Negative Regulator Of Abscisic Acid Signaling Research Articles

Exogenous preculture with sucrose and abscisic acid improves post-cryopreservation survival of eastern bracken fern gametophytes

Cryopreservation is an important technique used in the conservation of various plant tissues. This study proposes a cryopreservation method for the long-term conservation of eastern bracken fern gametophytes (Pteridium aquilinum var. latiusculum). Encapsulation–dehydration of the gametophytes was performed, and the exogenous sucrose and abscisic acid (ABA) preculture conditions were investigated. Gametophytes are sensitive to dehydration and drying, and the following treatment conditions were applied: encapsulation by alginate containing 0.75 M sucrose, 18-h loading treatment with 0.75 M sucrose, and 6-h drying treatment. The survival rate following cryopreservation was determined. The water content of < 27.5% in the alginate beads after dehydration and drying was found to be appropriate for ensuring survival. Additionally, performing an exogenous sucrose and ABA preculture was essential before encapsulation to achieve a survival of ≥ 90%. The high stress induced by cryopreservation and exogenous preculture regulated the expression of PaSuSy, PaLEA14, and PaABI1b and the endogenous ABA content. In eastern bracken gametophytes, ABI1 appears to be a negative regulator of ABA signaling. These results indicate that the encapsulation–dehydration method is effective for the long-term conservation of eastern bracken fern gametophytes, and exogenous preculture alleviates abiotic stress and increases the survival rate.

Silencing ZmPP2C-A10 with a foxtail mosaic virus (FoMV) derived vector benefits maize growth and development following water limitation.

Global climate change is causing more frequent and severe droughts, which can have negative impacts on plant growth and crop productivity. Under drought conditions, plants produce the hormone ABA (abscisic acid), which regulates adaptive responses, such as stomatal closure and root elongation. Plant viruses have been used in the lab to convey new traits to plants and could also be used to increase production of ABA or to enhance downstream plant drought resistance responses. In this study, foxtail mosaic virus (FoMV) was used to silence ZmPP2C-A10, a negative regulator of ABA signalling, in maize (Zea mays L.). Both silenced and control plants were exposed to an 8-day drought treatment, followed by a 30-day period of rewatering, after which indicators of drought resistance were measured. After drought treatment, we observed a nearly twofold increase in expression of a stress-mitigation gene, ZmRAB17, reduced chlorophyll fluorescence changes (indicator of stress), and increased plant biomass and development in the ZmPP2C-A10-silenced maize compared to controls. These results demonstrate that the FoMV systemcan be used to silence endogenous expression of ZmPP2C-A10 and increase maize tolerance to drought. This could offer a useful tool to improve crop traits and reduce yield loss during the growing season.

StHAB1, a negative regulatory factor in abscisic acid signaling, plays crucial roles in potato drought tolerance and shoot branching.

Abscisic acid (ABA) is critical in drought tolerance and plant growth. Group A protein type 2C phosphatases (PP2Cs) are negative regulators of ABA signaling and plant adaptation to stress. Knowledge about the functions of potato group A PP2Cs is limited. Here, we report that the potato group A PP2C StHAB1 is broadly expressed in potato plants and strongly induced by ABA and drought. Suppression of StHAB1 enhanced potato ABA sensitivity and drought tolerance, whereas overexpression of the dominant mutant StHAB1G276D compromised ABA sensitivity and drought tolerance. StHAB1 interacts with almost all ABA receptors and the Snf1-Related Kinase OST1. Suppressing StHAB1 and overexpressing StHAB1G276D alter potato growth morphology; notably, overexpression of StHAB1G276D causes excessive shoot branching. RNA-sequencing analyses identified that the auxin efflux carrier genes StPIN3, StPIN5, and StPIN8 were up-regulated in StHAB1G276D-overexpressing axillary buds. Correspondingly, the auxin concentration was reduced in StHAB1G276D-overexpressing axillary buds, consistent with the role of auxin in repressing lateral branch outgrowth. The expression of BRANCHED1s (StBRC1a and StBRC1b) was unchanged in StHAB1G276D-overexpressing axillary buds, suggesting that StHAB1G276D overexpression does not cause axillary bud outgrowth via regulation of BRC1 expression. Our findings demonstrate that StHAB1 is vital in potato drought tolerance and shoot branching.

