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The plant-specific VQ gene family participates in diverse physiological processes but little information is available on their role in leaf senescence. Here, we show that the VQ motif-containing proteins, Arabidopsis SIGMA FACTOR BINDING PROTEIN1 (SIB1) and SIB2 are negative regulators of abscisic acid (ABA)-mediated leaf senescence. Loss of SIB1 and SIB2 function resulted in increased sensitivity of ABA-induced leaf senescence. In contrast, overexpression of SIB1 significantly delayed this process. Moreover, biochemical studies revealed that SIBs interact with WRKY75 transcription factor. Loss of WRKY75 function decreased sensitivity to ABA-induced leaf senescence, while overexpression of WRKY75 significantly accelerated this process. Chromatin immunoprecipitation assays revealed that WRKY75 directly binds to the promoters of GOLDEN 2-LIKE1(GLK1) and GLK2, to repress their expression. SIBs repress the transcriptional function of WRKY75 and negatively regulate ABA-induced leaf senescence in a WRKY75-dependent manner. In contrast, WRKY75 positively modulates ABA-mediated leaf senescence in a GLK-dependent manner. In addition, SIBs inhibit WRKY75 function in ABA-mediated seed germination. These results demonstrate that SIBs can form a complex with WRKY75 to regulate ABA-mediated leaf senescence and seed germination., SIBs function as repressors of WRKY75 and regulate expression of GLKs in ABA-mediated leaf senescence and seed germination.

Fungal polypeptide-induced callose priming restricts the cell-to-cell movement of tobacco mosaic virus in N. benthamiana via the ABA biosynthetic pathway., The polypeptide extract of the dry mycelium of Penicillium chrysogenum (PDMP) can protect tobacco plants from tobacco mosaic virus (TMV), although the mechanism underlying PDMP-mediated TMV resistance remains unknown. In our study, we analysed a potential mechanism via RNA sequencing (RNA-seq) and found that the abscisic acid (ABA) biosynthetic pathway and β-1,3-glucanase, a callose-degrading enzyme, might play an important role in PDMP-induced priming of resistance to TMV. To test our hypothesis, we successfully generated a Nicotiana benthamiana ABA biosynthesis mutant and evaluated the role of the ABA pathway in PDMP-induced callose deposition during resistance to TMV infection. Our results suggested that PDMP can induce callose priming to defend against TMV movement. PDMP inhibited TMV movement by increasing callose deposition around plasmodesmata, but this phenomenon did not occur in the ABA biosynthesis mutant; moreover, these effects of PDMP on callose deposition could be rescued by treatment with exogenous ABA. Our results suggested that callose deposition around plasmodesmata in wild-type plants is mainly responsible for the restriction of TMV movement during the PDMP-induced defensive response to TMV infection, and that ABA biosynthesis apparently plays a crucial role in PDMP-induced callose priming for enhancing defence against TMV.

Centro de Biotecnología y Genómica de Plantas (CBGP), The evolutionary success of plants relies to a large extent on their extraordinary ability to adapt to changes in their environment. These adaptations require that plants balance their growth with their stress responses. Plant hormones are crucial mediators orchestrating the underlying adaptive processes. However, whether and how the growth-related hormone auxin and the stress-related hormones jasmonic acid, salicylic acid, and abscisic acid (ABA) are coordinated remains largely elusive. Here, we analyse the physiological role of AMIDASE 1 (AMI1) in Arabidopsis plant growth and its possible connection to plant adaptations to abiotic stresses. AMI1 contributes to cellular auxin homeostasis by catalysing the conversion of indole-acetamide into the major plant auxin indole-3-acetic acid. Functional impairment of AMI1 increases the plant's stress status rendering mutant plants more susceptible to abiotic stresses. Transcriptomic analysis of ami1 mutants disclosed the reprogramming of a considerable number of stress-related genes, including jasmonic acid and ABA biosynthesis genes. The ami1 mutants exhibit only moderately repressed growth but an enhanced ABA accumulation, which suggests a role for AMI1 in the crosstalk between auxin and ABA. Altogether, our results suggest that AMI1 is involved in coordinating the trade-off between plant growth and stress responses, balancing auxin and ABA homeostasis., This research was supported by grants from the German Research Foundation (DFG, SFB480/A10) and the Spanish Ministry of Economy, Industry and Competitiveness (MINECO, BFU2017-82826-R to SP and a grant from the Swedish Research Council (VR) to HA. JM was supported by the ‘Severo Ochoa Program for Centers of Excellence in R&D’ from the Agencia Estatal de Investigación of Spain, grant SEV-2016-0672 (2017-2021) to the CBGP., 17 Pág.

USB1, an exoribonuclease catalyzing U6 production for spliceosome assembly, negatively regulates, cooperatively with SOAR1, ABA signaling in early seedling development, probably by affecting spliceosome assembly., Signaling by the phytohormone abscisic acid (ABA) involves pre-mRNA splicing, a key process of post-transcriptional regulation of gene expression. However, the regulatory mechanism of alternative pre-mRNA splicing in ABA signaling remains largely unknown. We previously identified a pentatricopeptide repeat protein SOAR1 (suppressor of the ABAR-overexpressor 1) as a crucial player downstream of ABAR (putative ABA receptor) in ABA signaling. In this study, we identified a SOAR1 interaction partner USB1, which is an exoribonuclease catalyzing U6 production for spliceosome assembly. We reveal that together USB1 and SOAR1 negatively regulate ABA signaling in early seedling development. USB1 and SOAR1 are both required for the splicing of transcripts of numerous genes, including those involved in ABA signaling pathways, suggesting that USB1 and SOAR1 collaborate to regulate ABA signaling by affecting spliceosome assembly. These findings provide important new insights into the mechanistic control of alternative pre-mRNA splicing in the regulation of ABA-mediated plant responses to environmental cues.

Water stress is a primary trigger for reducing grain number per spike in wheat during the reproductive period. However, under stress conditions, the responses of plant organs and the interactions between them at the molecular and physiological levels remain unclear. In this study, when water stress occurred at the young microspore stage, RNA-seq data indicated that the spike had 970 differentially expressed genes, while the stem, comprising the two internodes below the spike (TIS), had 382. Abscisic acid (ABA) signal transduction genes were down-regulated by water stress in both these tissues, although to a greater extent in the TIS than in the spike. A reduction in sucrose was observed, and was accompanied by increases in cell wall invertase (CWIN) and sucrose:sucrose 1-fructosyl-transferase (1-SST) activities. Hexose and fructan were increased in the TIS but decreased in the spike. ABA was increased in the spike and TIS, and showed significant positive correlation with CWIN and 1-SST activities in the TIS. Overall, our results suggest that water stress induces the conversion of sucrose to hexose by CWIN, and to fructan by 1-SST, due to increased down-regulation of ABA signal transduction related-genes in the TIS; this leads to deficient sucrose supply to the spike and a decrease in grain number., Deficient sucrose and weak expression of ABA signal transduction genes in the spike may be the major reasons for the decrease in grain number under water stress.

PPC2, together with ABI5 and ABA, is involved in plant acclimation to low CO2 through linking photorespiratory metabolism with primary metabolism., Phosphoenolpyruvate carboxylase (PEPC) plays a pivotal role in the photosynthetic CO2 fixation of C4 plants. However, the functions of PEPCs in C3 plants are less well characterized, particularly in relation to low atmospheric CO2 levels. Of the four genes encoding PEPC in Arabidopsis, PPC2 is considered as the major leaf PEPC gene. Here we show that the ppc2 mutants suffered a growth arrest when transferred to low atmospheric CO2 conditions, together with decreases in the maximum efficiency of PSII (Fv/Fm) and lower levels of leaf abscisic acid (ABA) and carbohydrates. The application of sucrose, malate, or ABA greatly rescued the growth of ppc2 lines under low CO2 conditions. Metabolite profiling analysis revealed that the levels of glycine and serine were increased in ppc2 leaves, while the abundance of photosynthetic metabolites was decreased under these conditions. The transcript levels of encoding enzymes involved in glycine or serine metabolism was decreased in ppc2 in an ABI5-dependent manner. Like the ppc2 mutants, abi5-1 mutants had lower photosynthetic rates and Fv/Fm compared with the wild type under photorespiratory conditions (i.e. low CO2 availability). However, the growth of these mutants was similar to that of the wild type under non-photorespiratory (low O2) conditions. The constitutive expression of ABI5 prevented the growth arrest of ppc2 lines under low CO2 conditions. These findings demonstrate that PPC2 plays an important role in the acclimation of Arabidopsis plants to low CO2 availability by linking photorespiratory metabolism to primary metabolism, and that this is mediated, at least in part, through ABA- and ABI5-dependent processes.

