Regulation Of Stomatal Closure Research Articles

Phosphatidic acid produced by phospholipase Dα1 and Dδ is incorporated into the internal membranes but not involved in the gene expression of RD29A in the abscisic acid signaling network in Arabidopsis thaliana.

Core protein components of the abscisic acid (ABA) signaling network, pyrabactin resistance (PYR), protein phosphatases 2C (PP2C), and SNF1-related protein kinase 2 (SnRK2) are involved in the regulation of stomatal closure and gene expression downstream responses in Arabidopsis thaliana. Phosphatidic acid (PA) produced by the phospholipases Dα1 and Dδ (PLDs) in the plasma membrane has been identified as a necessary molecule in ABA-inducible stomatal closure. On the other hand, the involvement of PA in ABA-inducible gene expression has been suggested but remains a question. In this study, the involvement of PA in the ABA-inducible gene expression was examined in the model plant Arabidopsis thaliana and the canonical RD29A ABA-inducible gene that possesses a single ABA-responsive element (ABRE) in the promoter. The promoter activity and accumulation of the RD29A mRNA during ABA exposure to the plants were analyzed under conditions in which the production of PA by PLDs is abrogated through chemical and genetic modification. Changes in the subcellular localization of PA during the signal transduction were analyzed with confocal microscopy. The results obtained in this study suggest that inhibition of PA production by the PLDs does not affect the promoter activity of RD29A. PA produced by the PLDs and exogenously added PA in the plasma membrane are effectively incorporated into internal membranes to transduce the signal. However, exogenously added PA induces stomatal closure but not RD29A expression. This is because PA produced by the PLDs most likely inhibits the activity of not all but only the selected PP2C family members, the negative regulators of the RD29A promoter. This finding underscores the necessity for experimental verifications to adapt previous knowledge into a signaling network model before its construction.

Sinapate Esters Mediate UV-B-Induced Stomatal Closure by Regulating Nitric Oxide, Hydrogen Peroxide, and Malate Accumulation in Arabidopsis thaliana.

Sinapate esters, which are induced in plants under ultraviolet-B (UV-B) irradiation, have important roles not only in the protection against UV-B irradiation but also in the regulation of stomatal closure. Here, we speculated that sinapate esters would function in the stomatal closure of Arabidopsis thaliana in response to UV-B. We measured the stomatal aperture size of the wild-type (WT) and bright trichomes 1 (brt1) and sinapoylglucose accumulator 1 (sng1) mutants under UV-B irradiation; the latter two mutants are deficient in the conversion of sinapic acid to sinapoylglucose (SG) and SG to sinapoylmalate (SM), respectively. Both the brt1 and sng1 plants showed smaller stomatal apertures than the WT under normal light and UV-B irradiation conditions. The accumulation of SM and malate were induced by UV-B irradiation in WT and brt1 plants but not in sng1 plants. Consistently, exogenous malate application reduced UV-B-induced stomatal closure in WT, brt1 and sng1 plants. Nonetheless, levels of reactive oxygen species (ROS), nitric oxide (NO) and cytosolic Ca2+ were higher in guard cells of the sng1 mutant than in those of the WT under normal white light and UV-B irradiation, suggesting that disturbance of sinapate metabolism induced the accumulation of these signaling molecules that promote stomatal closure. Unexpectedly, exogenous sinapic acid application prevented stomatal closure of WT, brt1 and sng1 plants. In summary, we hypothesize that SG or other sinapate esters may promote the UV-B-induced malate accumulation and stomatal closure, whereas sinapic acid inhibits the ROS-NO pathway that regulates UV-B-induced cytosolic Ca2+ accumulation and stomatal closure.

Functional Characterization of Tomato Phytochrome A and B1B2 Mutants in Response to Heat Stress.

