Abscisic Acid Receptor Research Articles

RBR-Type E3 Ligases and the Ubiquitin-Conjugating Enzyme UBC26 Regulate Abscisic Acid Receptor Levels and Signaling.

The turnover of abscisic acid (ABA) signaling core components modulates the plant's response to ABA and is regulated by ubiquitination. We show that Arabidopsis (Arabidopsis thaliana) RING Finger ABA-Related1 (RFA1) and RFA4 E3 ubiquitin ligases, members of the RING between RING fingers (RBR)-type RSL1/RFA family, are key regulators of ABA receptor stability in root and leaf tissues, targeting ABA receptors for degradation in different subcellular locations. RFA1 is localized both in the nucleus and cytosol, whereas RFA4 shows specific nuclear localization and promotes nuclear degradation of ABA receptors. Therefore, members of the RSL1/RFA family interact with ABA receptors at plasma membrane, cytosol, and nucleus, targeting them for degradation via the endosomal/vacuolar RSL1-dependent pathway or 26S proteasome. Additionally, we provide insight into the physiological function of the relatively unexplored plant RBR-type E3 ligases, and through mutagenesis and biochemical assays we identified cysteine-361 in RFA4 as the putative active site cysteine, which is a distinctive feature of RBR-type E3 ligases. Endogenous levels of PYR1 and PYL4 ABA receptors were higher in the rfa1 rfa4 double mutant than in wild-type plants. UBC26 was identified as the cognate nuclear E2 enzyme that interacts with the RFA4 E3 ligase and forms UBC26-RFA4-receptor complexes in nuclear speckles. Loss-of-function ubc26 alleles and the rfa1 rfa4 double mutant showed enhanced sensitivity to ABA and accumulation of ABA receptors compared with the wild type. Together, our results reveal a sophisticated mechanism by which ABA receptors are targeted by ubiquitin at different subcellular locations, in which the complexity of the ABA receptor family is mirrored in the partner RBR-type E3 ligases.

Silicon-mediated alleviation of combined salinity and cadmium stress in date palm (Phoenix dactylifera L.) by regulating physio-hormonal alteration

We investigated the physio-molecular effects of separate and combined cadmium (Cd; 200 μM) and salinity (NaCl; 100 mM) stress on date palm during silicon (Si; 1.0 mM) applications. The results showed that exogenous Si led to significant improvements in plant growth, as well as physiology when compared with non-Si-treated seedling under stressed (Cd/NaCl) conditions. Interestingly, Si application led to lower metal (Cd) uptake and enhanced plant macronutrient uptake under combined stress, in turn, alleviating the combined salinity- and Cd-induced oxidative stress by lowering the lipid peroxidation rate, and peroxidase and catalase activities. Furthermore, ascorbate peroxidase level and the cytosolic Cu/Zn superoxide dismutase expression were significantly enhanced by Si application under combined stress. We further analyzed the effect of Si on modulation of stress-related hormonal crosstalk. Si markedly downregulated endogenous salicylic acid, jasmonic acid, and abscisic acid under NaCl stress and combined NaCl–Cd stress. However, during Cd toxicity alone, Si showed varying accumulation of these phytohormones. The results suggest that hindering the Cd uptake and enhancing silicon accumulation ultimately led to improvement of biomass and efficiency of the antioxidant system for alleviating combined stress. Moreover, higher transcript accumulation of PROLINE TRANSPORTER 2 and GAPDH and downregulation of ABA RECEPTOR by Si treatment under combined stress in date palm seedlings indicate the stress-ameliorative role of Si. The study provides evidence of the positive influence of Si on alleviating the combined toxicity of Cd and NaCl in date palm and can be further extended for field trials in Cd- and salinity-affected areas.

Sustainable bioenergy for climate mitigation: developing drought-tolerant trees and grasses.

