Major positive and negative regulators of ABA signaling
The ability of plants to respond to environmental stimuli is essential to plant survival. Abscisic acid (ABA) is a phytohormone with roles at various stages of plant development. ABA also plays a major role in mediating physiological responses to environmental stresses such as salt, osmotic, and cold stress. Plant responses to environmental stress have been widely studied in the model plant Arabidopsis thaliana and ABA signaling mechanisms elucidated. In general, the adaptive responses of plants to various stress conditions can be either ABA-dependent or ABA-independent. Here we focus on the role of ABA in stress signaling and abiotic stress tolerance. We describe the intrinsic mechanisms that confer stress tolerance via ABA, as well as how ABA-regulated gene products play a role in salt and drought tolerance at different stages of the life cycle. In addition, the contribution of ABA to regulation of stomatal aperture and therefore desiccation tolerance will be discussed. Understanding ABA signaling mechanisms in abiotic stress provides avenues for improving plant performance.
- ABA signaling
Due to their sessile nature, plants cannot avoid environmental stresses, thus they have evolved mechanisms to overcome the detrimental effects of stress. For example, plant endogenous developmental programs are modified such that structural and metabolic changes assist to overcome adverse environmental conditions such as salinity and drought. Failure to adapt to adverse environmental conditions can significantly reduce yield by impacting plant development and productivity. Abiotic stress conditions initiate a number of molecular, biochemical, and physiological changes at both the cellular and whole plant levels . One major biochemical change in response to stress is elevation of abscisic acid (ABA) levels, which in turn triggers expression of a cascade of stress-responsive genes . Cellular ABA levels are induced by environmental stimuli such as light, water, and salinity stress .
The plant hormone ABA has been identified as a key regulator of multiple stresses. In general, the adaptive responses of plants to various stress conditions can be either ABA-dependent or ABA-independent. However, there is no clear boundary between these two pathways and there is lot of crosstalk between the pathways and the components involved. This review will focus on recent advancements in ABA-mediated stress signaling and the role of ABA in abiotic stress tolerance in the model plant
2. The phytohormone Abscisic Acid (ABA)
ABA, a sesquiterpenoid (C15H20O4) with a 15-carbon ring (Figure 1), has a variety of biological functions and is found ubiquitously across several kingdoms, including cyanobacteria, sponges, algae, lichens, mosses, and mammals [4-7]. Discovered in the 1960s and initially named dormin or abscissin, ABA is now established as a widely occurring and important plant growth regulator. Although it was initially identified as an abscission-promoting hormone, later scientists discovered that this was partly due to an indirect effect of inducing ethylene biosynthesis . ABA is an important regulator of plant growth, including embryo and seed development, seedling establishment, vegetative and reproductive growth as well as promoting seed dormancy [9,10]. Seed maturation and promotion of dormancy are important in preventing preharvest sprouting. In addition, ABA has the ability to antagonize the germination promoting effects of gibberellin, regulate guard cells, and regulate stress-responsive gene expression under water-deprived conditions. ABA also has a role in plant pathogen responses in a pathosystem-dependent manner [4,5].
The molecular structure of ABA has several important features that facilitate its biological functions. The side chain with the two double bonds (Figure 1) and ABA’s stereocenter are two such important features. Exposure to UV light changes the conformation from active to inactive form .