Heat stress (HS) is a prevalent negative factor affecting plant growth and development, as it is predominant worldwide and threatens agriculture on a large scale. PHYTOCHROMES (PHYs) are photoreceptors that control plant growth and development, and the stress signaling response partially interferes with their activity. PHYA, B1, and B2 are the most well-known PHY types in tomatoes. Our study aimed to identify the role of tomato ‘Money Maker’ phyA and phyB1B2 mutants in stable and fluctuating high temperatures at different growth stages. In the seed germination and vegetative growth stages, the phy mutants were HS tolerant, while during the flowering stage the phy mutants revealed two opposing roles depending on the HS exposure period. The response of the phy mutants to HS during the fruiting stage showed similarity to WT. The most obvious stage that demonstrated phy mutants’ tolerance was the vegetative growth stage, in which a high degree of membrane stability and enhanced water preservation were achieved by the regulation of stomatal closure. In addition, both mutants upregulated the expression of heat-responsive genes related to heat tolerance. In addition to lower malondialdehyde accumulation, the phyA mutant enhanced proline levels. These results clarified the response of tomato phyA and phyB1B2 mutants to HS.

Biochemical and Molecular Effects Induced by Triacontanol in Acquired Tolerance of Rice to Drought Stress.

To assess the effect of triacontanol (TRIA) on rice plants grown under normal or drought conditions, rice seeds were presoaked in TRIA (35 ppm) for two hours. After 20 days of sowing, rice seedlings developed from TRIA-treated or untreated seeds were subjected to drought stress. After 10 days of plant exposure to drought stress, data of major growth attributes and the content of photosynthetic pigments were recorded. Moreover, the effect of drought stress on stomatal conductance and the photochemical efficiency of PSII (Fv/Fm) were followed. The data obtained indicated that the species of rice (Oryza sativa L.) cultivar Giza 177 under investigation was sensitive to drought stress where there were significant decreases in the fresh and dry weights of shoots and roots and in stomatal conductance, as well as in the content of chlorophyll a, chlorophyll b, and carotenoids. Seed priming with TRIA enhanced both growth and acquired plant tolerance to drought stress. Thus, TRIA via the enhancement of stomatal conductance through the regulation of stomatal closure, the rate of water loss, ABA metabolism, the accumulation of osmolytes, and the regulation of aquaporins genes improved the water status of plants grown under water scarcity. Moreover, TRIA via increasing the content of free amino acids and sugars under drought stress may increase the chance of plant tissues to retain more water under scarcity conditions.

Jasmonic Acid at the Crossroads of Plant Immunity and Pseudomonas syringae Virulence.

Sensing of pathogen infection by plants elicits early signals that are transduced to affect defense mechanisms, such as effective blockage of pathogen entry by regulation of stomatal closure, cuticle, or callose deposition, change in water potential, and resource acquisition among many others. Pathogens, on the other hand, interfere with plant physiology and protein functioning to counteract plant defense responses. In plants, hormonal homeostasis and signaling are tightly regulated; thus, the phytohormones are qualified as a major group of signaling molecules controlling the most widely tinkered regulatory networks of defense and counter-defense strategies. Notably, the phytohormone jasmonic acid mediates plant defense responses to a wide array of pathogens. In this review, we present the synopsis on the jasmonic acid metabolism and signaling, and the regulatory roles of this hormone in plant defense against the hemibiotrophic bacterial pathogen Pseudomonas syringae. We also elaborate on how this pathogen releases virulence factors and effectors to gain control over plant jasmonic acid signaling to effectively cause disease. The findings discussed in this review may lead to ideas for the development of crop cultivars with enhanced disease resistance by genetic manipulation.