Bioenergy crops are central to climate mitigation strategies that utilize biogenic carbon, such as BECCS (bioenergy with carbon capture and storage), alongside the use of biomass for heat, power, liquid fuels and, in the future, biorefining to chemicals. Several promising lignocellulosic crops are emerging that have no food role - fast-growing trees and grasses - but are well suited as bioenergy feedstocks, including Populus, Salix, Arundo, Miscanthus, Panicum and Sorghum. These promising crops remain largely undomesticated and, until recently, have had limited germplasm resources. In order to avoid competition with food crops for land and nature conservation, it is likely that future bioenergy crops will be grown on marginal land that is not needed for food production and is of poor quality and subject to drought stress. Thus, here we define an ideotype for drought tolerance that will enable biomass production to be maintained in the face of moderate drought stress. This includes traits that can readily be measured in wide populations of several hundred unique genotypes for genome-wide association studies, alongside traits that are informative but can only easily be assessed in limited numbers or training populations that may be more suitable for genomic selection. Phenotyping, not genotyping, is now the major bottleneck for progress, since in all lignocellulosic crops studied extensive use has been made of next-generation sequencing such that several thousand markers are now available and populations are emerging that will enable rapid progress for drought-tolerance breeding. The emergence of novel technologies for targeted genotyping by sequencing are particularly welcome. Genome editing has already been demonstrated for Populus and offers significant potential for rapid deployment of drought-tolerant crops through manipulation of ABA receptors, as demonstrated in Arabidopsis, with other gene targets yet to be tested. Bioenergy is predicted to be the fastest-developing renewable energy over the coming decade and significant investment over the past decade has been made in developing genomic resources and in collecting wild germplasm from within the natural ranges of several tree and grass crops. Harnessing these resources for climate-resilient crops for the future remains a challenge but one that is likely to be successful.

New ‘drug’ against drought

As climate change threatens farmers with more frequent and longer-lasting droughts, researchers are exploring how to help. Back in 2013, Sean R. Cutler’s group at the University of California, Riverside, identified a compound that mimicked the plant hormone abscisic acid (ABA). When applied to some plants, it helped them retain water (Proc. Natl. Acad. Sci. U.S.A. 2013, DOI: 10.1073/pnas.1305919110). But that compound, quinabactin, did not work well on some key crops, including wheat and tomato. The researchers performed structural studies to dig into why that might be and found that while ABA itself can bind to its receptors at two positions, in some ABA receptors, quinabactin binds to only one position. Now, the Cutler team has combined virtual screening and structure-guided design to develop a new compound it calls opabactin, which binds at both positions (Science 2019, DOI: 10.1126/science.aaw8848). The researchers tested opabactin on tomato and wheat plants in the

Abscisic Acid Receptors Modulate Metabolite Levels and Phenotype in Arabidopsis Under Normal Growing Conditions.

Abscisic acid (ABA) is a vital phytohormone that accumulates in response to various biotic and abiotic stresses, as well as plant growth. In Arabidopsis thaliana, there are 14 members of the ABA receptor family, which are key positive regulators involved in ABA signaling. Besides reduced drought stress tolerance, the quadruple and sextuple mutants (pyr1pyl1pyl2pyl4 (1124) and pyr1pyl1pyl2pyl4pyl5pyl8 (112458) show abnormal growth phenotypes, such as decreases in yield and height, under non-stress conditions. However, it remains unknown whether ABA receptors mediate ABA signaling to regulate plant growth and development. Here, we showed the primary metabolite profiles of 1124, 112458 and wild-type (WT) plants grown under normal conditions. The metabolic changes were significantly different between ABA receptor mutants and WT. Guanosine, for the biosynthesis of cyclic guanosine 3′,5′-monophosphate (cGMP), is an important second messenger that acts to regulate the level of ABA. In addition, other amino acids were increased in the 112458 mutant, including proline. These results, together with phenotype analysis, indicated that ABA receptors are involved in ABA signaling to modulate metabolism and plant growth under normal conditions.

Dynamic control of plant water use using designed ABA receptor agonists.

Drought causes crop losses worldwide, and its impact is expected to increase as the world warms. This has motivated the development of small-molecule tools for mitigating the effects of drought on agriculture. We show here that current leads are limited by poor bioactivity in wheat, a widely grown staple crop, and in tomato. To address this limitation, we combined virtual screening, x-ray crystallography, and structure-guided design to develop opabactin (OP), an abscisic acid (ABA) mimic with up to an approximately sevenfold increase in receptor affinity relative to ABA and up to 10-fold greater activity in vivo. Studies in Arabidopsis thaliana reveal a role of the type III receptor PYRABACTIN RESISTANCE-LIKE 2 for the antitranspirant efficacy of OP. Thus, virtual screening and structure-guided optimization yielded newly discovered agonists for manipulating crop abiotic stress tolerance and water use.

Discovery of Interacting Proteins of ABA Receptor PYL5 via Covalent Chemical Capture.