2.1. ABA signaling in plants
Although ABA has a broad range of functions in plant growth and development, its main function is to regulate plant water balance and osmotic stress tolerance . Thus, understanding ABA signaling is essential to improving plant performance. Genetic screens in
|ABA INSENSITIVE 1/2 (ABI1/2)||Various tissues and developmental stages||[13-16]|
|ABA HYPERSENSITIVE GERMINATION 1 (AHG1)|
|HYPERSENSITIVE TO ABA 1/2 (HAB1/2)|||
|SRK2D/SnRK2.2||Seeds and vegetative tissues|||
|SRK2I/SnRK2.3||Seeds and vegetative tissues|||
|SRK2E/OST1/SnRK2.6||Expressed in guard cells and involved in stomatal closure|||
Therefore, the ABA signaling complex/ABA signalosome is comprised of three major components: (a) RCAR/PYR/PYLs; (b) PP2Cs; and (c) SnRK2s assembled as a double negative regulatory system . In the absence of ABA, PP2Cs dephosphorylate SnRK2s inhibiting kinase activity and thereby preventing downstream gene expression (Figure 2). Several studies showed that these core components are essential for ABA signaling. For instance, Fujita
In guard cells (Figure 3), ABA binds to the PYR/PYL/RCAR receptor-PP2C complex and blocks its phosphatase activity. Consequently, activated protein kinase SnRK2.6/OPEN STOMATA 1 (OST1) phosphorylates and regulates the key target ion channels, SLOW ANION CHANNEL ASSOCIATED 1 (SLAC1) and K+ CHANNEL IN ARABIDOPSIS THALIANA 1 (KAT1). SnRK2.6/OST1 acts as a positive regulator of stomatal closure where it activates anion channel SLAC1 and inhibits cation channel KAT1 [23-25].
2.2. ABA-binding proteins and alternate ABA receptors
Identification of putative ABA receptors using forward genetic approaches was not successful for a long time due to genetic redundancy of the genes encoding ABA receptor proteins. However, biochemical approaches leading to purification and analysis of high-affinity ABA-binding proteins have been successful in identification of potential ABA receptor classes [5,26]. Some of these potential ABA receptors are cytosolic while others are on the cell surface, indicating there are extracellular as well as intracellular sites of ABA perception. Studies indicate that there can be multiple ABA receptors at different locations of the cell .
2.2.1. ChlH/ABAR (H subunit of the chloroplast magnesium chelatase/ ABA receptor)
The primary function of ChlH is chlorophyll synthesis. ChIH was initially identified as an ABA-binding protein in broad bean (
2.2.2. GTG1/GTG2 (G protein coupled receptor type G protein 1 and 2)
G protein coupled ABA receptors are plasma membrane localized cell surface receptors that are widely expressed in plants. Both GTG1 and GTG2 showed specific and saturable ABA-binding activity in direct ABA-binding assays. GTGs have GTPase activity and GDP bound GTGs have enhanced ABA-binding ability, which in turn initiates ABA signaling. GTG1/2 bind with GPA1 (G-PROTEIN α SUBUNIT 1), which abolishes its GTPase activity and represses ABA binding. GTP bound GPA1 represses ABA signalling. However, the downstream components of this pathway are not characterized yet [31,32].
2.3. Recent studies on ABA perception and signaling mechanisms
Recent findings that several ABA receptors exist in different parts of the cell provide evidence that ABA is active in a variety of subcellular compartments. ABA synthesis enzymes are present in different compartments, suggesting that ABA synthesis occurs in different parts of the cell and that these ABA levels contribute to overall ABA homeostasis. For example, the ABA biosynthesis enzyme AtABA1 is localized in the chloroplast, whereas AtABA2 is in the cytosol [33,34]. It has also been proposed that ABA produced in cytoplasm, plastids, vacuole, and other subcellular organelles may have different physiological roles initiated by signaling networks via different ABA receptors in each specific compartment .
Inhibition of PP2Cs results in autoactivation of SnRK2 kinases and thereby positive regulation of ABA signaling. Recently, the crystal structures of SnRK2.3 and SnRK2.6 were elucidated, providing evidence that kinase activation is a two-step mechanism as well as details of how the ABA signal is transmitted to downstream components . This study also showed that autophosphorylation of SnRK2.6 is more efficient than that of SnRK2.3.