Abscisic acid plays a positive role in regulating the OsSAPK9–OsbZIP20 pathway and elevating antioxidant and GS/GOGAT activities in rice to increase tolerance to high-NH4+ stress., Ammonium (NH4+) is one of the principal nitrogen (N) sources in soils, but is typically toxic already at intermediate concentrations. The phytohormone abscisic acid (ABA) plays a pivotal role in responses to environmental stresses. However, the role of ABA under high-NH4+ stress in rice (Oryza sativa L.) is only marginally understood. Here, we report that elevated NH4+ can significantly accelerate tissue ABA accumulation. Mutants with high (Osaba8ox) and low levels of ABA (Osphs3-1) exhibit elevated tolerance or sensitivity to high-NH4+ stress, respectively. Furthermore, ABA can decrease NH4+-induced oxidative damage and tissue NH4+ accumulation by enhancing antioxidant and glutamine synthetase (GS)/glutamate synthetasae (GOGAT) enzyme activities. Using RNA sequencing and quantitative real-time PCR approaches, we ascertain that two genes, OsSAPK9 and OsbZIP20, are induced both by high NH4+ and by ABA. Our data indicate that OsSAPK9 interacts with OsbZIP20, and can phosphorylate OsbZIP20 and activate its function. When OsSAPK9 or OsbZIP20 are knocked out in rice, ABA-mediated antioxidant and GS/GOGAT activity enhancement under high-NH4+ stress disappear, and the two mutants are more sensitive to high-NH4+ stress compared with their wild types. Taken together, our results suggest that ABA plays a positive role in regulating the OsSAPK9–OsbZIP20 pathway in rice to increase tolerance to high-NH4+ stress.

Transcription factors (TFs) help plants respond to environmental stresses by regulating gene expression. Up till now, studies on the MYB family of TFs have mainly focused on the highly abundant R2R3-subtype. While the less well-known 1R-subtype has been generally shown to enhance abscisic acid (ABA) sensitivity by acting as transcriptional activators, the mechanisms of their functions are unclear. Here we identified an ABA sensitivity-associated gene from soybean, ABA-Sensitive 1 (GmABAS1), of the 1R-subtype of MYB. Using the GFP-GmABAS1 fusion protein, we demonstrated that GmABAS1 is localized in the nucleus, and with yeast reporter systems, we showed that it is a transcriptional repressor. We then identified the target gene of GmABAS1 to be Glyma.01G060300, an annotated ABI five-binding protein 3 and showed that GmABAS1 binds to the promoter of Glyma.01G060300 both in vitro and in vivo. Furthermore, Glyma.01G060300 and GmABAS1 exhibited reciprocal expression patterns under osmotic stress, inferring that GmABAS1 is a transcriptional repressor of Glyma.01G060300. As a further confirmation, AtAFP2, an orthologue of Glyma.01G060300, was down-regulated in GmABAS1-transgenic Arabidopsis thaliana, enhancing the plant’s sensitivity to ABA. This is the first time a 1R-subtype of MYB from soybean has been reported to enhance ABA sensitivity by acting as a transcriptional repressor., ABAS1 negatively regulates the expression of a soybean AFP3 homologue and enhances ABA sensitivity. This is the first report on the transcriptional repressor activity of a 1R-subtype MYB from soybean.

Combined heat and drought stress can enhance phytohormone-dependent defenses of wheat against aphids and decrease aphid fitness, providing new insights on plant–insect interactions under conditions of complex stress., Plants are routinely subjected simultaneously to different abiotic and biotic stresses, such as heat, drought, and insect infestation. Plant–insect interactions in such complex stress situations are poorly understood. We evaluated the performance of the grain aphid (Sitobion avenae) in wheat (Triticum aestivum L.) exposed to a combination of heat and drought stresses. We also performed assays of the relative water content, nutritional quality, and responses of phytohormone signaling pathways. Lower relative water content and accumulation of soluble sugars and amino acids were observed in plants exposed to combined heat and drought stress. These conditions increased abscisic acid levels in the absence of aphids, as well as leading to higher levels of jasmonate-dependent transcripts. The grain aphid infestation further increased abscisic acid levels and the abundance of jasmonic acid- and salicylic acid-dependent defenses under the combined stress conditions. Aphids reared on plants grown under drought stress alone showed lower net reproductive rates, intrinsic rates of increase, and finite rates of increase compared with aphids reared on plants in the absence of stress. The heat-treated plants also showed a decreased aphid net reproductive rate. These findings demonstrate that exposure to a combination of stresses enhances plant defense responses against aphids as well as altering nutritional quality.

Shading of the soybean mother plant during seed development promotes subsequent germination by mediating the biosynthesis of pro-anthocyanidins, fatty acids, and the phytohormones ABA and GA., The effect of shading during seed development on subsequent germination remains largely unknown. In this study, two soybean (Glycine max) seed production systems, monocropping (MC) and maize–soybean intercropping (IC), were employed to examine the effects of shading of the mother plant on subsequent seed germination. Compared to the MC soybean seeds, which received light, the developing IC seeds were exposed to shade resulting from the taller neighboring maize plants. The IC seeds germinated faster than the MC seeds, although there was no significant difference in the thickness of the seed coat. The concentration of soluble pro-anthocyanidin in the IC seed coat was significantly lower than that in the MC seed coat. Changes in the concentrations of several types of fatty acids in IC seeds were also observed, the nature of which were consistent with the effect on germination. The expression levels of genes involved in abscisic acid (ABA) biosynthesis were down-regulated in IC seeds, while the transcription levels of the genes related to gibberellin (GA) biosynthesis were up-regulated. This was consistently reflected in decreased ABA concentrations and increased active GA4 concentrations in IC seeds, resulting in an increased GA4/ABA ratio. Our results thus indicated that shading of the mother plant during seed development in soybean promoted subsequent germination by mediating the biosynthesis of pro-anthocyanidins, fatty acids, and phytohormones.

Iron (Fe) is vital for plant growth. Plants balance the beneficial and toxic effects of this micronutrient, and tightly control Fe uptake and allocation. Here, we review the role of the basic helix–loop–helix (bHLH) transcription factor FIT (FER-LIKE FE DEFICIENCY-INDUCED TRANSCRIPTION FACTOR) in Fe acquisition. FIT is not only essential, it is also a central regulatory hub in root cells to steer and adjust the rate of Fe uptake by the root in a changing environment. FIT regulates a subset of root Fe deficiency (–Fe) response genes. Based on a combination of co-expression network and FIT-dependent transcriptome analyses, we defined a set of FIT-dependent and FIT-independent gene expression signatures and co-expression clusters that encode specific functions in Fe regulation and Fe homeostasis. These gene signatures serve as markers to integrate novel regulatory factors and signals into the –Fe response cascade. FIT forms a complex with bHLH subgroup Ib transcription factors. Furthermore, it interacts with key regulators from different signaling pathways that either activate or inhibit FIT function to adjust Fe acquisition to growth and environmental constraints. Co-expression clusters and FIT protein interactions suggest a connection of –Fe with ABA responses and root cell elongation processes that can be explored in future studies., FIT-dependent gene signatures and FIT protein interactions highlight links to developmental and stress signaling, and suggest a connection with abscisic acid responses and root cell elongation.

Ethylene controls post-germination starch degradation and seedling growth, but not radicle protrusion, in wheat through spatiotemporal alteration of the balance between abscisic acid and gibberellin., This study aimed to gain insights into the molecular mechanisms underlying the role of ethylene in regulating germination and seedling growth in wheat by combining pharmacological, molecular, and metabolomics approaches. Our study showed that ethylene does not affect radicle protrusion but controls post-germination endospermic starch degradation through transcriptional regulation of specific α-amylase and α-glucosidase genes, and this effect is mediated by alteration of endospermic bioactive gibberellin (GA) levels, and GA sensitivity via expression of the GA signaling gene, TaGAMYB. Our data implicated ethylene as a positive regulator of embryo axis and coleoptile growth through transcriptional regulation of specific TaEXPA genes. These effects were associated with modulation of GA levels and sensitivity, through expression of GA metabolism (TaGA20ox1, TaGA3ox2, and TaGA2ox6) and signaling (TaGAMYB) genes, respectively, and/or the abscisic acid (ABA) level and sensitivity, via expression of specific ABA metabolism (TaNCED2 or TaCYP707A1) and signaling (TaABI3) genes, respectively. Ethylene appeared to regulate the expression of TaEXPA3 and thereby root growth through its control of coleoptile ABA metabolism, and root ABA signaling via expression of TaABI3 and TaABI5. These results show that spatiotemporal modulation of ABA/GA balance mediates the role of ethylene in regulating post-germination storage starch degradation and seedling growth in wheat.

