from the mRNA isolated from leaves and embryonic roots to create 1587 nonredundant expressed sequence tags (ESTs). We developed a cDNA microarray containing 1506 ESTs from the leaves and embryonic roots, and 48 NBS-LRR (nucleotide-binding site leucine-rich repeat) gene fragments derived from 2 specific genomic libraries. Competitive hybridization between untreated and BTH-treated leaves resulted in 55 genes that were significantly overexpressed and 16 genes that were significantly underexpressed. In the roots, 37 and 42 genes were over and underexpressed, respectively. A general shift in metabolism from housekeeping to defence occurred in the leaves and roots after BTH treatment. We observed a systemic increase in pathogenesis-related protein synthesis, in the oxidative burst, and in the cell wall strengthening processes. Moreover, responses in the roots and leaves varied significantly. A2 - C1 - Molecular Signal Processing ER -