The interplay between hormone signaling and defense gene expression in grapevine genotypes carrying genetic resistance against Plasmopara viticola

The Arabidopsis mutant wrky33 is highly susceptible to Botrytis cinerea. We identified >1680 Botrytis-induced WRKY33 binding sites associated with 1576 Arabidopsis genes. Transcriptional profiling defined 318 functional direct target genes at 14 hr post inoculation. Comparative analyses revealed that WRKY33 possesses dual functionality acting either as a repressor or as an activator in a promoter-context dependent manner. We confirmed known WRKY33 targets involved in hormone signaling and phytoalexin biosynthesis, but also uncovered a novel negative role of abscisic acid (ABA) in resistance towards B. cinerea 2100. The ABA biosynthesis genes NCED3 and NCED5 were identified as direct targets required for WRKY33-mediated resistance. Loss-of-WRKY33 function resulted in elevated ABA levels and genetic studies confirmed that WRKY33 acts upstream of NCED3/NCED5 to negatively regulate ABA biosynthesis. This study provides the first detailed view of the genome-wide contribution of a specific plant transcription factor in modulating the transcriptional network associated with plant immunity.DOI: http://dx.doi.org/10.7554/eLife.07295.001

Lentil cultivation is often hampered by aphid population outspreads with detrimental impacts to crop development and production, challenging food safety and agriculture sustainability. The pea aphid (Acyrthosiphon pisum) is a significant threat to lentil in the temperate zone rainfed systems. A set of management practices including resilient cultivars and application of insecticides have effectively controlled aphid infestation. However, the plant defense against insect pests is scantily dissected and limited to the individual components including antibiosis, antixenosis and tolerance that constitute a combination of plant stress responses. Utilizing a lentil germplasm collection, we assessed the antixenosis and aphid tolerance mechanisms in association to important morphological parameters. Physiological parameters including relative water content (RWC) measured at 24h and 48h post-aphid infestation revealed genotype-specific responses. The contents of key plant hormones including salicylic acid (SA), jasmonic acid (JA), abscisic acid (ABA) and indoleacetic acid (IAA) implicated in defense signal-transduction pathways were also determined in lentil accessions after aphid herbivory infestation. In parallel, the expression of hallmark defense genes governed by SA- and JA-signaling pathways at 24h and 48h post aphid herbivory revealed significant differentiation patterns among the accessions. An interplay of hormone crosstalk is unveiled that possibly governs defense responses and aphid resistance. Besides the metabolomic profiling of accessions under aphid herbivory indicated the indispensable role of key secondary metabolites accumulation such as flavonoids, alkaloids, phenolics and fatty acids as a front line of plant defense and a potential integration of hormone signaling pathways in metabolome reprogramming. Overall, the study presents a panorama of distinct lentil responses to aphids and a critical view of the molecular mechanisms implicated in lentil insect defense to further our insight and advance crop protection and breeding approaches in a climate changing environment.

The plant hormones abscisic acid (ABA), jasmonic acid (JA), and ethylene are involved in diverse plant processes, including the regulation of gene expression during adaptive responses to abiotic and biotic stresses. Previously, ABA has been implicated in enhancing disease susceptibility in various plant species, but currently very little is known about the molecular mechanisms underlying this phenomenon. In this study, we obtained evidence that a complex interplay between ABA and JA-ethylene signaling pathways regulate plant defense gene expression and disease resistance. First, we showed that exogenous ABA suppressed both basal and JA-ethylene-activated transcription from defense genes. By contrast, ABA deficiency as conditioned by the mutations in the ABA1 and ABA2 genes, which encode enzymes involved in ABA biosynthesis, resulted in upregulation of basal and induced transcription from JA-ethylene responsive defense genes. Second, we found that disruption of AtMYC2 (allelic to JASMONATE INSENSITIVE1 [JIN1]), encoding a basic helix-loop-helix Leu zipper transcription factor, which is a positive regulator of ABA signaling, results in elevated levels of basal and activated transcription from JA-ethylene responsive defense genes. Furthermore, the jin1/myc2 and aba2-1 mutants showed increased resistance to the necrotrophic fungal pathogen Fusarium oxysporum. Finally, using ethylene and ABA signaling mutants, we showed that interaction between ABA and ethylene signaling is mutually antagonistic in vegetative tissues. Collectively, our results indicate that the antagonistic interactions between multiple components of ABA and the JA-ethylene signaling pathways modulate defense and stress responsive gene expression in response to biotic and abiotic stresses.