In Arabidopsis leaves, the stress-induced production of abscisic acid from its inactive glucose conjugate involves dynamic changes in the endoplasmic reticulum., The phytohormone abscisic acid (ABA) is produced via a multistep de novo biosynthesis pathway or via single-step hydrolysis of inactive ABA-glucose ester (ABA-GE). The hydrolysis reaction is catalyzed by β-glucosidase (BG, or BGLU) isoforms localized to various organelles, where they become activated upon stress, but the mechanisms underlying this organelle-specific activation remain unclear. We investigated the relationship between the subcellular distribution and stress-induced activation of BGLU18 (BG1), an endoplasmic reticulum enzyme critical for abiotic stress responses, in Arabidopsis thaliana leaves. High BGLU18 levels were present in leaf petioles, primarily in endoplasmic reticulum bodies. These Brassicaceae-specific endoplasmic reticulum-derived organelles responded dynamically to abiotic stress, particularly drought-induced dehydration, by changing in number and size. Under stress, BGLU18 distribution shifted toward microsomes, which was accompanied by increasing BGLU18-mediated ABA-GE hydrolytic activity and ABA levels in leaf petioles. Under non-stress conditions, impaired endoplasmic reticulum body formation caused a microsomal shift of BGLU18 and increased its enzyme activity; however, ABA levels increased only under stress, probably because ABA-GE is supplied to the endoplasmic reticulum only under these conditions. Loss of BGLU18 delayed dehydration-induced ABA accumulation, suggesting that ABA-GE hydrolysis precedes the biosynthesis. We propose that dynamics of the endoplasmic reticulum modulate ABA homeostasis and abiotic stress responses by activating BGLU18-mediated ABA-GE hydrolysis.

The dormancy-associated MADS-box (DAM) genes PpDAM5 and PpDAM6 have been shown to play important roles in bud endodormancy; however, their molecular regulatory mechanism in peach is unclear. In this study, by use of yeast one-hybrid screening, we isolated a TEOSINTE BRANCHED1/CYCLOIDEA/PROLIFERATING CELL FACTOR transcription factor, PpTCP20, in the peach cultivar ‘Zhongyou 4’ (Prunus persica var. nectarina). The protein was localized in the nucleus and was capable of forming a homodimer. Electrophoretic mobility shift assays demonstrated that PpTCP20 binds to a GCCCR element in the promoters of PpDAM5 and PpDAM6, and transient dual luciferase experiments showed that PpTCP20 inhibited the expression of PpDAM5 and PpDAM6 as the period of the release of flower bud endodormancy approached. In addition, PpTCP20 interacted with PpABF2 to form heterodimers to regulate bud endodormancy, and the content of abscisic acid decreased with the release of endodormancy. PpTCP20 also inhibited expression of PpABF2 to regulate endodormancy. Taken together, our results suggest that PpTCP20 regulates peach flower bud endodormancy by negatively regulating the expression of PpDAM5 and PpDAM6, and by interacting with PpABF2, thus revealing a novel regulatory mechanism in a perennial deciduous tree., PpTCP20 controls the release of endodormancy in peach flower buds by negatively regulating the expression of PpDAM5 and PpDAM6, and by interacting with PpABF2.

Gene regulation is central for growth, development, and adaptation to environmental changes in all living organisms. Many genes are induced by environmental cues, and the expression of these inducible genes is often repressed under normal conditions. Here, we show that the SHINY2 (SHI2) gene is important for repressing salt-inducible genes and also plays a role in cold response. The shi2 mutant displayed hypersensitivity to cold, abscisic acid (ABA), and LiCl. Map-based cloning demonstrates that SHI2 encodes a DEAD- (Asp-Glu-Ala-Asp) box RNA helicase with similarity to a yeast splicing factor. Transcriptomic analysis of the shi2 mutant in response to cold revealed that the shi2 mutation decreased the number of cold-responsive genes and the magnitude of their response, and resulted in the mis-splicing of some cold-responsive genes. Under salt stress, however, the shi2 mutation increased the number of salt-responsive genes but had a negligible effect on mRNA splicing. Our results suggest that SHI2 is a component in a ready-for-transcription repressor complex important for gene repression under normal conditions, and for gene activation and transcription under stress conditions. In addition, SHI2 also serves as a splicing factor required for proper splicing of cold-responsive genes and affects 5' capping and polyadenylation site selection., The DEAD-box RNA helix SHI2 represses expression of salt-inducible genes and modulates splicing of cold-responsive genes. SHI2 participates in transcription and post-transcriptional processes in response to stress conditions.

Auxin herbicides trigger a rapid down-regulation of photosynthetic-related gene expression and an up-regulation of abscisic acid biosynthesis independent of other plant hormones in Erigeron., The perception pathway for endogenous auxin has been well described, yet the mode of action of synthetic auxin herbicides, used for >70 years, remains uncharacterized. We utilized transcriptomics and targeted physiological studies to investigate the unknown rapid response to synthetic auxin herbicides in the globally problematic weed species Erigeron canadensis. Synthetic auxin herbicide application consistently and rapidly down-regulated the photosynthetic machinery. At the same time, there was considerable perturbation to the expression of many genes related to phytohormone metabolism and perception. In particular, auxin herbicide application enhanced the expression of the key abscisic acid biosynthetic gene, 9-cis-epoxycarotenoid deoxygenase (NCED). The increase in NCED expression following auxin herbicide application led to a rapid biosynthesis of abscisic acid (ABA). This increase in ABA levels was independent of a loss of cell turgor or an increase in ethylene levels, both proposed triggers for rapid ABA biosynthesis. The levels of ABA in the leaf after auxin herbicide application continued to increase as plants approached death, up to >3-fold higher than in the leaves of plants that were drought stressed. We propose a new model in which synthetic auxin herbicides trigger plant death by the whole-scale, rapid, down-regulation of photosynthetic processes and an increase in ABA levels through up-regulation of NCED expression, independent of ethylene levels or a loss of cell turgor.

The closely related sulphonamide safeners, metcamifen and cyprosulfamide, were tested for their ability to protect rice from clodinafop-propargyl, a herbicide normally used in wheat. While demonstrating that both compounds were equally bioavailable in planta, only metcamifen prevented clodinafop from damaging seedlings, and this was associated with the enhanced detoxification of the herbicide. Transcriptome studies in rice cultures demonstrated that whereas cyprosulfamide had a negligible effect on gene expression over a 4 h exposure, metcamifen perturbed the abundance of 590 transcripts. Changes in gene expression with metcamifen could be divided into three phases, corresponding to inductions occurring over 30 min, 1.5 h and 4 h. The first phase of gene induction was dominated by transcription factors and proteins of unknown function, the second by genes involved in herbicide detoxification, while the third was linked to cellular homeostasis. Analysis of the inducible genes suggested that safening elicited similar gene families to those associated with specific biotic and abiotic stresses, notably those elicited by abscisic acid, salicylic acid, and methyl jasmonate. Subsequent experiments with safener biomarker genes induced in phase 1 and 2 in rice cell cultures provided further evidence of similarities in signalling processes elicited by metcamifen and salicylic acid., Protection against herbicide damage afforded by the safener metcamifen in rice is associated with a phased induction of genes involved in signalling, xenobiotic detoxification, and cell homeostasis.

Activation of the kiwifruit suberin biosynthesis gene AchnFHT is coordinately controlled via repression by AchnMYB4 and promotion by AchnABF2, AchnMYB41, and AchnMYB107., Suberin is a cell-wall biopolymer with aliphatic and aromatic domains that is synthesized in the wound tissues of plants in order to restrict water loss and pathogen infection. ω-hydroxyacid/fatty alcohol hydroxycinnamoyl transferase (FHT) is required for cross-linking of the aliphatic and aromatic domains. ABA is known to play a positive role in suberin biosynthesis but it is not known how it interacts with FHT. In this study, the kiwifruit (Actinidia chinensis) AchnFHT gene was isolated and was found to be localized in the cytosol. Transient overexpression of AchnFHT in leaves of Nicotiana benthamiana induced massive production of ferulate, ω-hydroxyacids, and primary alcohols, consistent with the in vitro ability of AchnFHT to catalyse acyl-transfer from feruloyl-CoA to ω-hydroxypalmitic acid and 1-tetradecanol. A regulatory function of four TFs (AchnABF2, AchnMYB4, AchnMYB41, and AchnMYB107) on AchnFHT was identified. These TFs localized in the nucleus and directly interacted with the AchnFHT promoter in yeast one-hybrid assays. Dual-luciferase analysis indicated that AchnABF2, AchnMYB41, and AchnMYB107 activated the AchnFHT promoter while AchnMYB4 repressed it. These findings were supported by the results of transient overexpression in N. benthamiana, in which AchnABF2, AchnMYB41, and AchnMYB107 induced expression of suberin biosynthesis genes (including FHT) and accumulation of suberin monomers, whilst AchnMYB4 had the opposite effect. Exogenous ABA induced the expression of AchnABF2, AchnMYB41, AchnMYB107, and AchnFHT and induced suberin monomer formation, but it inhibited AchnMYB4 expression. In addition, fluridone (an inhibitor of ABA biosynthesis) was found to counter the inductive effects of ABA. Activation of suberin monomer biosynthesis by AchnFHT was therefore controlled in a coordinated way by both repression of AchnMYB4 and promotion of AchnABF2, AchnMYB41, and AchnMYB107.