3. ABA in stress signaling
In plants and other organisms, such as algae, cyanobacteria, and fungi, ABA levels tend to increase with exposure to stress, suggesting a potential role of ABA in stress signal transduction . Exogenous ABA application mimics stress conditions in plants and provides a useful means to study the effect of ABA on stress signaling and tolerance . ABA distributes throughout the plant as an inactive glucose sugar conjugate and is converted to the active form by β-glucosidase . ABA acts as an endogenous messenger and salt and drought stress signal transmission to initiate downstream gene expression occurs mainly via ABA signaling. However, cold stress signal transduction occurs in an ABA-independent manner via the C-REPEAT/DEHYDRATION RESPONSIVE ELEMENT BINDING FACTORS (CBFs/DREBs) signaling pathway .
A large number of ABA-responsive genes have a common
There are nine Group A bZIP transcription factors implicated in ABA signaling and they are subdivided into two groups based on where they are mainly expressed. The
SnRK2 III is the major subfamily of SnRKs involved in abiotic stress responses. SnRK2 protein kinases phosphorylate AREB/ABFs and regulate their function in ABA-regulated gene expression under stress conditions . SnRK2.6/OST1 is an important regulator of stomatal closure in drought stress. The role of SnRK2.2 and 2.3 is mainly to transmit the ABA signal to inhibit seed germination and seedling growth in response to stress. A decuple
3.1. ABA and abiotic stress tolerance
Drought and high salinity generate osmotic stress in plant cells. Endogenous ABA levels are elevated in response to osmotic stress, which in turn coordinates the plant’s response to reduced water availability. In addition, seed maturation and post-germinative growth creates cellular dehydration stress, which again results in accumulation of ABA in cells . The role of ABA in drought and salt stress is twofold: water balance and cellular dehydration tolerance. Water balance is achieved through guard cell regulation and the latter role by induction of genes that encode dehydration tolerance proteins in nearly all cells. ABA accumulation is induced by osmotic stress and this is as a result of activation of ABA biosynthesis as well as inhibition of ABA degradation . Thus, ABA-mediated adaptive stress responses of plants to environmental stimuli occur via ABA-responsive gene expression and regulation of stomatal pore size. ABA-responsive gene expression involves various transcription factors, ABA receptors, secondary messengers, protein kinase/phosphatase cascades, and chromatin remodeling factors .
Both drought stress and salinity stress upregulate osmotic stress responsive genes that are ABA-inducible. Most of the high-salinity-induced genes are also induced by drought, suggesting there is overlap between salt and drought stress tolerance mechanisms . A large number of transcription factors are induced by multiple stress conditions. AREB1/ABF2, AREB2/ABF4, ABF3, and MYB41 are some of the main transcription factors that are induced by both salt and drought in vegetative tissues .
Drought and salt stress results in osmotic imbalance; thus, salt and drought stress tolerance mechanisms aim at restoring cellular homeostasis. These mechanisms are adaptive responses that create either stress tolerance or avoidance of stress conditions. Modifications in metabolic pathways, synthesis of new proteins, changes in ion uptake, and free radical scavenging are some of the stress responses at the cellular level, immediately followed by stress signal transduction . High ABA levels in cells result in synthesis of storage proteins, desiccation tolerance, and dormancy via inhibition of seed germination . In the plant as a whole, key adaptive responses include induction of stomatal closure as well as control of seedling growth and lateral root formation. While the balance between ABA and auxin levels slightly affects primary root growth, ABA represses lateral root formation while auxin promotes it [4,48].
Inhibition of seed germination under abiotic stress is another function of ABA. Seed germination occurs when there is a balance between germination-promoting gibberellin and dormancy-promoting ABA. During late stages of maturation, seeds accumulate ABI5 which in turn activates transcription of LATE EMBRYOGENESIS ABUNDANT (LEA) proteins. LEA proteins confer osmotolerance to the embryo. ABA is necessary for activation of ABI5 via SnRK2.2 and SnRK2.3 phosphorylation of ABI5 [49,50]. When seeds are in unfavorable environmental conditions, elevated endogenous ABA levels results in ABI5 accumulation, preventing seeds from germinating.