Atmospheric H2S exposure does not affect stomatal aperture in maize

Main conclusionStomatal aperture in maize is not affected by exposure to a subtoxic concentration of atmospheric H2S. At least in maize, H2S, thus, is not a gaseous signal molecule that controls stomatal aperture.Sulfur is an indispensable element for the physiological functioning of plants with hydrogen sulfide (H2S) potentially acting as gasotransmitter in the regulation of stomatal aperture. It is often assumed that H2S is metabolized into cysteine to stimulate stomatal closure. To study the significance of H2S for the regulation of stomatal closure, maize was exposed to a subtoxic atmospheric H2S level in the presence or absence of a sulfate supply to the root. Similar to other plants, maize could use H2S as a sulfur source for growth. Whereas sulfate-deprived plants had a lower biomass than sulfate-sufficient plants, exposure to H2S alleviated this growth reduction. Shoot sulfate, glutathione, and cysteine levels were significantly higher in H2S-fumigated plants compared to non-fumigated plants. Nevertheless, this was not associated with changes in the leaf area, stomatal density, stomatal resistance, and transpiration rate of plants, meaning that H2S exposure did not affect the transpiration rate per stoma. Hence, it did not affect stomatal aperture, indicating that, at least in maize, H2S is not a gaseous signal molecule controlling this aperture.

The Tomato DELLA Protein PROCERA Promotes Abscisic Acid Responses in Guard Cells by Upregulating an Abscisic Acid Transporter.

Plants reduce transpiration through stomatal closure to avoid drought stress. While abscisic acid (ABA) has a central role in the regulation of stomatal closure under water-deficit conditions, we demonstrated in tomato (Solanum lycopersicum) that a gibberellin response inhibitor, the DELLA protein PROCERA (PRO), promotes ABA-induced stomatal closure and gene transcription in guard cells. To study how PRO affects stomatal closure, we performed RNA-sequencing analysis of isolated guard cells and identified the ABA transporters ABA-IMPORTING TRANSPORTER1 1 (AIT1 1) and AIT1 2, also called NITRATE TRANSPORTER1/PTR TRANSPORTER FAMILY4 6 in Arabidopsis (Arabidopsis thaliana), as being upregulated by PRO. Tomato has four AIT1 genes, but only AIT1 1 and AIT1 2 were upregulated by PRO, and only AIT1 1 exhibited high expression in guard cells. Functional analysis of AIT1 1 in yeast (Saccharomyces cerevisiae) confirmed its activity as an ABA transporter, possibly an importer. A clustered regularly interspaced short palindromic repeats-Cas9-derived ait1 1 mutant exhibited an increased transpiration, a larger stomatal aperture, and a reduced stomatal response to ABA. Moreover, ait1 1 suppressed the promoting effects of PRO on ABA-induced stomatal closure and gene expression in guard cells, suggesting that the effects of PRO on stomatal aperture and transpiration are AIT1.1-dependent. Previous studies suggest a negative crosstalk between gibberellin and ABA that is mediated by changes in hormone biosynthesis and signaling. The results of this study suggest this crosstalk is also mediated by changes in hormone transport.

Alleviation of drought stress in grapevine by foliar-applied strigolactones

Drought is one of the major abiotic stress factors that affect grape growth and yield, which in turn negatively affects the grape and wine production industry. Developing effective approaches to improve grapevine tolerance to drought stress is a priority for viticulture. Strigolactones, a newly discovered class of carotenoid-derived phytohormones, have been found to participate in various physiological processes. Herein, the effect of strigolactones (SLs) on grape seedlings under drought stress was investigated. Two-year-old grape seedlings (Vitis vinifera L.) were sprayed with 3 doses of rac-GR24 (1 μM, 3 μM and 5 μM), a synthesized strigolactone, and then were subjected to 7% (w/v) polyethylene glycol (PEG-6000) to simulate the drought conditions. Synthetic GR24 treated plants showed higher tolerance to drought stress with regard to lower electrolyte leakage, stomatal opening, reactive oxygen species (ROS), and higher relative water content, chlorophyll content, photosynthesis rate and malondialdehyde (MDA) content. GR24 application also decreased the levels of indoleacetic acid (IAA) and zeatin riboside (ZR), while increasing the level of abscisic acid (ABA), both in the roots and leaves under drought stress. These results suggested that foliar application of GR24 could ameliorate the adverse effects of drought due to its regulation of stomatal closure through ABA or ROS, and modulation of chlorophyll components and photosynthesis, as well as activation of the antioxidant defense capacity. Cross-talk with other hormones, especially ABA, was also suggested to be one of the important mechanism during this process. This study contributes to our current understanding of GR24-induced drought tolerance in grapevines.