Abscisic acid (ABA) is a key phytohormone with diverse functions in plants, and its signal transduction is mainly mediated by ABA receptors termed PYR/PYL/RCARs (hereafter referred to as PYLs) through the PYLs-PP2Cs-SnRK2s regulatory systems. However, the model failed to account for the roles of some important known regulators of ABA physiology. Given the central role of PYLs in ABA signal transduction, we therefore speculated that ABA receptors PYLs might be involved in regulatory pathways other than PP2Cs. Thus, a comprehensive analysis of PYLs-interacting partners could greatly facilitate the identification of unknown regulatory pathways, advancing our knowledge of the ABA signaling mechanism. Herein, we present a strategy involving covalent chemical capture coupled with HPLC-MS/MS analysis, to profile PYL5-interacting partners in plant cell lysates. With this strategy, three new PYL5-interacting partners, ubiquitin receptor RAD23C, COP9 signalosome complex subunit 1 (CSN1), and cyclase-associated protein 1 (CAP1), along with their key binding sites with PYL5 were identified. Among these proteins, CAP1 was verified to interact with PYL5 both in vitro and in vivo. The discovery of a new PYL5 binding partner showed the versatility of covalent chemical cross-linking and laid the foundation for future efforts to further elucidate the ABA signaling mechanism.

Wheat F-box Protein TaFBA1 Positively Regulates Plant Drought Tolerance but Negatively Regulates Stomatal Closure.

The phytohormone abscisic acid (ABA) regulates plant growth and development, as well as responses to various stresses, such as salt and drought. The wheat TaFBA1 gene, which encodes an F-box protein, was previously identified in our laboratory by homologous cloning. We previously found that TaFBA1 expression was induced by ABA and drought stress. In this study, wild-type (WT), TaFBA1 over-expressing (OEs), TaFBA1 homologous gene mutants, and TaFBA1 recovery (Rs) Arabidopsis plants were used. We found that the germination rate, the cotyledon greening rate, the root length, and the photosynthetic performance of TaFBA1 OE plants were better than those of WT under drought and ABA conditions, but mutant plants showed the opposite trend, and overexpression of TaFBA1 in mutants can recover their phenotype. In addition, TaFBA1 was found to be a negative regulator of ABA-induced stoma movement; mRNA transcription of certain ABA signaling-related genes was lower in TaFBA1 OE plants than in WT plants following ABA treatment. Further, we found that TaFBA1 can interact with RCAR1 (an ABA receptor) and ABI5. BiFC assay showed that TaFBA1 may interact with RCAR1 in the plasma membrane. In addition, accumulation of ROS and MDA in TaFBA1 OE plants was lower than that in the WT plants after ABA and drought treatments. Based on these results, we suggest that TaFBA1-regulated ABA insensitivity may be dependent on regulating ABA-mediated gene expression through interacting with RCAR1 and ABI5. Increased antioxidant competence and decreased ROS accumulation may be an important mechanism that underlies improved drought tolerance in TaFBA1 OE plants.

Responses of PYR/PYL/RCAR ABA Receptors to Contrasting stresses, Heat and Cold in Arabidopsis

ABSTRACTPlants growing in natural habitats have evolved a wide range of mechanisms to copy with environmental challenging, including biotic and abiotic stresses. Abiotic stresses-induced increases in Abscisic acid (ABA) levels in plants suffering from stresses, including drought, cold or heat stress. To explore the function of the core components in ABA signaling, we used the overexpression of RCARs transgenic plants to expose in heat or cold stress. In this study, overexpression of RCAR12 or RCAR13 (R12-OE or R13-OE) transgenic plants had higher germination and survival rate than the wild-type (WT) Arabidopsis, indicating that they are both positively responsive to the high temperature. And the heat shock genes HSP18.2 and HSP70 were significantly induced by RCAR12 or RCAR13. Further, the results inferred that the over-expression of RCAR12 or RCAR13 could tolerance the cold stress, through induction CBFs expressions, the cold-responsive genes when plants were challenged the cold tress. And when complementation of RCAR12 to the 1124 mutant (R12:1124), the results indicated that RCAR12 could recover the insensitivity of 1124 to heat and cold stresses. Hence, we propose that RCAR12 and RCAR13, the ABA receptors, may play the positive roles in regulating the extreme temperature, including cold and high temperature in Arabidopsis.