3.1.1. ABA and salt tolerance
Salt stress severely impacts plant growth by affecting metabolic processes and photosynthetic efficiency. NaCl initially induces osmotic stress and eventually accumulation of both Na+ and Cl- ions generates ionic stress . However, some responses are salt-specific and distinct from responses to osmotic stress (reviewed in ). High salinity in the soil is first sensed by the plant roots. Salt and drought stress induce a rapid increase in cytosolic Ca2+ levels in the root cells. Ca2+ acts a second messenger, inducing salt- and drought-responsive genes [53,54]. Hyperosmotic stress is coupled with Ca2+ signaling and Reactive Oxygen Species (ROS) signaling, thereby inducing a cascade of signaling events, which results in downstream gene expression .
Biochemical and molecular mechanisms of salt tolerance in plants include exclusion of salt ions, production of suitable osmolytes, changing the structure of the membranes to control ion uptake, and induction of enzymes that produce antioxidants and phytohormones. To manage salt or drought stress, cellular ABA levels increase dramatically. The plant cuticle has been shown to mediate stress signaling as well as ABA biosynthesis and signaling. In addition to its primary function, providing mechanical support to the cell wall and plasma membrane, the cuticle has been implicated in osmotic stress regulation. CED1 (9-CIS EPOXYCAROTENOID DIOXYGENASE DEFECTIVE 1) is an essential protein in cuticle biogenesis.
ABA regulates root growth and architecture in plants under stress. Duan
ABA regulates expression of many salt-stress-responsive genes via transcription factors that are elevated in response to salt. For instance, ABF2/AREB1, ABF3, ABF4/AREB2, ABRE BINDING PROTEIN 9 (ABP9), and MYC/MYB, WRKY, and APETALA2/ETHYLENE RESPONSE FACTOR (AP2/ERF) are some of the salt-stress-responsive transcription factors that enhance stress tolerance. A recent study showed that the PYL8/RCAR3 ABA receptor has a role in ABA-mediated inhibition of primary root growth and also recovery of lateral root growth on exposure to ABA. PYL8/RCAR3 combines the action of ABA and auxin through direct interaction with MYB transcription factors during growth recovery of postemergence lateral roots [48,57].
There are proteins in the cell that are produced in an ABA-dependent manner that have a role in osmotic tolerance. For example, ABI5 activates transcription of LEA proteins. LEA proteins are highly hydrophilic small proteins shown to have an osmoprotectant role against cellular dehydration during late embryogenesis. LEA proteins also have a role in salt stress tolerance . Due to their hydrophilic nature, LEA proteins can sequester ions accumulating in the cell, as well as act as chaperones and retain water molecules to prevent protein aggregation and inactivation of cellular enzymes . In Arabidopsis, 51 LEA proteins have been identified that belong to nine different groups . Jia
In addition, ABA has been implicated in histone H3 acetylation and methylation, thereby regulating stress-inducible gene expression at the epigenetic level. Chen
3.1.2. ABA and drought tolerance
Drought is lack of water in the soil. Drought stress in plants arises due to water deficit conditions and results in removal of water from the cell membranes, disrupting the lipid bilayer structure. In addition, protein denaturation and accumulation of cellular electrolytes results in disruption of cellular metabolism . Therefore, drought causes osmotic stress, and osmotic stress causes dehydration and inhibition of water uptake in plants. ABA accumulates under osmotic stress conditions and plays an important role in the stress response and tolerance of plants. In addition to autoactivation of SnRK2s by inhibition of PP2Cs in the ABA signaling cascade, hyperosmotic stress activates SnRK2s . SnRK2 kinases are a major component of the osmotic stress signaling pathway. The Arabidopsis triple mutant
ABA induces expression of many transcription factors as well as genes that encode enzymes in the synthesis of osmoprotectants . Osmolytes are compatible solutes such as amino acids (proline), sugar alcohols (mannitol, pinitol), and other sugars that accumulate without disrupting the function of proteins. Osmolytes make an osmotic adjustment facilitating a favorable water potential gradient and promote stress tolerance .