A multifaceted rhizobacterium Bacillus licheniformis functions as a fungal antagonist and a promoter of plant growth and abiotic stress tolerance

Plant-growth-promoting rhizobacteria play a beneficial role in plant growth through various mechanisms. In this study, an isolated rhizobacterial strain, Bacillus licheniformis CH102 (hereafter CH102), exhibited antifungal activity and harbored genes required for lantibiotic and 2,3-butanediol production. In Arabidopsis, increased root hair number and fresh weight were obtained in the CH102-inoculated seedlings. Moreover, increases in chlorophyll content, H2O2 concentration and activities of antioxidant enzymes including catalase and POD were observed in the treated seedlings. Exposure to CH102 increased the tolerance toward high temperatures and water deficits in Arabidopsis seedlings. CH102 induced gene expression involved in induced disease resistance, nitrogen assimilation, antioxidants, photosystem II, ion transport and synthesis of secondary metabolites. Arabidopsis transcriptomic analysis revealed the CH102-upregulated genes were associated with several cellular pathways including the PEP-PEPR system, cell wall modification, and biosynthesis of osmoprotectants including choline, proline, and galactinol. The presence of CH102 elicited increased response of Arabidopsis genes associated with the jasmonic acid (JA) and abscisic acid (ABA) signaling pathways. Moreover, the upregulated expression of SLAC1 HOMOLOGUE 3 (SLAH3) function in the regulation of stomatal closure was detected. Furthermore, CH102 inoculation increased expression of transcription factors implicated in the regulation of abiotic stress responses, including DDF2, CBF1, DREB2A, HSFA2, WRKY30 and WRKY33. In summary, CH102 exerts effects on multiple cellular pathways involved in defense response, metabolism, antioxidant system and hormone signaling such as JA and ABA to regulate plant growth and tolerance to heat and drought stress.

Gene Expression Pattern and Protein Localization of Arabidopsis Phospholipase D Alpha 1 Revealed by Advanced Light-Sheet and Super-Resolution Microscopy.

Phospholipase D alpha 1 (PLDα1, At3g15730) and its product phosphatidic acid (PA) are involved in a variety of cellular and physiological processes, such as cytoskeletal remodeling, regulation of stomatal closure and opening, as well as biotic and abiotic stress signaling. Here we aimed to study developmental expression patterns and subcellular localization of PLDα1 in Arabidopsis using advanced microscopy methods such as light-sheet fluorescence microscopy (LSFM) and structured illumination microscopy (SIM). We complemented two knockout pldα1 mutants with a YFP-tagged PLDα1 expressed under the PLDα1 native promoter in order to study developmental expression pattern and subcellular localization of PLDα1 in Arabidopsis thaliana under natural conditions. Imaging of tissue-specific and developmentally-regulated localization of YFP-tagged PLDα1 by LSFM in roots of growing seedlings showed accumulation of PLDα1-YFP in the root cap and the rhizodermis. Expression of PLDα1-YFP in the rhizodermis was considerably higher in trichoblasts before and during root hair formation and growth. Thus, PLDα1-YFP accumulated in emerging root hairs and in the tips of growing root hairs. PLDα1-YFP showed cytoplasmic subcellular localization in root cap cells and in cells of the root transition zone. In aerial parts of plants PLDα1-YFP was also localized in the cytoplasm showing enhanced accumulation in the cortical cytoplasmic layer of epidermal non-dividing cells of hypocotyls, leaves, and leaf petioles. However, in dividing cells of root apical meristem and leaf petiole epidermis PLDα1-YFP was enriched in mitotic spindles and phragmoplasts, as revealed by co-visualization with microtubules. Finally, super-resolution SIM imaging revealed association of PLDα1-YFP with both microtubules and clathrin-coated vesicles (CCVs) and pits (CCPs). In conclusion, this study shows the developmentally-controlled expression and subcellular localization of PLDα1 in dividing and non-dividing Arabidopsis cells.