The unique sweet potato NAC transcription factor IbNAC3 modulates combined salt and drought stresses.

Plants often simultaneously experience combined stresses rather than a single stress, causing more serious damage, but the underlying mechanisms remain unknown. Here, we identified the stress-induced IbNAC3 from sweet potato (Ipomoea batatas) as a nucleus-localized transcription activator. IbNAC3 contains a unique activation domain whose MKD sequence confers transactivation activities to multiple other TFs and is essential for the activated expression of downstream target genes. Ectopic expression of IbNAC3 conferred tolerance to single and combined salt and drought stresses in Arabidopsis (Arabidopsis thaliana), and a group of NAM, ATAF1/2, and CUC2 (NAC) TFs, including ANAC011, ANAC072, ANAC083, ANAC100, and NAP, interacted with IbNAC3, and the specific domains responsible for each interaction varied. Intriguingly, IbNAC3 repressed the interaction among the five NACs, and knockout or mutation of ANAC011 and ANAC072 dramatically impaired combined stress tolerance. IbNAC3-ANAC072 and IbNAC3-NAP modules synergistically activated the MICROTUBULE-RELATED E3 LIGASE57 (MREL57) gene. Consistently, mutation of MREL57 and overexpression of WAVE-DAM-PENED2-LIKE7, encoding a target protein of MREL57, both remarkably impaired combined stress tolerance. Moreover, transgenic plants displayed abscisic acid (ABA) hyposensitivity by directly promoting the transcription of ENHANCED RESPONSE TO ABA 1, a key negative regulator of ABA signaling. The data unravel the unique IbNAC3 TF functions as a pivotal component in combined stress tolerance by integrating multiple regulatory events and ubiquitin pathways, which is essential for developing high-tolerant plants in natural environments.

Arabidopsis ACINUS is O-glycosylated and regulates transcription and alternative splicing of regulators of reproductive transitions

O-GlcNAc modification plays important roles in metabolic regulation of cellular status. Two homologs of O-GlcNAc transferase, SECRET AGENT (SEC) and SPINDLY (SPY), which have O-GlcNAc and O-fucosyl transferase activities, respectively, are essential in Arabidopsis but have largely unknown cellular targets. Here we show that AtACINUS is O-GlcNAcylated and O-fucosylated and mediates regulation of transcription, alternative splicing (AS), and developmental transitions. Knocking-out both AtACINUS and its distant paralog AtPININ causes severe growth defects including dwarfism, delayed seed germination and flowering, and abscisic acid (ABA) hypersensitivity. Transcriptomic and protein-DNA/RNA interaction analyses demonstrate that AtACINUS represses transcription of the flowering repressor FLC and mediates AS of ABH1 and HAB1, two negative regulators of ABA signaling. Proteomic analyses show AtACINUS’s O-GlcNAcylation, O-fucosylation, and association with splicing factors, chromatin remodelers, and transcriptional regulators. Some AtACINUS/AtPININ-dependent AS events are altered in the sec and spy mutants, demonstrating a function of O-glycosylation in regulating alternative RNA splicing.

Bacterial Quorum-Sensing Signaling-Related drr1 Mutant Influences Abscisic Acid Responsiveness in Arabidopsis thaliana L.