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., ABA receptor PYL9 plays a role in the regulation of flower abscission and fruit ripening in tomato by modulating ABA signaling components, which regulates expression of related genes.

Plant aquaporins and hormones regulate root water uptake in mildly stressed poplar together with the participation of fungal aquaporins from the ectomycorrhizal fungus Laccaria bicolor., The relatively better performance of mycorrhizal plants subjected to drought stress has commonly been linked to improved root water uptake through the fungal regulation of plant aquaporins and hormones. In this study, we examined the role of ectomycorrhizal fungi in plant water relations and plant hormonal balance under mild drought using split-root seedlings of Populus trichocarpa × deltoides either with or without inoculation with Laccaria bicolor. The root compartments where the drought treatment was applied had higher ABA and lower cytokinin tZR contents, and greater expression of the plant aquaporins PtPIP1;1, PtPIP1;2, PtPIP2;5, and PtPIP2;7. On the other hand, the presence of L. bicolor within the roots down-regulated PtPIP1;4, PtPIP2;3, and PtPIP2;10, and reduced the abundance of PIP2 proteins. In addition, expression of the fungal aquaporins JQ585595 and JQ585596 were positively correlated with root ABA content, while tZR content was positively correlated with PtPIP1;4 and negatively correlated with PtPIP2;7. The results demonstrate a coordinated plant–fungal system that regulates the different mechanisms involved in water uptake in ectomycorrhizal poplar plants.

ZmPTF1 regulates drought tolerance in maize by promoting root development and ABA synthesis, by binding to the G-box in the promoter and activating the expression of NCEDs, CBF4, NAC081, and NAC30., Drought stress is the most important environmental stress limiting maize production. ZmPTF1, a phosphate starvation-induced basic helix-loop-helix (bHLH) transcription factor, contributes to root development and low-phosphate tolerance in maize. Here, ZmPTF1 expression, drought tolerance, and the underlying mechanisms were studied by using maize ZmPTF1 overexpression lines and mutants. ZmPTF1 was found to be a positive regulator of root development, ABA synthesis, signalling pathways, and drought tolerance. ZmPTF1 was also found to bind to the G-box element within the promoter of 9-cis-epoxycarotenoid dioxygenase (NCED), C-repeat-binding factor (CBF4), ATAF2/NAC081, NAC30, and other transcription factors, and to act as a positive regulator of the expression of those genes. The dramatically upregulated NCEDs led to increased abscisic acid (ABA) synthesis and activation of the ABA signalling pathway. The up-regulated transcription factors hierarchically regulate the expression of genes involved in root development, stress responses, and modifications of transcriptional regulation. The improved root system, increased ABA content, and activated ABA-, CBF4-, ATAF2-, and NAC30-mediated stress responses increased the drought tolerance of the ZmPTF1 overexpression lines, while the mutants showed opposite trends. This study describes a useful gene for transgenic breeding and helps us understand the role of a bHLH protein in plant root development and stress responses.

Overexpression of a novel peanut CLAVATA1-like gene significantly enhanced the resistance to Ralstonia solanacearum in tobacco via defense response signaling associated with EDS1-mediated R-gene pathways., Bacterial wilt caused by Ralstonia solanacearum is a devastating disease affecting hundreds of plant species, yet the host factors remain poorly characterized. The leucine-rich repeat receptor-like kinase gene AhRLK1, characterized as CLAVATA1, was found to be up-regulated in peanut upon inoculation with R. solanacearum. The AhRLK1 protein was localized in the plasma membrane and cell wall. qPCR results showed AhRLK1 was induced in a susceptible variety but little changed in a resistant cultivar after inoculated with R. solanacearum. Hormones such as salicylic acid, abscisic acid, methyl jasmonate, and ethephon induced AhRLK1 expression. In contrast, AhRLK1 expression was down-regulated under cold and drought treatments. Transient overexpression of AhRLK1 led to a hypersensitive response (HR) in Nicotiana benthamiana. Furthermore, AhRLK1 overexpression in tobacco significantly increased the resistance to R. solanacearum. Besides, the transcripts of most representative defense responsive genes in HR and hormone signal pathways were significantly increased in the transgenic lines. EDS1 and PAD4 in the R gene signaling pathway were also up-regulated, but NDR1 was down-regulated. Accordingly, AhRLK1 may increase the defense response to R. solanacearum via HR and hormone defense signaling, in particular through the EDS1 pathway of R gene signaling. These results provide a new understanding of the CLAVATA1 function and will contribute to genetic enhancement of peanut.

The APETALA1 (AP1)/FRUITFULL (FUL)-like transcription factor OsMADS18 plays diverse functions in rice development, but the underlying molecular mechanisms are far from fully understood. Here, we report that down-regulation of OsMADS18 expression in RNAi lines caused a delay in seed germination and young seedling growth, whereas the overexpression of OsMADS18 produced plants with fewer tillers. In targeted OsMADS18 genome-edited mutants (osmads18-cas9), an increased number of tillers, altered panicle size, and reduced seed setting were observed. The EYFP-OsMADS18 (full-length) protein was localized to the nucleus and plasma membrane but the EYFP-OsMADS18-N (N-terminus) protein mainly localized to the nucleus. The expression of OsMADS18 could be stimulated by abscisic acid (ABA), and ABA stimulation triggered the cleavage of HA-OsMADS18 and the translocation of OsMADS18 from the plasma membrane to the nucleus. The inhibitory effect of ABA on seedling growth was less effective in the OsMADS18-overexpressing plants. The expression of a set of ABA-responsive genes was significantly reduced in the overexpressing plants. The phenotypes of transgenic plants expressing EYFP-OsMADS18-N resembled those observed in the osmads18-cas9 mutants. Analysis of the interaction of OsMADS18 with OsMADS14, OsMADS15, and OsMADS57 strongly suggests an essential role for OsMADS18 in rice development., OsMADS18 plays an essential role in rice development, including seed maturation and germination, tiller formation, and the response to ABA

In contrast to the light requirement for Arabidopsis seed germination, the germination of several Aethionema arabicum accessions is inhibited by light, due to antipodal transcriptional regulation of hormone balance., The timing of seed germination is crucial for seed plants and is coordinated by internal and external cues, reflecting adaptations to different habitats. Physiological and molecular studies with lettuce and Arabidopsis thaliana have documented a strict requirement for light to initiate germination and identified many receptors, signaling cascades, and hormonal control elements. In contrast, seed germination in several other plants is inhibited by light, but the molecular basis of this alternative response is unknown. We describe Aethionema arabicum (Brassicaceae) as a suitable model plant to investigate the mechanism of germination inhibition by light, as this species has accessions with natural variation between light-sensitive and light-neutral responses. Inhibition of germination occurs in red, blue, or far-red light and increases with light intensity and duration. Gibberellins and abscisic acid are involved in the control of germination, as in Arabidopsis, but transcriptome comparisons of light- and dark-exposed A. arabicum seeds revealed that, upon light exposure, the expression of genes for key regulators undergo converse changes, resulting in antipodal hormone regulation. These findings illustrate that similar modular components of a pathway in light-inhibited, light-neutral, and light-requiring germination among the Brassicaceae have been assembled in the course of evolution to produce divergent pathways, likely as adaptive traits.

MYB-type transcription factors (TFs) play important roles in plant growth and development, and in the responses to several abiotic stresses. In rice (Oryza sativa), the roles of MYB-related TFs in leaf senescence are not well documented. Here, we examined rice MYB TF gene OsMYB102 and found that an OsMYB102 T-DNA activation-tagged line (termed osmyb102-D), which constitutively expresses OsMYB102 under the control of four tandem repeats of the 35S promoter, and OsMYB102-overexpressing transgenic lines (35S:OsMYB102 and 35S:GFP-OsMYB102) maintain green leaves much longer than the wild-type under natural, dark-induced, and abscisic acid (ABA)-induced senescence conditions. Moreover, an osmyb102 knockout mutant showed an accelerated senescence phenotype under dark-induced and ABA-induced leaf senescence conditions. Microarray analysis showed that a variety of senescence-associated genes (SAGs) were down-regulated in the osmyb102-D line. Further studies demonstrated that overexpression of OsMYB102 controls the expression of SAGs, including genes associated with ABA degradation and ABA signaling (OsABF4, OsNAP, and OsCYP707A6), under dark-induced senescence conditions. OsMYB102 inhibits ABA accumulation by directly activating the transcription of OsCYP707A6, which encodes the ABA catabolic enzyme ABSCISIC ACID 8′-HYDROXYLASE. OsMYB102 also indirectly represses ABA-responsive genes, such as OsABF4 and OsNAP. Collectively, these results demonstrate that OsMYB102 plays a critical role in leaf senescence by down-regulating ABA accumulation and ABA signaling responses., Rice transcription factor OsMYB102 delays leaf senescence by decreasing ABA accumulation through up-regulation of OsCYP707A6, which encodes an ABA catalytic enzyme.