Mitochondrial pyruvate carrier 1 mediates abscisic acid-regulated stomatal closure and the drought response by affecting cellular pyruvate content in Arabidopsis thaliana

BackgroundStomata are micropores surrounded by pairs of guard cells, and their opening is finely controlled to balance water vapor loss as transpiration and CO2 absorption for photosynthesis. The regulatory signaling network for stomatal movement is complicated, and increasing numbers of new genes have been shown to be involved in this process. Our previous study indicated that a member of the plant putative mitochondrial pyruvate carrier (MPC) family, NRGA1, is a negative regulator of guard cell abscisic acid (ABA) signaling. In this study, we identified novel physiological roles of pyruvate and MPC1, another member of the MPC family, in the regulation of stomatal closure in Arabidopsis.ResultsLoss-of-function mutants of MPC1 (mpc1) were hypersensitive to ABA-induced stomatal closure and ABA-activated guard cell slow-type anion currents, and showed a reduced rate of water loss upon drought treatment compared with wild-type plants. In contrast, plants overexpressing MPC1 showed a hyposensitive ABA response and increased sensitivity to drought stress. In addition, mpc1 mutants accumulated more pyruvate after drought or ABA treatment. The increased pyruvate content also induced stomatal closure and activated the slow-type anion channels of guard cells, and this process was dependent on the function of RbohD/F NADPH oxidases and reactive oxygen species concentrations in guard cells.ConclusionsOur findings revealed the essential roles of MPC1 and pyruvate in stomatal movement and plant drought resistance.

Biology of SLAC1-type anion channels - from nutrient uptake to stomatal closure.

Contents 46 I. 46 II. 47 III. 50 IV. 53 V. 56 VI. 57 58 58 References 58 SUMMARY: Stomatal guard cells control leaf CO2 intake and concomitant water loss to the atmosphere. When photosynthetic CO2 assimilation is limited and the ratio of CO2 intake to transpiration becomes suboptimal, guard cells, sensing the rise in CO2 concentration in the substomatal cavity, deflate and the stomata close. Screens for mutants that do not close in response to experimentally imposed high CO2 atmospheres identified the guard cell-expressed Slowly activating anion channel, SLAC1, as the key player in the regulation of stomatal closure. SLAC1 evolved, though, before the emergence of guard cells. In Arabidopsis, SLAC1 is the founder member of a family of anion channels, which comprises four homologues. SLAC1 and SLAH3 mediate chloride and nitrate transport in guard cells, while SLAH1, SLAH2 and SLAH3 are engaged in root nitrate and chloride acquisition, and anion translocation to the shoot. The signal transduction pathways involved in CO2 , water stress and nutrient-sensing activate SLAC/SLAH via distinct protein kinase/phosphatase pairs. In this review, we discuss the role that SLAC/SLAH channels play in guard cell closure, on the one hand, and in the root-shoot continuum on the other, along with the molecular basis of the channels' anion selectivity and gating.

Mitogen-activated protein kinase 4-like carrying an MEY motif instead of a TXY motif is involved in ozone tolerance and regulation of stomatal closure in tobacco.