Comprehensive survey of the VxGΦL motif of PP2Cs from Oryza sativa reveals the critical role of the fourth position in regulation of ABA responsiveness

Regulation of abscisic acid (ABA) signaling is crucial in balancing responses to abiotic stresses and retaining growth in planta. An ABA receptor (PYL/RCAR) and a protein phosphatase (PP2C), a co-receptor, form a complex upon binding to ABA. Previously we reported that the second and fourth positions in the VxGΦL motif of PP2Cs from Oryza sativa are critical in the interaction of PP2Cs with PYL/RCARs. Considering substantial effects of the VxGΦL motif on ABA signaling outputs, further comprehensive characterization of residues in the second and fourth positions are required. Here we surveyed the second and fourth positions of the VxGΦL motif by combination of biochemical, structural and physiological analyses. We found that the fourth position of the VxGΦL motif, highly conserved to small hydrophobic residues, was a key determinant of the OsPP2C50:OsPYL/RCAR interactions across subfamilies. Large hydrophobic or any hydrophilic residues in the fourth position abrogated ABA responsiveness. Analysis of crystal structures of OsPP2C50 mutants, S265L/I267V ("LV"), I267L ("SL") and I267W ("SW"), in complex with ABA and OsPYL/RCAR3, along with energy calculation of the complexes, uncovered that a bulky hydrophobic residue in the fourth position of the VxGΦL motif pushed away side chains of nearby residues, conferring side-chain rotameric energy stress. Hydrophilic residues in this position imposed solvation energy stress to the PP2C:PYL/RCAR complex. Germination and gene expression analyses corroborated that OsPP2C50 AS and AK mutants modulated ABA responsiveness in Arabidopsis. Our results suggest that ABA responsiveness could be fine-tuned by the fourth position of the VxGΦL motif on PP2Cs. KEY MESSAGE: We comprehensively surveyed the VxGΦL motif to find that the fourth position, highly conserved to small hydrophobic residues, was critical in regulating ABA responsiveness.

In silico study revealed major conserve architectures and novel features of pyrabactin binding to Oryza sativa ABA receptors compare to the Arabidopsis thaliana

Enhancing water use efficiency (WUE) of crops in irrigated agriculture and drought tolerance in rain-fed agriculture is the major goal for sustaining and enhancing agricultural productivity in the future. The phytohormone abscisic acid (ABA) signaling pathway is a major target for the agronomic management of WUE and genetic improvement of drought tolerance in crops. The START domain proteins PYRABACTIN RESISTANCE1 (PYR1)/PYR1-like (PYL)/Regulatory Components of ABA Receptors (RCARs) of the model plant Arabidopsis thaliana have been characterized as bona fide ABA receptors (ABARs). ABA signaling pathway can be activated or repressed by using specific agonist and antagonist against ABAR and therefore, can be used to control ABA-mediated physiological changes in plants. In the present work, we have reported the 3 D structure models of three ABARs (OsPYL1-3) from drought-tolerant Indica rice N22 (Oryza sativa L. sp. Indica cv N22) in apo- and ligand-bound conformations developed using comparative modeling techniques. Subsequently, these models were used in docking study to investigate the binding mode of known ABAR agonists and antagonists. Further, molecular dynamics studies on the selected systems verified the residues involved in protein-ligand interactions. The study identified the important ligand-binding features for the future development of specific agonists/antagonists to modulate the ABA activity in O. sativa and provides in silico models for designing and virtual screening to identify potent ABA receptor ligands.Communicated by Ramaswamy H. Sarma

Abscisic Acid Derivatives with Different Alkyl Chain Lengths Activate Distinct Abscisic Acid Receptor Subfamilies.

The plant hormone abscisic acid (ABA) regulates the development of various plant organs including seeds, roots, and fruits, and significantly contributes to abiotic stress responses, especially to drought. Since recent climate changes are adversely affecting crop cultivation, enhancement of plant stress tolerance by regulation of ABA signaling would be an important strategy. In the plant genome, ABA receptors are encoded by multiple genes constituting three subfamilies; however, functional differences among them remain unclear. To enhance desired effects of ABA, the biological functions of the receptor family warrant clarification. This study aimed to determine the functional differences among ABA receptors in plants. We screened small-molecule ligands binding to specific receptors, using a chemical array. In vitro evaluation of hit compounds using 11 Arabidopsis ABA receptors revealed that (+)-3'-alkyl ABAs served as agonists for different receptors depending on the length of their 3'-alkyl chains. Combinatorial in vitro and physiological effects of these compounds on the stomata, seeds, and seedlings indicated that, along with subfamily III, receptors of subfamily II are important to induce strong drought responses. Among (+)-3'-alkyl ABAs assessed herein, (+)-3'-butyl ABA induced a transcriptional response and stomatal closure but only slightly inhibited seed germination and growth, suggesting that it enhances drought tolerance. In silico docking simulation and site-directed mutagenesis revealed the amino acid residues contributing to the selective agonist activity of the (+)-3'-alkyl ABAs. These results provide novel insights into the structure and biological effects of 3'-derivatives of ABA and a basis for agrochemical development.