Carbohydrate accumulation in inferior grains in rice panicles subjected to moderate post-anthesis water stress is regulated by NAC transcription factors that activate the expression of WAXY., Moderate soil drying imposed at the post-anthesis stage significantly increases starch accumulation in inferior grains of rice, but how this process is regulated at the level of gene expression remains unclear. In this study, we applied moderate drying (MD) treatments to the soil at the post-anthesis stage and followed the dynamics of the conversion process of soluble sugars to starch in inferior grains using RNA-seq analysis. An elevated level of ABA induced by MD was consistently associated with down-regulation of ABA8ox2, suggesting that lower expression of this gene may be responsible for the higher ABA content, potentially resulting in better filling in inferior grains. In addition, MD treatments up-regulated genes encoding five key enzymes involved sucrose-to-starch conversion and increased the activities of enzymes responsible for soluble-sugar reduction and starch accumulation in inferior grains. Differentially expressed transcription factors, including NAC, GATA, WRKY, and M-type MADS, were predicted to interact with other proteins in mediating filling of inferior grains as a response to MD. Transient expression analysis showed that NAC activated WAXY expression by binding to its promoter, indicating that NAC played a key role in starch synthesis of inferior grains under MD treatment. Our results provide new insights into the molecular mechanisms that regulate grain filling in inferior grains of rice under moderate soil drying.

Strigolactones positively regulate defense against infection by root-knot nematode in tomato through a process that is dependent on MYC2 integrating SL, ABA, and JA signaling, Strigolactones (SLs) are carotenoid-derived phytohormones that are known to influence various aspects of plant growth and development. As root-derived signals, SLs can enhance symbiosis between plants and arbuscular mycorrhizal fungi (AMF). However, little is known about the roles of SLs in plant defense against soil-borne pathogens. Here, we determined that infection with root-knot nematodes (RKNs; Meloidogyne incognita) induced SL biosynthesis in roots of tomato (Solanum lycopersicum). Silencing of SL biosynthesis genes compromised plant defense against RKNs, whilst application of the SL analog racGR24 enhanced it. Accumulation of endogenous jasmonic acid (JA) and abscisic acid (ABA) in the roots in response to RKN infection was enhanced by silencing of SL biosynthetic genes and was suppressed by application of racGR24. Genetic evidence showed that JA was a positive regulator of defense against RKNs while ABA was a negative regulator. In addition, racGR24 enhanced the defense against nematode in a JA-deficient mutant but not in an ABA-deficient mutant. Silencing of SL biosynthetic genes resulted in up-regulation of MYC2, which negatively regulated defense against RKNs. Our results demonstrate that SLs play a positive role in nematode defense in tomato and that MYC2 negatively regulates this defense, potentially by mediating hormone crosstalk among SLs, ABA and JA.

GhNAC83 inhibits corm dormancy release in Gladiolus hybridus by promoting ABA signaling and inhibiting CK biosynthesis., Corm dormancy is an important trait for breeding in many bulb flowers, including the most cultivated Gladiolus hybridus. Gladiolus corms are modified underground stems that function as storage organs and remain dormant to survive adverse environmental conditions. Unlike seed dormancy, not much is known about corm dormancy. Here, we characterize the mechanism of corm dormancy release (CDR) in Gladiolus. We identified an important ABA (abscisic acid) signaling regulator, GhPP2C1 (protein phosphatase 2C1), by transcriptome analysis of CDR. GhPP2C1 expression increased during CDR, and silencing of GhPP2C1 expression in dormant cormels delayed CDR. Furthermore, we show that GhPP2C1 expression is directly regulated by GhNAC83, which was identified by yeast one-hybrid library screening. In planta assays show that GhNAC83 is a negative regulator of GhPP2C1, and silencing of GhNAC83 promoted CDR. As expected, silencing of GhNAC83 decreased the ABA level, but also dramatically increased cytokinin (CK; zeatin) content in cormels. Binding assays demonstrate that GhNAC83 associates with the GhIPT (ISOPENTENYLTRANSFERASE) promoter and negatively regulates zeatin biosynthesis. Taken together, our results reveal that GhNAC83 promotes ABA signaling and synthesis, and inhibits CK biosynthesis pathways, thereby inhibiting CDR. These findings demonstrate that GhNAC83 regulates the ABA and CK pathways, and therefore controls corm dormancy.

ADF5 promotes stomatal closure by regulating actin filament dynamics, and members of the ABF/AREB transcription factor family may serve as potential upstream regulators of ADF5 in the drought stress/ABA signaling pathway., Stomatal movement plays an essential role in plant responses to drought stress, and the actin cytoskeleton and abscisic acid (ABA) are two important components of this process. Little is known about the mechanism underlying actin cytoskeleton remodeling and the dynamic changes occurring during stomatal movement in response to drought stress/ABA signaling. Actin-depolymerizing factors (ADFs) are conserved actin severing/depolymerizing proteins in eukaryotes, and in angiosperms ADFs have evolved actin-bundling activity. Here, we reveal that the transcriptional expression of neofunctionalized Arabidopsis ADF5 was induced by drought stress and ABA treatment. Furthermore, we demonstrated that ADF5 loss-of-function mutations increased water loss from detached leaves, reduced plant survival rates after drought stress, and delayed stomatal closure by regulating actin cytoskeleton remodeling via its F-actin-bundling activity. Biochemical assays revealed that an ABF/AREB transcription factor, DPBF3, could bind to the ADF5 promoter and activate its transcription via the ABA-responsive element core motif ACGT/C. Taken together, our findings indicate that ADF5 participates in drought stress by regulating stomatal closure, and may also serve as a potential downstream target of the drought stress/ABA signaling pathway via members of the ABF/AREB transcription factors family.

This review focuses on recent advances relating to H2S signaling mechanisms, and highlights interconnections between H2S and H2O2 at the post-translational modification level and with ABA in stomatal movement., Hydrogen sulfide (H2S) is a signaling molecule that regulates critical processes and allows plants to adapt to adverse conditions. The molecular mechanism underlying H2S action relies on its chemical reactivity, and the most-well characterized mechanism is persulfidation, which involves the modification of protein thiol groups, resulting in the formation of persulfide groups. This modification causes a change of protein function, altering catalytic activity or intracellular location and inducing important physiological effects. H2S cannot react directly with thiols but instead can react with oxidized cysteine residues; therefore, H2O2 signaling through sulfenylation is required for persulfidation. A comparative study performed in this review reveals 82% identity between sulfenylome and persulfidome. With regard to abscisic acid (ABA) signaling, widespread evidence shows an interconnection between H2S and ABA in the plant response to environmental stress. Proteomic analyses have revealed persulfidation of several proteins involved in the ABA signaling network and have shown that persulfidation is triggered in response to ABA. In guard cells, a complex interaction of H2S and ABA signaling has also been described, and the persulfidation of specific signaling components seems to be the underlying mechanism.

The chloride component of NaCl salinity causes the leaf apoplast to transiently alkalinize. This transition in pH reduces stomatal aperture. However, whether this apoplastic pH (pHapo) transient initiates stomatal closure by interacting with other chloride stress-induced responses or whether the pH transient alone initiates stomatal closure is unknown. To clarify the problem, the transient alkalinization of the leaf apoplast was mimicked in intact maize (Zea mays L.) by infiltrating near-neutral pH buffers into the leaf apoplast. Effects of the pHapo transient could thus be investigated independently from other chloride stress-derived effects. Microscopy-based ratiometric live pHapo imaging was used to monitor pHapoin planta. LC-MS/MS and real-time quantitative reverse transcription–PCR leaf analyses showed that the artificially induced pHapo transient led to an increase in the concentrations of the stomata-regulating plant hormone abscisic acid (ABA) and in transcripts of the key ABA-synthesizing gene ZmVp14 in the leaf. Since stomatal aperture and stomatal conductance decreased according to pHapo, we conclude that the pHapo transient alone initiates stomatal closure. Therefore, the functionality does not depend on interactions with other compounds induced by chloride stress. Overall, our data indicate that the pH of the leaf apoplast links chloride salinity with the control of stomatal aperture via effects exerted on the transcription of ABA., In maize, the transient alkalinization of the leaf apoplast acts as a switch that increases transcription of the ABA-synthesizing key gene viviparous 14 and reduces transpiration.