N-acyl-L-homoserine lactones (AHLs) are involved in cell-to-cell communication in Gram-negative bacteria through a process termed quorum-sensing (QS). In this report, we evaluated the response of Arabidopsis thaliana primary roots to abscisic acid (ABA) in wild-type (WT) and decanamide resistant root 1 (drr1) mutant, previously reported to be resistant to N-decanoyl-L-homoserine lactone (C10-HL). When compared to WT seedlings, drr1 mutants were hypersensitive to ABA and had primary roots shorter, which correlated with lower cell division in meristems, a higher concentration of endogenous ABA, and a greater expression of ABI5 gene; and this shortened primary root phenotype was reversed in drr1abi5 double mutants. An analysis of expression of ABSCISIC ACID INSENSITIVE 4 (ABI4) showed an ABA-inducible pattern in primary root tips, which was further increased in drr1 mutant seedlings. Comparison of seed germination in WT, drr1, abi5, and drr1abi5 lines showed higher germination percentages in the following order under control condition: abi5 > drr1abi5 > WT > drr1, while abi5 and drr1abi5 germinated faster and drr1 germinated slower with respect to WT under ABA condition. Taken together, our results suggest that DRR1 is a negative regulator of ABA signaling probably acting upstream of the transcription factors ABI4 and ABI5, which influence ABA responsiveness in primary roots and seed germination.

AcoMYB4, an Ananas comosus L. MYB Transcription Factor, Functions in Osmotic Stress through Negative Regulation of ABA Signaling.

Drought and salt stress are the main environmental cues affecting the survival, development, distribution, and yield of crops worldwide. MYB transcription factors play a crucial role in plants’ biological processes, but the function of pineapple MYB genes is still obscure. In this study, one of the pineapple MYB transcription factors, AcoMYB4, was isolated and characterized. The results showed that AcoMYB4 is localized in the cell nucleus, and its expression is induced by low temperature, drought, salt stress, and hormonal stimulation, especially by abscisic acid (ABA). Overexpression of AcoMYB4 in rice and Arabidopsis enhanced plant sensitivity to osmotic stress; it led to an increase in the number stomata on leaf surfaces and lower germination rate under salt and drought stress. Furthermore, in AcoMYB4 OE lines, the membrane oxidation index, free proline, and soluble sugar contents were decreased. In contrast, electrolyte leakage and malondialdehyde (MDA) content increased significantly due to membrane injury, indicating higher sensitivity to drought and salinity stresses. Besides the above, both the expression level and activities of several antioxidant enzymes were decreased, indicating lower antioxidant activity in AcoMYB4 transgenic plants. Moreover, under osmotic stress, overexpression of AcoMYB4 inhibited ABA biosynthesis through a decrease in the transcription of genes responsible for ABA synthesis (ABA1 and ABA2) and ABA signal transduction factor ABI5. These results suggest that AcoMYB4 negatively regulates osmotic stress by attenuating cellular ABA biosynthesis and signal transduction pathways.

ABA-dependent bZIP transcription factor, CsbZIP18, from Camellia sinensis negatively regulates freezing tolerance in Arabidopsis.

Overexpression of the tea plant gene CsbZIP18 in Arabidopsis impaired freezing tolerance, and CsbZIP18 is a negative regulator of ABA signaling and cold stress. Basic region/leucine zipper (bZIP) transcription factors play important roles in the abscisic acid (ABA) signaling pathway and abiotic stress response in plants. However, few bZIP transcription factors have been functionally characterized in tea plants (Camellia sinensis). In this study, a bZIP transcription factor, CsbZIP18, was found to be strongly induced by natural cold acclimation, and the expression level of CsbZIP18 was lower in cold-resistant cultivars than in cold-susceptible cultivars. Compared with wild-type (WT) plants, Arabidopsis plants constitutively overexpressing CsbZIP18 exhibited decreased sensitivity to ABA, increased levels of relative electrolyte leakage (REL) and reduced values of maximal quantum efficiency of photosystem II (Fv/Fm) under freezing conditions. The expression of ABA homeostasis- and signal transduction-related genes and abiotic stress-inducible genes, such as RD22, RD26 and RAB18, was suppressed in overexpression lines under freezing conditions. However, there was no significant change in the expression of genes involved in the C-repeat binding factor (CBF)-mediated ABA-independent pathway between WT and CsbZIP18 overexpression plants. These results indicate that CsbZIP18 is a negative regulator of freezing tolerance via an ABA-dependent pathway.