PYL9 is involved in the regulation of ABA signaling during tomato fruit ripening.

Abscisic acid (ABA) regulates fruit ripening, yet little is known about the exact roles of ABA receptors in fruit. In this study, we reveal the role of SlPYL9, a tomato pyrabactin resistance (PYR)/pyrobactin resistance-like (PYL)/regulatory component of ABA receptors (RCAR) protein, as a positive regulator of ABA signaling and fruit ripening. SlPYL9 inhibits protein phosphatase-type 2C (PP2C2/6) in an ABA dose-dependent way, and it interacts physically with SlPP2C2/3/4/5 in an ABA-dependent manner. Expression of SlPYL9 was observed in the seeds, flowers, and fruits. Overexpression and suppression of SlPYL9 induced a variety of phenotypes via altered expression of ABA signaling genes (SlPP2C1/2/9, SlSnRK2.8, SlABF2), thereby affecting expression of ripening-related genes involved in ethylene release and cell wall modification. SlPYL9-OE/RNAi plants showed a typical ABA hyper-/hypo-sensitive phenotype in terms of seed germination, primary root growth, and response to drought. Fruit ripening was significantly accelerated in SlPYL9-OE by 5–7 d as a result of increased endogenous ABA accumulation and advanced release of ethylene compared with the wild-type. In the SlPYL9-RNAi lines, fruit ripening was delayed, mesocarp thickness was enhanced, and petal abscission was delayed compared with the wild-type, resulting in conical/oblong and gourd-shaped fruits. These results suggest that SlPYL9 is involved in ABA signaling, thereby playing a role in the regulation of flower abscission and fruit ripening in tomato.

The role of Arabidopsis ABA receptors from the PYR/PYL/RCAR family in stomatal acclimation and closure signal integration.

Stomata are microscopic pores found on the surfaces of leaves that act to control CO2 uptake and water loss. By integrating information derived from endogenous signals with cues from the surrounding environment, the guard cells, which surround the pore, 'set' the stomatal aperture to suit the prevailing conditions. Much research has concentrated on understanding the rapid intracellular changes that result in immediate changes to the stomatal aperture. In this study, we look instead at how stomata acclimate to longer timescale variations in their environment. We show that the closure-inducing signals abscisic acid (ABA), increased CO2, decreased relative air humidity and darkness each access a unique gene network made up of clusters (or modules) of common cellular processes. However, within these networks some gene clusters are shared amongst all four stimuli. All stimuli modulate the expression of members of the PYR/PYL/RCAR family of ABA receptors. However, they are modulated differentially in a stimulus-specific manner. Of the six members of the PYR/PYL/RCAR family expressed in guard cells, PYL2 is sufficient for guard cell ABA-induced responses, whereas in the responses to CO2, PYL4 and PYL5 are essential. Overall, our work shows the importance of ABA as a central regulator and integrator of long-term changes in stomatal behaviour, including sensitivity, elicited by external signals. Understanding this architecture may aid in breeding crops with improved water and nutrient efficiency.

Identification of recombinant AtPYL2, an abscisic acid receptor, in E. coli using a substrate-derived bioactive small molecule, a biotin linker with alkyne and amino groups, and a protein cross-linker.

Target protein identification of bioactive small molecules is one of the most important research in forward chemical genetics. The affinity chromatography technique to use a resin bound with a small molecule is often used for identification of a target protein of a bioactive small molecule. Here we report a new method to isolate a protein targeted with a bioactive small molecule using a biotin linker with alkyne and amino groups, protein cross-linker containing disulfide bond, and a bioactive small molecule with an azido group (azido probe). After an azido probe is associated with a target protein, the complex of a target protein and azido probe is covalently bound through the biotin linker by azide-alkyne Huisgen cycloaddition and protein cross-linker containing disulfide bond. This ternary complex is immobilized on an affinity matrix with streptavidin, and then the target protein is selectively eluted with a buffer containing a reducing agent for cleavage of disulfide bonds. This method uses a probe having an azido group, which a small functional group, and has the possibility to be a solution strategy to overcome the hindrance of a functional group introduced into the probe that reduces association a target protein. The effectiveness of the method in this study was shown using linker 1, 3'-azidoabscisic acid 3, and protein cross-linker containing a disulfide bond (DTSSP 5).