The mitogen-activated protein kinases (MAPKs/MPKs) are important factors in the regulation of signal transduction in response to biotic and abiotic stresses. Previously, we characterized a MAPK from tobacco, Nicotiana tabacum MPK4 (NtMPK4). Here, we found a highly homologous gene, NtMPK4-like (NtMPK4L), in tobacco as well as other species in Solanaceae and Gramineae. Deduced amino acid sequences of their translation products carried MEY motifs instead of conserved TXY motifs of the MAPK family. We isolated the full length NtMPK4L gene and examined the physiological functions of NtMPK4L. We revealed that NtMPK4L was activated by wounding, like NtMPK4. However, a constitutively active salicylic acid-induced protein kinase kinase (SIPKK(EE)), which phosphorylates NtMPK4, did not phosphorylate NtMPK4L. Moreover, a tyrosine residue in the MEY motif was not involved in NtMPK4L activation. We also found that NtMPK4L-silenced plants showed rapid transpiration caused by remarkably open stomata. In addition, NtMPK4L-silenced plants completely lost the ability to close stomata upon ozone treatment and were highly sensitive to ozone, suggesting that this atypical MAPK plays a role in ozone tolerance through stomatal regulation.

Arabidopsis HY1-Modulated Stomatal Movement: An Integrative Hub Is Functionally Associated with ABI4 in Dehydration-Induced ABA Responsiveness.

Heme oxygenase (HO; EC 1.14.99.3) has recently been proposed as a novel component in mediating wide ranges of the plant adaptive signaling processes. However, the physiological significance and molecular basis underlying Arabidopsis (Arabidopsis thaliana) HO1 (HY1) functioning in drought tolerance remained unclear. Here, we report that mutation of HY1 promoted, but overexpression of this gene impaired, Arabidopsis drought tolerance. This was attributed to the abscisic acid (ABA)-hypersensitive or -hyposensitive phenotypes, with the regulation of stomatal closure in particular. However, comparative transcriptomic profile analysis showed that the induction of numerous ABA/stress-dependent genes in dehydrated wild-type plants was differentially impaired in the hy1 mutant. In agreement, ABA-induced ABSCISIC ACID-INSENSITIVE4 (ABI4) transcript accumulation was strengthened in the hy1 mutant. Genetic analysis further identified that the hy1-associated ABA hypersensitivity and drought tolerance were arrested in the abi4 background. Moreover, the promotion of ABA-triggered up-regulation of RbohD abundance and reactive oxygen species (ROS) levels in the hy1 mutant was almost fully blocked by the mutation of ABI4, suggesting that the HY1-ABI4 signaling in the wild type involved in stomatal closure was dependent on the RbohD-derived ROS production. However, hy1-promoted stomatal closure was not affected by a nitric oxide scavenger. Correspondingly, ABA-insensitive behaviors in rbohD stomata were not affected by either the mutation of HY1 or its ectopic expression in the rbohD background, both of which responded significantly to exogenous ROS. These data indicate that HY1 functioned negatively and acted upstream of ABI4 in drought signaling, which was casually dependent on the RbohD-derived ROS in the regulation of stomatal closure.

Arabidopsis RZFP34/CHYR1, a Ubiquitin E3 Ligase, Regulates Stomatal Movement and Drought Tolerance via SnRK2.6-Mediated Phosphorylation

Abscisic acid (ABA) is a phytohormone that plays a fundamental role in plant development and stress response, especially in the regulation of stomatal closure in response to water deficit stress. The signal transduction that occurs in response to ABA and drought stress is mediated by protein phosphorylation and ubiquitination. This research identified Arabidopsis thaliana RING ZINC-FINGER PROTEIN34 (RZP34; renamed here as CHY ZINC-FINGER AND RING PROTEIN1 [CHYR1]) as an ubiquitin E3 ligase. CHYR1 expression was significantly induced by ABA and drought, and along with its corresponding protein, was expressed mainly in vascular tissues and stomata. Analysis of CHYR1 gain-of-function and loss-of-function plants revealed that CHYR1 promotes ABA-induced stomatal closure, reactive oxygen species production, and plant drought tolerance. Furthermore, CHYR1 interacted with SNF1-RELATED PROTEIN KINASE2 (SnRK2) kinases and could be phosphorylated by SnRK2.6 on the Thr-178 residue. Overexpression of CHYR1(T178A), a phosphorylation-deficient mutant, interfered with the proper function of CHYR1, whereas CHYR1(T178D) phenocopied the gain of function of CHYR1. Thus, this study identified a RING-type ubiquitin E3 ligase that functions positively in ABA and drought responses and detailed how its ubiquitin E3 ligase activity is regulated by SnRK2.6-mediated protein phosphorylation.