MYB96 recruits the HDA15 protein to suppress negative regulators of ABA signaling in Arabidopsis

Unlike activation of target genes in response to abscisic acid (ABA), how MYB96 transcription factor represses ABA-repressible genes to further enhance ABA responses remains unknown. Here, we show MYB96 interacts with the histone modifier HDA15 to suppress negative regulators of early ABA signaling. The MYB96-HDA15 complex co-binds to the promoters of a subset of RHO GTPASE OF PLANTS (ROP) genes, ROP6, ROP10, and ROP11, and represses their expression by removing acetyl groups of histone H3 and H4 from the cognate regions, particularly in the presence of ABA. In support, HDA15-deficient mutants display reduced ABA sensitivity and are susceptible to drought stress with derepression of the ROP genes, as observed in the myb96-1 mutant. Biochemical and genetic analyses show that MYB96 and HDA15 are interdependent in the regulation of ROP suppression. Thus, MYB96 confers maximal ABA sensitivity by regulating both positive and negative regulators of ABA signaling through distinctive molecular mechanisms.

Salinity and ABA Seed Responses in Pepper: Expression and Interaction of ABA Core Signaling Components.

Abscisic acid (ABA) plays an important role in various aspects of plant growth and development, including adaptation to stresses, fruit development and ripening. In seeds, ABA participates through its core signaling components in dormancy instauration, longevity determination, and inhibition of germination in unfavorable environmental conditions such as high soil salinity. Here, we show that seed germination in pepper was delayed but only marginally reduced by ABA or NaCl with respect to control treatments. Through a similarity search, pepper orthologs of ABA core signaling components PYL (PYRABACTIN RESISTANCE1-LIKE), PP2C (PROTEIN PHOSPHATASE2C), and SnRK2 (SUCROSE NONFERMENTING1 (SNF1)-RELATED PROTEIN KINASE2) genes were identified. Gene expression analyses of selected members showed a low abundance of PYL and SnRK2 transcripts in dry seeds compared to other tissues, and an up-regulation at high concentrations of ABA and/or NaCl for both positive and negative regulators of ABA signaling. As expected, in hydroponically-grown seedlings exposed to NaCl, only PP2C encoding genes were up-regulated. Yeast two hybrid assays performed among putative pepper core components and with Arabidopsis thaliana orthologs confirmed the ability of the identified proteins to function in ABA signaling cascade, with the exception of a CaABI isoform cloned from seeds. BiFC assay in planta confirmed some of the interactions obtained in yeast. Altogether, our results indicate that a low expression of perception and signaling components in pepper seeds might contribute to explain the observed high percentages of seed germination in the presence of ABA. These results might have direct implications on the improvement of seed longevity and vigor, a bottleneck in pepper breeding.

Abscisic Acid Inhibits Rice Protein Phosphatase PP45 via H2O2 and Relieves Repression of the Ca2+/CaM-Dependent Protein Kinase DMI3.

In plants, Ca2+/calmodulin-dependent protein kinase (CCaMK) is a positive regulator of abscisic acid (ABA) responses, including root growth, antioxidant defense, and tolerance of both water stress and oxidative stress. However, the underlying molecular mechanisms are poorly understood. Here, we show a direct interaction between DMI3 (Doesn't Make Infections 3), a rice (Oryza sativa) CCaMK and PP45, a type 2C protein phosphatase in rice (PP2C). This interaction involves the CaM binding domain of DMI3 and the PP2C domain of PP45. In the absence of ABA, PP45 directly inactivates DMI3 by dephosphorylating Thr-263 in DMI3. However, in the presence of ABA, ABA-induced H2O2 production by the NADPH oxidases RbohB/E inhibits the activity of PP45 not only by inhibiting the expression of PP45 but also by oxidizing Cys-350 and Cys-428 residues to form PP45 intermolecular dimers. ABA-induced oxidation of Cys-350 and Cys-428 in PP45 blocked the interaction between PP45 and DMI3 and substantially prevented PP45-mediated inhibition in DMI3 activity. Genetic analysis indicated that PP45 is an important negative regulator of ABA signaling. These results reveal important pathways for the inhibition of DMI3 under the basal state and for its ABA-induced activation in rice.