The ABA receptor-like gene VyPYL9 from drought-resistance wild grapevine confers drought tolerance and ABA hypersensitivity in Arabidopsis

The PYR/PYL/RCAR (hereafter referred to PYLs) proteins, which act as abscisic acid (ABA) receptors, have been reported to play a crucial role in response to drought stress. In this study, we used a drought-resistant accession of the wild Chinese grapevine, Vitis yeshanensis-’Yanshan-1’, as a model to understand the function of PYLs in grape and other fruit crops. We cloned six PYL genes from V. yeshanensis. Phylogenetic analysis showed that the encoded proteins could be classified into three subfamilies. All six grapevine PYL proteins contained conserved amino acids at positions implicated in ‘gate-and-latch’ mechanism of ABA binding and in the interaction with PP2C proteins. Gene expression profiles of VyPYLs in various tissues of ‘Yanshan-1’ plants revealed they were relatively strongly expressed in old leaves compared to young leaves. Analysis of the subcellular localization of the VyPYL proteins showed that they were localized in the cytoplasm and the nucleus. Yeast two-hybrid assays revealed that six VyPYL proteins selectively interacted with three VyPP2C proteins in an ABA-dependent or ABA-independent manner. Expression of one of the genes, VyPYL9, was induced by exogenous ABA and drought conditions. Heterologous expression of VyPYL9 in Arabidopsis enhanced ABA sensitivity during seed germination and primary root growth. Moreover, VyPYL9-expressing Arabidopsis plants showed enhanced drought tolerance, and this was associated with decreased water loss and increased activity of antioxidant defense systems. In addition, we found that ABA promoted leaf senescence in Arabidopsis plants expressing VyPYL9. Taken together, our results suggest that VyPYL9 may be involved in response of grapevine to drought stress via the ABA signaling pathway. VyPYL9-expressing Arabidopsis plants showed enhanced drought tolerance, and this was associated with decreased water loss and increased activity of antioxidant defense systems. In addition, we found that ABA promoted leaf senescence in Arabidopsis plants expressing VyPYL9.

GmWRKY54 improves drought tolerance through activating genes in abscisic acid and Ca2+ signaling pathways in transgenic soybean.

WRKY transcription factors play important roles in response to various abiotic stresses. Previous study have proved that soybean GmWRKY54 can improve stress tolerance in transgenic Arabidopsis. Here, we generated soybean transgenic plants and further investigated roles and biological mechanisms of GmWRKY54 in response to drought stress. We demonstrated that expression of GmWRKY54, driven by either a constitutive promoter (pCm) or a drought-induced promoter (RD29a), confers drought tolerance. GmWRKY54 is a transcriptional activator and affects a large number of stress-related genes as revealed by RNA sequencing. Gene ontology (GO) enrichment and co-expression network analysis, together with measurement of physiological parameters, supported the idea that GmWRKY54 enhances stomatal closure to reduce water loss, and therefore confers drought tolerance in soybean. GmWRKY54 directly binds to the promoter regions of genes including PYL8, SRK2A, CIPK11 and CPK3 and activates them. Therefore GmWRKY54 achieves its function through abscisic acid (ABA) and Ca2+ signaling pathways. It is valuable that GmWRKY54 activates an ABA receptor and an SnRK2 kinase in the upstream position, unlike other WRKY proteins that regulate downstream genes in the ABA pathway. Our study revealed the role of GmWRKY54 in drought tolerance and further manipulation of this gene should improve growth and production in soybean and other legumes/crops under unfavorable conditions.