ABA produced in the fig fruit triggers ethylene production in the inflorescence and induces non-climacteric ripening in the receptacle tissue., The common fig bears a unique closed inflorescence structure, the syconium, composed of small individual drupelets that develop from the ovaries, which are enclosed in a succulent receptacle of vegetative origin. The fig ripening process is traditionally classified as climacteric; however, recent studies have suggested that distinct mechanisms exist in its reproductive and non-reproductive parts. We analysed ABA and ethylene production, and expression of ABA-metabolism, ethylene-biosynthesis, MADS-box, NAC, and ethylene response-factor genes in inflorescences and receptacles of on-tree fruit treated with ABA, ethephon, fluridone, and nordihydroguaiaretic acid (NDGA). Exogenous ABA and ethephon accelerated fruit ripening and softening, whereas fluridone and NDGA had the opposite effect, delaying endogenous ABA and ethylene production compared to controls. Expression of the ABA-biosynthesis genes FcNCED2 and FcABA2, ethylene-biosynthesis genes FcACS4, FcACOL, and FcACO2, FcMADS8, 14, 15, FcNAC1, 2, 5, and FcERF9006 was up-regulated by exogenous ABA and ethephon. NDGA down-regulated FcNCED2 and FcABA2, whereas fluridone down-regulated FcABA2; both down-regulated the ethylene-related genes. These results demonstrate the key role of ABA in regulation of ripening by promoting ethylene production, as in the climacteric model plant tomato, especially in the inflorescence. However, increasing accumulation of endogenous ABA until full ripeness and significantly low expression of ethylene-biosynthesis genes in the receptacle suggests non-climacteric, ABA-dependent ripening in the vegetative-originated succulent receptacle part of the fruit.

An NO-sensing mechanism controlling hypocotyl growth in etiolated seedlings requires the biosynthesis and/or signaling of ethylene, strigolactones, salicylate, abscisic acid, and brassinosteroids., Plants are often exposed to high levels of nitric oxide (NO) that affects development and stress-triggered responses. However, the way in which plants sense NO is still largely unknown. Here we combine the analysis of early changes in the transcriptome of plants exposed to a short acute pulse of exogenous NO with the identification of transcription factors (TFs) involved in NO sensing. The NO-responsive transcriptome was enriched in hormone homeostasis- and signaling-related genes. To assess events involved in NO sensing in hypocotyls, we used a functional sensing assay based on the NO-induced inhibition of hypocotyl elongation in etiolated seedlings. Hormone-related mutants and the TRANSPLANTA collection of transgenic lines conditionally expressing Arabidopsis TFs were screened for NO-triggered hypocotyl shortening. These approaches allowed the identification of hormone-related TFs, ethylene perception and signaling, strigolactone biosynthesis and signaling, and salicylate production and accumulation that are essential for or modulate hypocotyl NO sensing. Moreover, NO inhibits hypocotyl elongation through the positive and negative regulation of some abscisic acid (ABA) receptors and transcripts encoding brassinosteroid signaling components thereby also implicating these hormones in NO sensing.

Silencing the calcium-dependent protein kinase CPK27 in tomato decreases cold-induced ABA, H2O2 and NO production, as well as decreasing activation of mitogen-activated protein kinase, compromising cold acclimation., Cold acclimation-induced cold tolerance is associated with the generation of reactive oxygen species (ROS), nitric oxide (NO), and mitogen-activated protein kinases (MPKs) in plants. Here, we hypothesized that calcium-dependent protein kinases (CPKs) induce a crosstalk among ROS, NO, and MPKs, leading to the activation of abscisic acid (ABA) signaling in plant adaptation to cold stress. Results showed that cold acclimation significantly increased the transcript levels of CPK27 along with the biosynthesis of ABA in tomato (Solanum lycopersicum). Silencing of CPK27 compromised acclimation-induced cold tolerance, generation of hydrogen peroxide (H2O2) in the apoplast, NO and ABA accumulation, and the activation of MPK1/2. Crosstalk among H2O2, NO, and MPK1/2 contributes to the homeostasis of H2O2 and NO, activation of MPK1/2, and cold tolerance. ABA is also critical for CPK27-induced cold tolerance, generation of H2O2 and NO, and the activation of MPK1/2. These results strongly suggest that CPK27 may function as a positive regulator of ABA generation by activating the production of ROS and NO as well as MPK1/2 in cold adaptation.

Acclimation of wheat to waterlogging is mediated by regulation of hormonal metabolism and transport in adventitious roots and during their emergence., To gain insights into the molecular mechanisms underlying hormonal regulation in adventitious roots and during their emergence under waterlogged conditions in wheat, the present study investigated transcriptional regulation of genes related to hormone metabolism and transport in the root and stem node tissues. Waterlogging-induced inhibition of axile root elongation and lateral root formation, and promotion of surface adventitious and axile root emergence and aerenchyma formation are associated with enhanced expression levels of ethylene biosynthesis genes, ACS7 and ACO2, in both tissues. Inhibition of axile root elongation is also related to increased root indole acetic acid (IAA) and jasmonate (JA) levels that are associated with up-regulation of specific IAA biosynthesis/transport (TDC, YUC1, and PIN9) and JA metabolism (LOX8, AOS1, AOC1, and JAR1) genes, and transcriptional alteration of gibberellin (GA) metabolism genes (GA3ox2 and GA2ox8). Adventitious root emergence from waterlogged stem nodes is associated with increased levels of IAA and GA but decreased levels of cytokinin and abscisic acid (ABA), which are regulated through the expression of specific IAA biosynthesis/transport (TDC, YUC1, and PIN9), cytokinin metabolism (IPT5-2, LOG1, CKX5, and ZOG2), ABA biosynthesis (NCED1 and NCED2), and GA metabolism (GA3ox2 and GA2ox8) genes. These results enhance our understanding of the molecular mechanisms underlying the adaptive response of wheat to waterlogging.

Gibberellin has an inhibitory effect during initial activation of dormant grapevine buds, and at this stage its level is down-regulated. At a later stage, gibberellin level increases and enhances bud regrowth., The molecular mechanism regulating dormancy release in grapevine buds is as yet unclear. It has been hypothesized that (i) abscisic acid (ABA) represses bud–meristem activity; (ii) perturbation of respiration induces an interplay between ethylene and ABA metabolism, which leads to removal of repression; and (iii) gibberellin (GA)-mediated growth is resumed. The first two hypothesis have been formally supported. The current study examines the third hypothesis regarding the potential involvement of GA in dormancy release. We found that during natural dormancy induction, levels of VvGA3ox, VvGA20ox, and VvGASA2 transcripts and of GA1 were decreased. However, during dormancy release, expression of these genes was enhanced, accompanied by decreased expression of the bud-expressed GA-deactivating VvGA2ox. Despite indications for its positive role during natural dormancy release, GA application had inhibitory effects on bud break. Hydrogen cyanamide up-regulated VvGA2ox and down-regulated VvGA3ox and VvGA20ox expression, reduced GA1 levels, and partially rescued the negative effect of GA. GA had an inhibitory effect only when applied simultaneously with bud-forcing initiation. Given these results, we hypothesize that during initial activation of the dormant bud meristem, the level of GA must be restricted, but after meristem activation an increase in its level serves to enhance primordia regrowth.

A leucine-rich repeat receptor-like kinase in strawberry, FaRIPK1, interacts with a putative ABA receptor, FaABAR, and forms a newly identified ABA signaling pathway involved in the regulation of fruit ripening., Strawberry (Fragaria×ananassa) is a model plant for studying non-climacteric fruit ripening regulated by abscisic acid (ABA); however, its exact molecular mechanisms are yet not fully understood. In this study, a predicted leu-rich repeat (LRR) receptor-like kinase in strawberry, red-initial protein kinase 1 (FaRIPK1), was screened and, using a yeast two-hybrid assay, was shown to interact with a putative ABA receptor, FaABAR. This association was confirmed by bimolecular fluorescence complementation and co-immunoprecipitation assays, and shown to occur in the nucleus. Expression analysis by real-time PCR showed that FaRIPK1 is expressed in roots, stems, leaves, flowers, and fruit, with a particularly high expression in white fruit at the onset of coloration. Down-regulation of FaRIPK1 expression in strawberry fruit, using Tobacco rattle virus-induced gene silencing, inhibited ripening, as evidenced by suppression of ripening-related physiological changes and reduced expression of several genes involved in softening, sugar content, pigmentation, and ABA biosynthesis and signaling. The yeast-expressed LRR and STK (serine/threonine protein kinase) domains of FaRIPK1 bound ABA and showed kinase activity, respectively. A fruit disc-incubation test revealed that FaRIPK1 expression was induced by ABA and ethylene. The synergistic action of FaRIPK1 with FaABAR in regulation of strawberry fruit ripening is discussed.

This study synchronously investigated maize growth and physiological responses to progressive soil drying. It indicates that hydraulic and chemical changes may regulate plant development and functioning during onset of drought., A better understanding of physiological responses of crops to drought stress is important for ensuring sustained crop productivity under climate change. Here, we studied the effect on 15-day-old maize (Zea mays L.) plants of a 6 d non-lethal period of soil drying [soil water potential (SWP) decreased from –0.20 MPa to –0.81 MPa]. Root growth was initially stimulated during drying (when SWP decreased from –0.31 MPa to –0.38 MPa, compared with –0.29 MPa in well-watered pots), followed by inhibition during Days 5–6 (SWP from –0.63 MPa to –0.81 MPa). Abscisic acid (ABA) in the root began to accumulate as the root water potential declined during Days 2–3. Leaf elongation was inhibited from Day 4 (SWP less than –0.51 MPa), just after leaf ABA content began to increase, but coinciding with a decline in leaf water potential. The stomatal conductance was restricted earlier in the younger leaf (fourth) (on Day 3) than in the older leaf (third). The ethylene content of leaves and roots decreased during drying, but after the respective increase in ABA contents. This work identified critical timing of hydraulic and chemical changes at the onset of soil drying, which can be important in initiating early stomatal and growth responses to drought.