Arabidopsis small ubiquitin-related modifier protease ASP1 positively regulates abscisic acid signaling during early seedling development.

The small ubiquitin-related modifier (SUMO) modification plays an important role in the regulation of abscisic acid (ABA) signaling, but the function of the SUMO protease, in ABA signaling, remains largely unknown. Here, we show that the SUMO protease, ASP1 positively regulates ABA signaling. Mutations in ASP1 resulted in an ABA-insensitive phenotype, during early seedling development. Wild-type ASP1 successfully rescued, whereas an ASP1 mutant (C577S), defective in SUMO protease activity, failed to rescue, the ABA-insensitive phenotype of asp1-1. Expression of ABI5 and MYB30 target genes was attenuated in asp1-1 and our genetic analyses revealed that ASP1 may function upstream of ABI5 and MYB30. Interestingly, ASP1 accumulated upon ABA treatment, and ABA-induced accumulation of ABI5 (a positive regulator of ABA signaling) was abolished, whereas ABA-induced accumulation of MYB30 (a negative regulator of ABA signaling) was increased in asp1-1. These findings support the hypothesis that increased levels of ASP1, upon ABA treatment, tilt the balance between ABI5 and MYB30 towards ABI5-mediated ABA signaling.

EAR1 Negatively Regulates ABA Signaling by Enhancing 2C Protein Phosphatase Activity.

The reversible phosphorylation of proteins by kinases and phosphatases is an antagonistic process that modulates many cellular functions. Protein phosphatases are usually negatively regulated by inhibitor proteins. During abscisic acid (ABA) signaling, these inhibitor proteins comprise PYR1/PYL/RCAR ABA receptors, which inhibit the core negative regulators, the clade A type 2C protein phosphatases (PP2Cs). However, it is not known whether these PP2Cs are positively regulated by other proteins. Here, we identified an Arabidopsis thaliana ear1 (enhancer of aba co-receptor1) mutant that exhibits pleiotropic ABA-hypersensitive phenotypes. EAR1 encodes an uncharacterized protein that is conserved in both monocots and dicots. EAR1 interacts with the N-terminal inhibition domains of all six PP2Cs, ABA INSENSITIVE1 (ABI1), ABI2, HYPERSENSITIVE TO ABA1 (HAB1), HAB2, ABA-HYPERSENSITIVE GERMINATION1 (AHG1), and AHG3, during ABA signaling and enhances the activity of PP2Cs both in vitro and in vivo. ABA treatment caused EAR1 to accumulate in the nucleus. These results indicate that EAR1 is a negative regulator of ABA signaling that enhances the activity of PP2Cs by interacting with and releasing the N-terminal autoinhibition of these proteins.

Chemical Control of ABA Receptors to Enable Plant Protection Against Water Stress.

This chapter describes procedures to find small molecules that bind to abscisic acid (ABA) receptors and improve plant tolerance to water stress. Abscisic acid promotes the interaction between ABA receptors and protein phosphatase 2Cs (PP2Cs), which are negative regulators of ABA signaling. This receptor-mediated ABA-dependent inhibition of PP2C activity is required for ABA action in vivo. ABA agonists can be identified by high-throughput screening for molecules that promote agonist-induced ABA receptor-PP2C interactions using yeast two-hybrid assays. In addition to in vitro assays, an in vivo test to evaluate ABA agonist activity has been devised in which an ABA-inducible GUS reporter strain is used to evaluate the effect of each ABA agonist. The effects of ABA agonists can also be determined using thermal imaging analyses and a water loss assay of stomata. Finally, the ability of each ABA agonist to confer drought stress tolerance upon plants grown in soil is determined. These methods can be used to identify various ABA agonists that protect plants against water stress.