Towards Understanding Extracellular ROS Sensory and Signaling Systems in Plants

Reactive Oxygen Species (ROS) are ubiquitous metabolites in all aerobic organisms. Traditionally ROS have been considered as harmful, accidental byproducts of cellular functions involving electron transport chains or electron transfer. However, it is now recognized that controlled production of ROS has significant signaling functions, for example, in pathogen defense, in the regulation of stomatal closure, or in cell-to-cell signaling. ROS formation in subcellular compartments is critical to act as “alarm” signal in the response to stress, and the concept of ROS as primarily signaling substances has emerged. The involvement of ROS in several developmental and inducible processes implies that there must be coordinated function of signaling network(s) that govern ROS responses and subsequent processes. The air pollutant ozone can be used as a useful tool to elucidate the function of apoplastic ROS: O3degrades in cell wall into various ROS which are interpreted as ROS with signaling function inducing downstream responses. We have used ozone as a tool in mutant screens and transcript profiling-reverse genetics to identify genes involved in processes related to the signaling function of ROS. We review here our recent findings in the elucidation of apoplastic ROS sensing, signaling, and interaction with various symplastic components.

Ethylene-Induced Flavonol Accumulation in Guard Cells Suppresses Reactive Oxygen Species and Moderates Stomatal Aperture

Guard cell swelling controls the aperture of stomata, pores that facilitate gas exchange and water loss from leaves. The hormone abscisic acid (ABA) has a central role in regulation of stomatal closure through synthesis of second messengers, which include reactive oxygen species (ROS). ROS accumulation must be minimized by antioxidants to keep concentrations from reaching damaging levels within the cell. Flavonols are plant metabolites that have been implicated as antioxidants; however, their antioxidant activity in planta has been debated. Flavonols accumulate in guard cells of Arabidopsis thaliana, but not surrounding pavement cells, as visualized with a flavonol-specific dye. The expression of a reporter driven by the promoter of CHALCONE SYNTHASE, a gene encoding a flavonol biosynthetic enzyme, in guard cells, but not pavement cells, suggests guard cell-specific flavonoid synthesis. Increased levels of ROS were detected using a fluorescent ROS sensor in guard cells of transparent testa4-2, which has a null mutation in CHALCONE SYNTHASE and therefore synthesizes no flavonol antioxidants. Guard cells of transparent testa4-2 show more rapid ABA-induced closure than the wild type, suggesting that flavonols may dampen the ABA-dependent ROS burst that drives stomatal closing. The levels of flavonols are positively regulated in guard cells by ethylene treatment in the wild type, but not in the ethylene-insensitive2-5 mutant. In addition, in both ethylene-overproducing1 and ethylene-treated wild-type plants, elevated flavonols lead to decreasing ROS and slower ABA-mediated stomatal closure. These results are consistent with flavonols suppressing ROS accumulation and decreasing the rate of ABA-dependent stomatal closure, with ethylene-induced increases in guard cell flavonols modulating these responses.