Potassium maintains cotton fiber cell elongation under conditions of calcium deficiency, through the CBL-INTERACTING PROTEIN KINASE 6 (GhCIPK6) gene., Calcium (Ca2+) is necessary for fiber cell development in cotton (Gossypium hirsutum), both as a cell wall structural component and for environmental signaling responses. It is also known that potassium (K+) plays a critical role in cotton fiber cell elongation. However, it is unclear whether Ca2+ integrates its activities with K+ to regulate fiber elongation. Here, we report the novel discovery that Ca2+ deficiency, when integrated with K+ signaling, promotes fiber elongation. Using inductively coupled plasma–mass spectrometry (ICP-MS), we determined dynamic profiles of the ionome in ovules and fibers at different developmental stages, and found that a high accumulation of macro-elements, but not Ca2+, was associated with longer fibers. Using an in vitro ovule culture system, we found that under Ca2+-deficient conditions, sufficient K+ (52 mM) rapidly induced ovule and fiber browning, while reduced K+ (2 or 27 mM) not only suppressed tissue browning but also altered fiber elongation. Reduced K+ also enhanced reactive oxygen species scavenging ability and maintained abscisic acid and jasmonic acid levels, which in turn compensated for Ca2+ deficiency. Ca2+ deficiency combined with reduced K+ (0 mM Ca2+ and 27 mM K+) produced longer fibers in cultured ovules, due to cell wall loosening by phytosulfokine (PSK), expansin (EXP), and xyloglucan endotransglycosylase/hydrolase (XTH), and an increase of the K+ content of fiber cells. Using transgenic cotton, we showed that the CBL-INTERACTING PROTEIN KINASE 6 (GhCIPK6) gene mediates the uptake of K+ under Ca2+-deficient conditions. This study establishes a new link between Ca2+, K+, and fiber elongation.

RNAi-repression of vacuolar HvSUT2 in transgenic barley demonstrates its indispensable role for proper grain filling and for the control of sucrose homeostasis in concert with the plasma-membrane localised HvSUT1., Sucrose transport and partitioning are crucial for seed filling. While many plasma-membrane-localised sucrose transporters (SUT1 family members) have been analysed in seeds, the functions of vacuolar SUT2 members are still obscure. In barley grains, expression of HvSUT1 and HvSUT2 overlap temporally and spatially, suggesting concerted functions to regulate sucrose homeostasis. Using HvSUT2-RNAi plants, we found that grains were also deficient in HvSUT1 expression and seemingly sucrose-limited during mid-to-late grain filling. Transgenic endosperms accumulated less starch and dry weight, although overall sucrose and hexose contents were higher. Comprehensive transcript and metabolite profiling revealed that genes related to glycolysis, the tricarboxylic acid cycle, starch and amino acid synthesis, grain maturation, and abscisic acid signalling were down-regulated together with most glycolytic intermediates and amino acids. Sucrose was increased along the sucrose delivery route in the nucellar projection, the endosperm transfer cells, and the starchy endosperm, indicating that suppressed transporter activity diminished sucrose efflux from vacuoles, which generated sugar deficiency in the cytoplasm. Thus, endosperm vacuoles may buffer sucrose concentrations to regulate homeostasis at grain filling. Transcriptional changes revealed that limited endosperm sucrose initiated sugar starvation responses, such as sugar recycling from starch, hemicelluloses and celluloses together with vacuolar protein degradation, thereby supporting formation of nucleotide sugars. Barley endosperm cells can thus suppress certain pathways to retrieve resources to maintain essential cell functions.

ACCERBATIN is an ethylene-mimicking small molecule that affects auxin homeostasis and ROS accumulation in etiolated seedlings. In light-grown plants, it exhibits auxin-like herbicidal properties., The volatile two-carbon hormone ethylene acts in concert with an array of signals to affect etiolated seedling development. From a chemical screen, we isolated a quinoline carboxamide designated ACCERBATIN (AEX) that exacerbates the 1-aminocyclopropane-1-carboxylic acid-induced triple response, typical for ethylene-treated seedlings in darkness. Phenotypic analyses revealed distinct AEX effects including inhibition of root hair development and shortening of the root meristem. Mutant analysis and reporter studies further suggested that AEX most probably acts in parallel to ethylene signaling. We demonstrated that AEX functions at the intersection of auxin metabolism and reactive oxygen species (ROS) homeostasis. AEX inhibited auxin efflux in BY-2 cells and promoted indole-3-acetic acid (IAA) oxidation in the shoot apical meristem and cotyledons of etiolated seedlings. Gene expression studies and superoxide/hydrogen peroxide staining further revealed that the disrupted auxin homeostasis was accompanied by oxidative stress. Interestingly, in light conditions, AEX exhibited properties reminiscent of the quinoline carboxylate-type auxin-like herbicides. We propose that AEX interferes with auxin transport from its major biosynthesis sites, either as a direct consequence of poor basipetal transport from the shoot meristematic region, or indirectly, through excessive IAA oxidation and ROS accumulation. Further investigation of AEX can provide new insights into the mechanisms connecting auxin and ROS homeostasis in plant development and provide useful tools to study auxin-type herbicides.

Decreased leaf turgor triggers up-regulation of the rate-limiting NCED3 gene for de novo biosynthesis of ABA within the narrow time-frame required to initiate stomatal closure at increased vapour pressure deficit., A major environmental signal influencing day-time stomatal aperture is the vapour pressure deficit between the leaf and atmosphere (VPD). In angiosperms, increased VPD triggers biosynthesis of abscisic acid (ABA), prompting rapid stomatal closure. Altered cell turgor has been proposed as the trigger for ABA biosynthesis, but the timing and nature of the genetic signals linking these processes have remained uncertain. We investigated this in Arabidopsis by examining changes induced by a decrease in leaf turgor, simulating a natural increase in VPD. We found that the rate-limiting gene within the de novo ABA biosynthesis pathway, 9-cis-epoxycarotenoid dioxygenase 3 (NCED3), was induced and ABA levels increased within just 5 min of decreased leaf turgor. This rapid induction matches the time-frame for initiation of stomatal closure in response to a doubling in VPD. We further examined Arabidopsis histidine kinase1 (AHK1) as the most likely candidate for the turgor-sensing receptor involved, but found no significant difference between wild-type and an ahk1 null mutant in the induction of ABA-biosynthetic genes, ABA production, or stomatal behaviour. We show that decreased leaf turgor triggers de novo ABA biosynthesis within the time-frame of the stomatal response to VPD, but that AHK1 does not fulfil a critical role as a turgor-sensing receptor within this pathway.

Highlight PCA22 negatively regulates the drought response through the modulation of osmolytic components and regulates pollen tube length in Arabidopsis., Proline metabolism is important for environmental responses, plant growth, and development. However, its precise roles in plant abiotic stress tolerance are not well understood. Mutants are valuable for the identification of new genes and for elucidating their roles in physiological mechanisms. We applied a suppressor mutation approach to identify novel genes involved in the regulation of proline metabolism in Arabidopsis. Using the atrzf1 (Arabidopsis thaliana ring zinc finger 1) mutant as a parental line for activation tagging mutagenesis, we selected several mutants with suppressed induction of proline accumulation under dehydration conditions. One of the selected mutants [proline content alterative 22 (pca22)] appeared to have reduced proline contents compared with the atrzf1 mutant under drought stress. Generally, pca22 mutant plants displayed suppressed atrzf1 insensitivity to dehydration and abscisic acid during early seedling growth. Additionally, the pca22 mutant exhibited shorter pollen tube length than wild-type (WT) and atrzf1 plants. Furthermore, PCA22-overexpressing plants were more sensitive to dehydration stress than the WT and RNAi lines. Green fluorescent protein-tagged PCA22 was localized to the cytoplasm of transgenic Arabidopsis cells. Collectively, these results suggest that pca22 acts as dominant suppressor mutant of atrzf1 in the abiotic stress response.

Highlight Auxin and jasmonates are involved in whole-plant regulation of growth and secondary metabolites, whereas abscisic and salicylic acid may be associated with leaf oxidative stress in response to low carbon availability., Phytohormones play important roles in plant acclimation to changes in environmental conditions. However, their role in whole-plant regulation of growth and secondary metabolite production under increasing atmospheric CO2 concentrations ([CO2]) is uncertain but crucially important for understanding plant responses to abiotic stresses. We grew winter wheat (Triticum aestivum) under three [CO2] (170, 390, and 680 ppm) over 10 weeks, and measured gas exchange, relative growth rate (RGR), soluble sugars, secondary metabolites, and phytohormones including abscisic acid (ABA), auxin (IAA), jasmonic acid (JA), and salicylic acid (SA) at the whole-plant level. Our results show that, at the whole-plant level, RGR positively correlated with IAA but not ABA, and secondary metabolites positively correlated with JA and JA-Ile but not SA. Moreover, soluble sugars positively correlated with IAA and JA but not ABA and SA. We conclude that increasing carbon availability stimulates growth and production of secondary metabolites via up-regulation of auxin and jasmonate levels, probably in response to sugar-mediated signalling. Future low [CO2] studies should address the role of reactive oxygen species (ROS) in leaf ABA and SA biosynthesis, and at the transcriptional level should focus on biosynthetic and, in particular, on responsive genes involved in [CO2]-induced hormonal signalling pathways.