The effects of exogenous methyl jasmonate (MeJA) on the resistance levels to quinclorac and endogenous hormone levels were examined using the resistant and sensitive biotypes of barnyardgrass with the same genetic background. The results showed that exogenous MeJA could significantly increase the resistance of resistant plants to quinclorac, but did not affect the resistance of sensitive plants. There were significant differences in the contents of indole-3-acetic acid (IAA), abscisic acid (ABA), salicylic acid (SA), and jasmonic acid (JA) among the tested materials. Quinclorac treatment could significantly increase the contents of ABA, SA, and JA in barnyardgrass, with stronger increase of ABA and JA contents in the sensitive plants. Exogenous MeJA could significantly enhance the induction of ABA and JA contents by quinclorac, especially in the sensitive biotypes. These results showed that rapid hormone variations in sensitive plants inhibited their response to herbicide, while gentle hormone variations endowed resistant plants with a strong adapta-bility to herbicide, which closely related to the enhanced resistance by MeJA. ABA and JA hormone signals might play an important role in the resistance of barnyardgrass to quinclorac stress.

<i>Moniliophthora perniciosa</i>produces hormones and alters endogenous auxin and salicylic acid in infected cocoa leaves

Phytohormones play essential roles in plant-nematode interactions through complex crosstalk. Although these hormones often accumulate in nematode-resistant plants, the roles of abscisic acid (ABA) and ethylene (ET) in rice (Oryza sativa) resistance to the root-knot nematode (RKN) Meloidogyne graminicola (Mg) remain unclear, particularly regarding concentration dependency and underlying mechanisms. Using exogenous hormone gradient treatments, we show that only high concentrations of ABA (200 µM) and ET-releasing compound ethephon (Eth, 500 µM) induce systemic nematode resistance. High-concentration ET triggers endogenous systemic accumulation of ET and jasmonic acid (JA), accompanied by transient suppression followed by delayed accumulation of ABA, and induces JA-, ABA-, and salicylic acid (SA)-associated transcriptional responses. Exogenous ABA leads to endogenous ABA and SA accumulation and increased expression of related genes, while it suppresses ET biosynthesis gene expression and levels, highlighting a negative feedback effect of ABA on ET. Both hormones converge on a common mitogen-activated protein kinase 5 (OsMPK5)-dependent transcriptional and translational module. Low doses of ABA (50 µM) failed to activate this module and induced plant susceptibility, highlighting a threshold requirement for immune activation. Offspring of rice plants treated bi-weekly with high doses of ABA or ET were less susceptible to nematodes. This intergenerational acquired resistance was also OsMPK5-dependent. Our findings reveal concentration-dependent systemic effects of ABA and ET, whereby high-dose ABA and ET converge on OsMPK5 to reprogram translation and defense gene expression, underpinning both immediate and heritable resistance to root-knot nematodes.