Regulation of water, salinity, and cold stress responses by salicylic acid

Salicylic acid (SA) is a naturally occurring phenolic compound. SA plays an important role in the regulation of plant growth, development, ripening, and defense responses. The role of SA in the plant–pathogen relationship has been extensively investigated. In addition to defense responses, SA plays an important role in the response to abiotic stresses, including drought, low temperature, and salinity stresses. It has been suggested that SA has great agronomic potential to improve the stress tolerance of agriculturally important crops. However, the utility of SA is dependent on the concentration of the applied SA, the mode of application, and the state of the plants (e.g., developmental stage and acclimation). Generally, low concentrations of applied SA alleviate the sensitivity to abiotic stresses, and high concentrations of applied induce high levels of oxidative stress, leading to a decreased tolerance to abiotic stresses. In this article, the effects of SA on the water stress responses and regulation of stomatal closure are reviewed.

PYR/PYL/RCAR abscisic acid receptors regulate K+ and Cl- channels through reactive oxygen species-mediated activation of Ca2+ channels at the plasma membrane of intact Arabidopsis guard cells.

The discovery of the START family of abscisic acid (ABA) receptors places these proteins at the front of a protein kinase/phosphatase signal cascade that promotes stomatal closure. The connection of these receptors to Ca(2+) signals evoked by ABA has proven more difficult to resolve, although it has been implicated by studies of the pyrbactin-insensitive pyr1/pyl1/pyl2/pyl4 quadruple mutant. One difficulty is that flux through plasma membrane Ca(2+) channels and Ca(2+) release from endomembrane stores coordinately elevate cytosolic free Ca(2+) concentration ([Ca(2+)]i) in guard cells, and both processes are facilitated by ABA. Here, we describe a method for recording Ca(2+) channels at the plasma membrane of intact guard cells of Arabidopsis (Arabidopsis thaliana). We have used this method to resolve the loss of ABA-evoked Ca(2+) channel activity at the plasma membrane in the pyr1/pyl1/pyl2/pyl4 mutant and show the consequent suppression of [Ca(2+)]i increases in vivo. The basal activity of Ca(2+) channels was not affected in the mutant; raising the concentration of Ca(2+) outside was sufficient to promote Ca(2+) entry, to inactivate current carried by inward-rectifying K(+) channels and to activate current carried by the anion channels, both of which are sensitive to [Ca(2+)]i elevations. However, the ABA-dependent increase in reactive oxygen species (ROS) was impaired. Adding the ROS hydrogen peroxide was sufficient to activate the Ca(2+) channels and trigger stomatal closure in the mutant. These results offer direct evidence of PYR/PYL/RCAR receptor coupling to the activation by ABA of plasma membrane Ca(2+) channels through ROS, thus affecting [Ca(2+)]i and its regulation of stomatal closure.

Overexpression of the trehalase gene AtTRE1 leads to increased drought stress tolerance in Arabidopsis and is involved in abscisic acid-induced stomatal closure.

Introduction of microbial trehalose biosynthesis enzymes has been reported to enhance abiotic stress resistance in plants but also resulted in undesirable traits. Here, we present an approach for engineering drought stress tolerance by modifying the endogenous trehalase activity in Arabidopsis (Arabidopsis thaliana). AtTRE1 encodes the Arabidopsis trehalase, the only enzyme known in this species to specifically hydrolyze trehalose into glucose. AtTRE1-overexpressing and Attre1 mutant lines were constructed and tested for their performance in drought stress assays. AtTRE1-overexpressing plants had decreased trehalose levels and recovered better after drought stress, whereas Attre1 mutants had elevated trehalose contents and exhibited a drought-susceptible phenotype. Leaf detachment assays showed that Attre1 mutants lose water faster than wild-type plants, whereas AtTRE1-overexpressing plants have a better water-retaining capacity. In vitro studies revealed that abscisic acid-mediated closure of stomata is impaired in Attre1 lines, whereas the AtTRE1 overexpressors are more sensitive toward abscisic acid-dependent stomatal closure. This observation is further supported by the altered leaf temperatures seen in trehalase-modified plantlets during in vivo drought stress studies. Our results show that overexpression of plant trehalase improves drought stress tolerance in Arabidopsis and that trehalase plays a role in the regulation of stomatal closure in the plant drought stress response.