Highlight Grapevine red blotch interferes with the hormonal and transcriptional regulation of berry development, leading to suppression of secondary metabolism and induction of photosynthesis at late ripening stages., Grapevine red blotch-associated virus (GRBaV) is a major threat to the wine industry in the USA. GRBaV infections (aka red blotch disease) compromise crop yield and berry chemical composition, affecting the flavor and aroma properties of must and wine. In this study, we combined genome-wide transcriptional profiling with targeted metabolite analyses and biochemical assays to characterize the impact of the disease on red-skinned berry ripening and metabolism. Using naturally infected berries collected from two vineyards, we were able to identify consistent berry responses to GRBaV across different environmental and cultural conditions. Specific alterations of both primary and secondary metabolism occurred in GRBaV-infected berries during ripening. Notably, GRBaV infections of post-véraison berries resulted in the induction of primary metabolic pathways normally associated with early berry development (e.g. thylakoid electron transfer and the Calvin cycle), while inhibiting ripening-associated pathways, such as a reduced metabolic flux in the central and peripheral phenylpropanoid pathways. We show that this metabolic reprogramming correlates with perturbations at multiple regulatory levels of berry development. Red blotch caused the abnormal expression of transcription factors (e.g. NACs, MYBs, and AP2-ERFs) and elements of the post-transcriptional machinery that function during red-skinned berry ripening. Abscisic acid, ethylene, and auxin pathways, which control both the initiation of ripening and stress responses, were also compromised. We conclude that GRBaV infections disrupt normal berry development and stress responses by altering transcription factors and hormone networks, which result in the inhibition of ripening pathways involved in the generation of color, flavor, and aroma compounds.

Highlight Increasing the ratio of red to far red light promotes axillary bud outgrowth and suppresses bud abscisic acid signalling within 3 h, before stem auxin signalling is affected., Arabidopsis thaliana shoot branching is inhibited by a low red light to far red light ratio (R:FR, an indicator of competition), and by loss of phytochrome B function. Prior studies have shown that phytochrome B deficiency suppresses bud growth by elevating systemic auxin signalling, and that increasing the R:FR promotes the growth of buds suppressed by low R:FR by inhibiting bud abscisic acid (ABA) accumulation and signalling. Here, systemic auxin signalling and bud ABA signalling were examined in the context of rapid bud responses to an increased R:FR. Increasing the R:FR promoted the growth of buds inhibited by a low R:FR within 6 h. Relative to a low R:FR, bud ABA accumulation and signalling in plants given a high R:FR showed a sustained decline within 3 h, prior to increased growth. Main stem auxin levels and signalling showed a weak, transient response. Systemic effects and those localised to the bud were further examined by decapitating plants maintained either under a low R:FR or provided with a high R:FR. Increasing the R:FR promoted bud growth before decapitation, but decapitated plants eventually formed longer branches. The data suggest that rapid responses to an increased R:FR may be mediated by changes in bud ABA physiology, although systemic auxin signalling is necessary for sustained bud repression under a low R:FR.

A novel NAC transcription factor, CrNAC036, can interact with CrMYB68 to synergistically regulate the ripening process of citrus fruit by down-regulating the expression of 9-cis-epoxycarotenoid dioxygenase 5., Although abscisic acid (ABA) is a vital regulator of fruit ripening and several transcription factors have been reported to regulate ABA biosynthesis, reports of the effect of ABA on citrus ripening and the regulation of its biosynthesis by a multiple-transcription-factor complex are scarce. In the present study, a systematic metabolic, cytological, and transcriptome analysis of an ABA-deficient mutant (MT) of Citrus reticulata cv. Suavissima confirmed the positive effect of ABA on the citrus ripening process. The analysis of transcriptome profiles indicated that CrNAC036 played an important role in the ABA deficiency of the mutant, most likely due to an effect on the expression of 9-cis-epoxycarotenoid dioxygenase 5 (CrNCED5). Electrophoretic mobility shift assays and dual luciferase assays demonstrated that CrNAC036 can directly bind and negatively regulate CrNCED5 expression. Furthermore, yeast two-hybrid, bimolecular fluorescence complementation, and dual luciferase assays demonstrated that CrNAC036 interacted with CrMYB68, also down-regulating the expression of CrNCED5. Taken together, our results suggest that CrNAC036 and CrMYB68 synergistically inhibit ABA biosynthesis in citrus fruit by regulating the expression of CrNCED5.

Moderate water stress in rice induces rhizosheath formation by modulating the abscisic acid and auxin responses to regulate root and root hair growth., The rhizosheath is known to be beneficial for drought resistance in many plants, but the regulation of rhizosheath formation in rice plants is unclear. Here, we investigate rhizosheath formation in different rice varieties and root hair mutants. Our results showed that moderate water stress in rice induced rhizosheath formation. The soil porosity and water content were higher in the rice rhizosheath than in the rice bulk soil under moderate water stress. Additionally, rhizosheath formation in short root hair mutants was lower than in wild-type rice under moderate water stress. Moreover, transcriptomic results indicated that abscisic acid (ABA) and auxin were involved in root and root hair responses in rhizosheath formation. Further, blocking ABA and auxin pathways in wild type and in rhl1-1, the shortest root hair mutant, rhizosheath formation and root hair length were significantly decreased under moderate water stress. However, wild type plants maintained a higher root ABA content, root basipetal auxin transport, root hair length, and amount of rhizosheath than did rhl1-1. Our results suggest that moderate water stress in rice induces rhizosheath formation by modulating the ABA and auxin responses to regulate root and root hair growth, which may be used to breed rice varieties resistant to drought.

Seed germination is a developmental process regulated by numerous internal and external cues. Our previous studies have shown that calcium influx mediated by the Arabidopsis glutamate receptor homolog 3.5 (AtGLR3.5) modulates the expression of the ABSCISIC ACID INSENSITIVE 4 (ABI4) transcription factor during germination and that L-methionine (L-Met) activates AtGLR3.1/3.5 Ca2+ channels in guard cells. However, it is not known whether L-Met participates in regulation of germination and what cellular mechanism is responsible for Met production during germination. Here, we describe Arabidopsis methionine synthase 1 (AtMS1), which acts in the final step of Met biosynthesis, synthesizes the Met required for the activation of AtGLR3.5 Ca2+ channels whose expression is up-regulated during germination, leading to the regulation of seed germination. We show that exogenous L-Met promotes germination in an AtGRL3.5-dependent manner. We also demonstrate that L-Met directly regulates the AtGLR3.5-mediated increase in cytosolic Ca2+ level in seedlings. We provide pharmacological and genetic evidence that Met synthesized via AtMS1 acts upstream of the AtGLR3.5-mediated Ca2+ signal and regulates the expression of ABI4, a major regulator in the abscisic acid response in seeds. Overall, our results link AtMS1, L-Met, the AtGLR3.5 Ca2+ channel, Ca2+ signals, and ABI4, and shed light on the physiological role and molecular mechanism of L-Met in germination., Methionine synthase 1 provides methionine that activates the glutamate receptor homolog 3.5 calcium channel during seed germination in Arabidopsis.

cis-12-Oxo-phytodienoic acid acts in addition to ABA to repress seed germination under far-red light conditions. The response to both these phytohormones is integrated by MFT, a negative regulator of germination., Light-dependent seed germination is induced by gibberellins (GA) and inhibited by abscisic acid (ABA). The widely accepted view of the GA/ABA ratio controlling germination does not, however, explain the fact that seeds deficient in ABA still germinate poorly under shade conditions that repress germination. In Arabidopsis, MOTHER-OF-FT-AND-TFL1 (MFT) acts as a key negative regulator of germination, modulating GA and ABA responses under shade conditions. Under full light the oxylipin cis-12-oxo-phytodienoic acid (OPDA), a precursor of the stress-related phytohormone jasmonic acid, interacts with ABA and MFT to repress germination. Here, we show that under shade conditions both OPDA and ABA repress germination to varying extents. We demonstrate that the level of shade-induced MFT expression influences the ability of OPDA and/or ABA to fully repress germination. We also found that MFT expression decreases with seed age and this again correlates with the response of seeds to OPDA and ABA. We conclude that OPDA plays an essential role alongside ABA in repressing germination in response to shade and the combined effect of these phytohormones is integrated to a significant extent through MFT.