Dehydrins and LEA-like proteins act as cellular chaperones that protect cellular membranes and macromolecules in the cell . During seed maturation seeds undergo dehydration stress. LEA proteins accumulate in the embryo as a result of osmotic stress and their functions include protection of enzymes, lipids, and mRNAs from dehydration. LEA proteins have been found to protect mitochondrial membranes from damage. LEA proteins are produced in an ABA-dependent and ABA-independent manner under osmotic stress [47,58].
Under moderate water stress conditions plant root growth has to be maintained in order to keep the plants alive. ABA accumulates under moderate water stress and mediates auxin transport in the root tip, which enhances the proton pumps in the plasma membrane. Proton secretions in the root tip play an important role in primary root growth and root hair development under moderate drought stress .
Based on the critical water level, drought tolerance is considered to be mechanisms that confer tolerance to moderate dehydration. Further dehydration requires desiccation tolerance mechanisms in order to restore the ability of cells to rehydrate successfully .
3.1.3. ABA and desiccation tolerance
Water loss results in a change in turgor pressure that affects the cell walls. Desiccation tolerance is defined as evolution of cell walls that can withstand extensive water loss without damaging its structure or polymer organization. Desiccation tolerance mechanisms aim to restructure the cells walls and maintain normal growth under water stress conditions .
Regulation of the stomatal pore is crucial in adapting plants to abiotic stress by reducing extensive water loss. Stomatal opening and closing occurs as a result of turgor pressure differences in the surrounding guard cells . In response to water stress, ABA concentration is increased in the guard cell cytoplasm and apoplast, which results in a decrease in the turgor pressure due to activation of the K+ outward rectifying channel and inhibition of the K+ inward rectifying channel (KAT1 and 2). ABA also induces the anion channel SLAC1 resulting in release of anionic organic acids from the vacuole to the cytoplasm . Reduced turgor pressure initiates closure of stomata as a mechanism of minimizing water loss from the plant. ABA levels rise in leaves immediately following water stress. CHLH/ABAR has been proposed as the chloroplast ABA receptor that links ABA signaling within the chloroplast with ABA signaling in the nucleus. Overexpression of CHLH promotes stomatal closure and thereby dessication tolerance .
SnRK2 OPEN STOMATA 1 (OST1) is a key SnRK2 protein kinase involved in regulation of the stomatal aperture by movement of guard cells during ABA signaling . OST1 is activated by ABA, low humidity, and osmotic stress and is an important kinase found in guard cells preventing rapid water loss. Loss of function mutants of SnRK2 do not exhibit ABA-mediated stomatal closure activity and showed a wilty phenotype under dehydration stress conditions [72,73]. Also SnRK2.6/OST1 physically interacts with ABI1 and ABI2. ABI1 is required for ABA-dependent activation of OST1 and both ABI and ABI2 are required for osmotic-stress-induced activation of OST1 . Thus, SnRK2.6/OST1 acts as a positive regulator in ABA-induced stomatal closure. Moreover, Yoshida
Reactive Oxygen Species (ROS) have also been identified as secondary messengers in ABA signaling in guard cells. In Arabidopsis, two partially redundant guard cell expressed NADPH oxidase catalytic subunit genes,
ABA has a wide range of functions from plant development to biotic and abiotic stress signaling and tolerance. The primary functions of ABA in salt, drought, and desiccation tolerance act via inhibiting seed germination, altering root architecture, and inducing stress-responsive genes as well as gene products that act as osmoprotectants. ABA signaling cascades and stress tolerance mechanisms studied in Arabidopsis provide insight into application of stress tolerance strategies to commercial crops. While ABA is not the only plant hormone involved in stress responses, many of these responses occur in an ABA-dependent manner, indicating the importance of ABA in plant stress response and tolerance.