Rebuilding core abscisic acid signaling pathways of Arabidopsis in yeast

The phytohormone abscisic acid (ABA) regulates plant responses to abiotic stress, such as drought and high osmotic conditions. The multitude of functionally redundant components involved in ABA signaling poses a major challenge for elucidating individual contributions to the response selectivity and sensitivity of the pathway. Here, we reconstructed single ABA signaling pathways in yeast for combinatorial analysis of ABA receptors and coreceptors, downstream‐acting SnRK2 protein kinases, and transcription factors. The analysis shows that some ABA receptors stimulate the pathway even in the absence of ABA and that SnRK2s are major determinants of ABA responsiveness by differing in the ligand‐dependent control. Five SnRK2s, including SnRK2.4 known to be active under osmotic stress in plants, activated ABA‐responsive transcription factors and were regulated by ABA receptor complexes in yeast. In the plant tissue, SnRK2.4 and ABA receptors competed for coreceptor interaction in an ABA‐dependent manner consistent with a tight integration of SnRK2.4 into the ABA signaling pathway. The study establishes the suitability of the yeast system for the dissection of core signaling cascades and opens up future avenues of research on ligand‐receptor regulation.

Arabidopsis ALIX Regulates Stomatal Aperture and Turnover of Abscisic Acid Receptors.

The plant endosomal trafficking pathway controls the abundance of membrane-associated soluble proteins, as shown for abscisic acid (ABA) receptors of the PYRABACTIN RESISTANCE1/PYR1-LIKE/REGULATORY COMPONENTS OF ABA RECEPTORS (PYR/PYL/RCAR) family. ABA receptor targeting for vacuolar degradation occurs through the late endosome route and depends on FYVE DOMAIN PROTEIN REQUIRED FOR ENDOSOMAL SORTING1 (FYVE1) and VACUOLAR PROTEIN SORTING23A (VPS23A), components of the ENDOSOMAL SORTING COMPLEX REQUIRED FOR TRANSPORT-I (ESCRT-I) complexes. FYVE1 and VPS23A interact with ALG-2 INTERACTING PROTEIN-X (ALIX), an ESCRT-III-associated protein, although the functional relevance of such interactions and their consequences in cargo sorting are unknown. In this study we show that Arabidopsis (Arabidopsis thaliana) ALIX directly binds to ABA receptors in late endosomes, promoting their degradation. Impaired ALIX function leads to altered endosomal localization and increased accumulation of ABA receptors. In line with this activity, partial loss-of-function alix-1 mutants display ABA hypersensitivity during growth and stomatal closure, unveiling a role for the ESCRT machinery in the control of water loss through stomata. ABA-hypersensitive responses are suppressed in alix-1 plants impaired in PYR/PYL/RCAR activity, in accordance with ALIX affecting ABA responses primarily by controlling ABA receptor stability. ALIX-1 mutant protein displays reduced interaction with VPS23A and ABA receptors, providing a molecular basis for ABA hypersensitivity in alix-1 mutants. Our findings unveil a negative feedback mechanism triggered by ABA that acts via ALIX to control the accumulation of specific PYR/PYL/RCAR receptors.

RNA-Seq identification of candidate defense genes by analyzing Mythimna separata feeding-damage induced systemic resistance in balsas teosinte.

Armyworm (Mythimna separata) is a destructive herbivore for maize. Balsas teosinte (Zea mays ssp. parviglumis), the direct wild ancestor of cultivated maize, has shown great potential to defend against herbivory. Here, based on armyworm bioassay, we compared responses of teosinte and B73 maize inbred during armyworm attack in their transcriptome profiles to elucidate the gene expression changes involved in teosinte responses to armyworm attack. The goal of this study was to identify novel resistance alleles that could serve as valuable resources for modern maize breeding. Our bioassay revealed that armyworm larvae grew less on teosinte than on maize. A follow-up transcriptomic comparison showed more down-regulated genes in maize B73 and similar numbers of up-regulated genes in both genotypes under armyworm attack. The up-regulated genes in teosinte were markedly enriched in MAPK cascade-mediated signaling pathway and phytohormone pathway. Gene ontology and Kyoto Encyclopedia of Genes and Genomes pathway analysis showed that phytohormones jasmonic acid, ethylene, salicylic acid and abscisic acid (ABA) were actively involved in armyworm resistance of teosinte plants, and so were transcription factors such as MYBs, WRKYs and TIFYs. Interestingly, teosinte also showed high regulation in three ABA receptor PYLs. Based on differential expression analysis, we identified 30 candidate defense-related genes in teosinte, which belong to 11 gene families and the majority of the genes were up-regulated, while some of them were nonresponsive in maize. This study demonstrates that teosinte showed more vigorous defense response than maize toward armyworm attack and might be a beneficial genetic resource to improve pest resistance in cultivated maize. © 2019 Society of Chemical Industry.