To characterize cultivar variations in hormonal regulation of the transition between pattern-triggered immunity (PTI) and effector-triggered immunity or susceptibility (ETI or ETS), the responses of resistance (R-) genes, hydrogen peroxide, and proline metabolism in two Brassica napus cultivars to contrasting disease susceptibility (resistant cv. Capitol vs. susceptible cv. Mosa) were interpreted as being linked to those of endogenous hormonal levels and signaling genes based on a time course of disease symptom development. Disease symptoms caused by the Xanthomonas campestris pv. campestris (Xcc) infections were much more developed in cv. Mosa than in cv. Capitol, as shown by an earlier appearance (at 3 days postinoculation [3 DPI]) and larger V-shaped necrosis lesions (at 9–15 DPI) in cv. Mosa. The cultivar variations in the R-genes, hormone status, and proline metabolism were found in two different phases (early [0–3 DPI] and later [9–15 DPI]). In the early phase, Xcc significantly upregulated PTI-related cytoplasmic kinase (Botrytis-induced kinase-1 [BIK1]) expression (+6.3-fold) with salicylic acid (SA) accumulation in cv. Capitol, while relatively less (+2.6-fold) with highly increased jasmonic acid (JA) level in cv. Mosa. The Xcc-responsive proline accumulation in both cultivars was similar to upregulated expression of proline synthesis-related genes (P5CS2 and P5CR). During the later phase in cv. Capitol, Xcc-responsive upregulation of ZAR1 (a coiled-coil-nucleotide binding site-leucine-rich repeat [CC-NB-LRR-type R-gene]) was concomitant with a gradual increase in JA levels without additional proline accumulation. However, in cv. Mosa, upregulation of TAO1 (a toll/interleukin-1 receptor-nucleotide binding site-leucine-rich repeat [TIR-NB-LRR-type R-gene]) was consistent with an increase in SA and abscisic acid (ABA) levels and resulted in an antagonistic depression of JA, which led to a proline accumulation. These results indicate that Xcc-induced BIK1- and ZAR1-mediated JA signaling interactions provide resistance and confirm ETI, whereas BIK1- and TAO1-enhanced SA- and/or ABA-mediated proline accumulation is associated with disease susceptibility (ETS).

Grapevine downy mildew, caused by Plasmopara viticola, poses significant threats to viticulture, necessitating sustainable alternatives to chemical fungicides. This study investigates the efficacy of exogenous 24-epibrassinolide (EBR), a brassinosteroid, in enhancing disease resistance across two grape cultivars: susceptible Vitis vinifera Cabernet Sauvignon and resistant Ecolly. EBR application reduced P. viticola colonization in both cultivars, suppressing hyphal growth and sporulation. In Cabernet Sauvignon, EBR delayed pathogen establishment by modulating stomatal closure and boosting antioxidant enzymes (superoxide dismutase, catalase, ascorbate peroxidase, peroxidase) and pathogenesis-related proteins (chitinase, β-1,3-glucanase). Ecolly exhibited inherent resistance, characterized by localized hypersensitive responses and elevated salicylic acid (SA) and jasmonic acid (JA) levels post-EBR treatment, which amplified systemic acquired resistance (SAR) via upregulation of PAD4, EDS1, and PRs genes. Stomatal regulation in Cabernet Sauvignon correlated with EBR-induced abscisic acid (ABA) dynamics, while Ecolly maintained constitutive stomatal defense. Transcriptional analysis revealed cultivar-specific phytohormonal interplay, with SA pathways dominating in Ecolly and delayed SA accumulation in Cabernet Sauvignon. These findings demonstrate that EBR enhances resistance through multifaceted mechanisms-pathogen inhibition, redox homeostasis, hormonal signaling, and genetic priming-with efficacy modulated by genetic background. This study underscores brassinosteroid as a sustainable tool for integrated disease management in grapevines.

Response of phytohormones and correlation of SAR signal pathway genes to the different resistance levels of grapevine against Plasmopara viticola infection

Plants can sometimes be under the influence of more than one stress factor. In this case, combined stress factors can cause different responses in plants. In this study, the phytohormonal activity and hormone gene expressions of beans (Phaseolus vulgaris L.) were investigated in different drought levels and soils with different levels of cadmium accumulation. Plants were grown at 4 different cadmium (Cd) levels (Cd: 0, Cd1: 100, Cd2: 150, and Cd3: 200 mg kg$^{-1}$) and 3 irrigation levels (D0: 100%, D1: 75%, and D2: 50% of field capacity). Abscisic acid (ABA), indole acetic acid (IAA), gibberellic acid (GA), salicylic acid (SA), cytokinin, zeatin, and jasmonic acid contents and gene expressions related to them were investigated. As a result of the study, we determined that there was a significant decrease in the content of IAA, GA, SA, cytokinin, zeatin, and jasmonic acid with increasing doses of drought and Cd. Especially with the coexistence of both stress factors, this reducing effect (the reductions in these hormones were 86.6%, 64.1%, 71.9%, 66.7%, 45.5%, and 73.9% as compared to D0, respectively) on the measured parameters was greater. However, ABA content increased significantly with drought and/or Cd stresses, especially with Cd3D2, about a thousand times higher than the control. On the other hand, mRNA transcript levels of some hormone genes (ABA 8'-hydroxylase, adenylate isopentenyltransferase (AIPT3), auxin-related protein (AUX/ IAA), MeJA biosynthesis and gibberellin 20-oxidase and NRAMP2 gene known to be involved in Cd transport, were detected. According to the results, while ABA 8'-hydroxylase, gibberellin 20-oxidase, and NRAMP2 genes were overexpressed in all stress treatments, AIPT3 and AUX/IAA genes were down-regulated and also, MeJA biosynthesis gene varied depending on dose and stress.