The sHSP22 Heat Shock Protein Requires the ABI1 Protein Phosphatase to Modulate Polar Auxin Transport and Downstream Responses.

The phytohormones abscisic acid (ABA) and indole-3-acetic acid (IAA) response pathways interact synergistically or antagonistically to regulate plant development and environmental adaptation. Here, we show that ABI1, a key negative regulator of ABA signaling, is essential for auxin-modulated root development. We performed a microarray analysis using the loss-of-function mutant abi1-3 and Col-0 seedlings treated with IAA. For sHSP22, an endoplasmic reticulum (ER) small heat shock protein-encoding gene, the induction by IAA was dependent on ABI1shsp22 displayed enhanced sensitivity to ABA in primary root growth. In contrast, overexpression of full-length, but not truncated sHSP22 lacking signal peptide and ER-retention sequences, resulted in decreased ABA sensitivity. Overexpressed (OX) sHSP22 partially rescued the ABA hypersensitivity of abi1-3 In addition, sHSP22 is involved in auxin-regulated hypocotyl elongation at high temperature treatment. sHSP22 also affected accumulation of auxin efflux carrier PIN proteins due to potentiated intracellular trafficking. And sHSP22 OX lines initiated more lateral roots after auxin application. Our results suggest that sHSP22 regulates auxin response through modulating auxin polar transport, and ABI1-sHSP22 provides a novel module orchestrating ABA and auxin signaling crosstalk in Arabidopsis (Arabidopsis thaliana).

Trithorax-group proteins ARABIDOPSIS TRITHORAX4 (ATX4) and ATX5 function in abscisic acid and dehydration stress responses.

Trithorax-group proteins (TrxGs) play essential regulatory roles in chromatin modification to activate transcription. Although TrxGs have been shown to be extensively involved in the activation of developmental genes, how the specific TrxGs function in the dehydration and abscisic acid (ABA)-mediated modulation of downstream gene expression remains unknown. Here, we report that two evolutionarily conserved Arabidopsis thaliana TrxGs, ARABIDOPSIS TRITHORAX4 (ATX4) and ATX5, play essential roles in the drought stress response. atx4 and atx5 single loss-of-function mutants showed drought stress-tolerant and ABA-hypersensitive phenotypes during seed germination and seedling development, while the atx4 atx5 double mutant displayed further exacerbation of the phenotypes. Genome-wide RNA-sequencing analyses showed that ATX4 and ATX5 regulate the expression of genes functioning in dehydration stress. Intriguingly, ABA-HYPERSENSITIVE GERMINATION 3 (AHG3), an essential negative regulator of ABA signaling, acts genetically downstream of ATX4 and ATX5 in response to ABA. ATX4 and ATX5 directly bind to the AHG3 locus and trimethylate histone H3 of Lys 4 (H3K4). Moreover, ATX4 and ATX5 occupancies at AHG3 are dramatically increased under ABA treatment, and are also essential for RNA polymerase II (RNAPII) occupancies. Our findings reveal novel molecular functions of A. thaliana TrxGs in dehydration stress and ABA responses.

Members of the abscisic acid co-receptor PP2C protein family mediate salicylic acid-abscisic acid crosstalk.

The interplay between abscisic acid (ABA) and salicylic acid (SA) influences plant responses to various (a)biotic stresses; however, the underlying mechanism for this crosstalk is largely unknown. Here, we report that type 2C protein phosphatases (PP2Cs), some of which are negative regulators of ABA signaling, bind SA. SA binding suppressed the ABA‐enhanced interaction between these PP2Cs and various ABA receptors belonging to the PYR/PYL/RCAR protein family. Additionally, SA suppressed ABA‐enhanced degradation of PP2Cs and ABA‐induced stabilization of SnRK2s. Supporting SA's role as a negative regulator of ABA signaling, exogenous SA suppressed ABA‐induced gene expression, whereas the SA‐deficient sid2‐1 mutant displayed heightened PP2C degradation and hypersensitivity to ABA‐induced suppression of seed germination. Together, these results suggest a new molecular mechanism through which SA antagonizes ABA signaling. A better understanding of the crosstalk between these hormones is important for improving the sustainability of agriculture in the face of climate change.