Phytohormones play critical roles in regulating plant responses to stress. We investigated the effects of salinity on abscisic acid (ABA), indole-3-acetic acid (IAA), salicylic acid (SA), and jasmonic acid (JA) in leaves, stalks, fruits, and seeds of Iris hexagona, a native wetland species. Using gas chromatography-mass spectroscopy with selected ion monitoring, our experiments demonstrated significant and different short- and long-term changes in iris phytohormones. ABA and JA generally increased and IAA and SA declined in response to salinity. We conclude that these phytohormones may have separate and interactive effects on how plants respond and adapt to stress in natural environments.

An efficient simplified isotope dilution method was developed to determine four carboxyl containing phytohormones simultaneously in 200 mg of fresh tomato tissues using ultra high performance liquid chromatography-triple quadrupole mass spectrometry (UPLC-MS/MS) with negative electrospray ionization. The four phytohormones are indole-3-acetic acid (IAA), abscisic acid (ABA), jasmonic acid (JA), and salicylic acid (SA). Only one purification step of Oasis MAX solid phase extraction (SPE) was employed to enrich target phytohormones after crude extraction. In addition, two endogenous isomers of JA, (-)-JA and (+)-7-iso-JA, were separated directly. The validated method has been applied to monitor changes of JA, SA, IAA, and ABA in both local and systemic leaves of wild-type and transgenic 35S::prosystemin (35S::PS) tomato lines. Meanwhile, the JA burst amplified by the overexpressed prosystemin in 35S::PS was verified. Furthermore, the spatial and temporal changes of JA, SA, ABA, and IAA were analyzed.

The greenbug aphid Schizaphis graminum causes significant damage to wheat crops, so increasing plant resistance against aphids is one of the primary tasks. It has been shown that colonization by aphids to trigger both jasmonate/ethylene- and salicylate-dependent defense responses in plants. Ethylene is produced when insects attack. It has been suggested that ethylene plays an important role in inducing resistance against pests, but the mechanism of action of ethylene is not fully understood. In this work, the role of ethylene in the induction of hormonal signaling pathways in wheat plants during the development of resistance against the greenbug aphid S. graminum was revealed by treating plants with the ethylene receptor inhibitor 1-MCP (1-methylcyclopropene) or an ethylene precursor (ethephon - ET). Analysis of the transcriptional activity of the genes of the hormonal signaling pathways ethylene, salicylic acid (SA), jasmonic acid (JA), cytokinins (CK) and abscisic acid (ABA) showed that treatment of plants with ET activated the genes SA-, JA-, ABA- and ethylene- signaling pathways (TaEIN3, TaERF1, TaWRKY53b, TaPR3, TaPR1, TaWRKY13, TaPR6, TaABI5, TaABAI and TaNCED).The study of the endogenous level of phytohormones using enzyme immunoassay showed that treatment of plants with ET induced the accumulation of ABA and indolylacetic acid (IAA), but not the content of CK after the plants were colonized by aphids. Thus, the positive effect of ethylene on the resistance of wheat plants against the greenbug aphid was proven through the synergistic effect of ethylene with SA and JA and the positive or negative regulation of the activity of the components of these signaling pathways by the phytohormones CK, ABA and IAA.

Hormonal signaling regulates photosynthetic function of alfalfa (Medicago sativa L.) under NaHCO3 stress

Supramolecular solvent-based high-throughput sample treatment for monitoring phytohormones in plant tissues