AIK1, A Mitogen-Activated Protein Kinase, Modulates Abscisic Acid Responses through the MKK5-MPK6 Kinase Cascade.

The mitogen-activated protein kinase (MAPK) cascade is an evolutionarily conserved signal transduction module involved in transducing extracellular signals to the nucleus for appropriate cellular adjustment. This cascade essentially consists of three components: a MAPK kinase kinase (MAPKKK), a MAPK kinase, and a MAPK, connected to each other by the event of phosphorylation. Here, we report the characterization of a MAPKKK, ABA-INSENSITIVE PROTEIN KINASE1 (AIK1), which regulates abscisic acid (ABA) responses in Arabidopsis (Arabidopsis thaliana). T-DNA insertion mutants of AIK1 showed insensitivity to ABA in terms of both root growth and stomatal response. AIK1 functions in ABA responses via regulation of root cell division and elongation, as well as stomatal responses. The activity of AIK1 is induced by ABA in Arabidopsis and tobacco (Nicotiana benthamiana), and the Arabidopsis protein phosphatase type 2C, ABI1, a negative regulator of ABA signaling, restricts AIK1 activity by dephosphorylation. Bimolecular fluorescence complementation analysis showed that MPK3, MPK6, and AIK1 interact with MKK5. The single mutant seedlings of mpk6 and mkk5 have similar phenotypes to aik1, but mkk4 does not. AIK1 was localized in the cytoplasm and shown to activate MKK5 by protein phosphorylation, which was an ABA-activated process. Constitutively active MKK5 in aik1 mutant seedlings complements the ABA-insensitive root growth phenotype of aik1 The activity of MPK6 was increased by ABA in wild-type seedlings, but its activation by ABA was impaired in aik1 and aik1 mkk5 mutants. These findings clearly suggest that the AIK1-MKK5-MPK6 cascade functions in the ABA regulation of primary root growth and stomatal response.

FERONIA interacts with ABI2-type phosphatases to facilitate signaling cross-talk between abscisic acid and RALF peptide in Arabidopsis

Receptor-like kinase FERONIA (FER) plays a crucial role in plant response to small molecule hormones [e.g., auxin and abscisic acid (ABA)] and peptide signals [e.g., rapid alkalinization factor (RALF)]. It remains unknown how FER integrates these different signaling events in the control of cell growth and stress responses. Under stress conditions, increased levels of ABA will inhibit cell elongation in the roots. In our previous work, we have shown that FER, through activation of the guanine nucleotide exchange factor 1 (GEF1)/4/10-Rho of Plant 11 (ROP11) pathway, enhances the activity of the phosphatase ABA Insensitive 2 (ABI2), a negative regulator of ABA signaling, thereby inhibiting ABA response. In this study, we found that both RALF and ABA activated FER by increasing the phosphorylation level of FER. The FER loss-of-function mutant displayed strong hypersensitivity to both ABA and abiotic stresses such as salt and cold conditions, indicating that FER plays a key role in ABA and stress responses. We further showed that ABI2 directly interacted with and dephosphorylated FER, leading to inhibition of FER activity. Several other ABI2-like phosphatases also function in this pathway, and ABA-dependent FER activation required PYRABACTIN RESISTANCE (PYR)/PYR1-LIKE (PYL)/REGULATORY COMPONENTS OF ABA RECEPTORS (RCAR)-A-type protein phosphatase type 2C (PP2CA) modules. Furthermore, suppression of RALF1 gene expression, similar to disruption of the FER gene, rendered plants hypersensitive to ABA. These results formulated a mechanism for ABA activation of FER and for cross-talk between ABA and peptide hormone RALF in the control of plant growth and responses to stress signals.

Negative regulation of ABA signaling by WRKY33 is critical for Arabidopsis immunity towards Botrytis cinerea 2100.

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