The phytohormone abscisic acid (ABA) modulates the expression of many genes important to plant growth and development and to stress adaptation. In this study, we found that an APETALA2/EREBP-type transcription factor, AtERF7, plays an important role in ABA responses. AtERF7 interacts with the protein kinase PKS3, which has been shown to be a global regulator of ABA responses. AtERF7 binds to the GCC box and acts as a repressor of gene transcription. AtERF7 interacts with the Arabidopsis thaliana homolog of a human global corepressor of transcription, AtSin3, which in turn may interact with HDA19, a histone deacetylase. The transcriptional repression activity of AtERF7 is enhanced by HDA19 and AtSin3. Arabidopsis plants overexpressing AtERF7 show reduced sensitivity of guard cells to ABA and increased transpirational water loss. By contrast, AtERF7 and AtSin3 RNA interference lines show increased sensitivity to ABA during germination. Together, our results suggest that AtERF7 plays an important role in ABA responses and may be part of a transcriptional repressor complex and be regulated by PKS3.

Drought stress is an important environmental factor limiting plant productivity. In this study, we screened drought-resistant transgenic plants from 65 promoter-pyrabactin resistance 1-like (PYL) abscisic acid (ABA) receptor gene combinations and discovered that pRD29A::PYL9 transgenic lines showed dramatically increased drought resistance and drought-induced leaf senescence in both Arabidopsis and rice. Previous studies suggested that ABA promotes senescence by causing ethylene production. However, we found that ABA promotes leaf senescence in an ethylene-independent manner by activating sucrose nonfermenting 1-related protein kinase 2s (SnRK2s), which subsequently phosphorylate ABA-responsive element-binding factors (ABFs) and Related to ABA-Insensitive 3/VP1 (RAV1) transcription factors. The phosphorylated ABFs and RAV1 up-regulate the expression of senescence-associated genes, partly by up-regulating the expression of Oresara 1. The pyl9 and ABA-insensitive 1-1 single mutants, pyl8-1pyl9 double mutant, and snrk2.2/3/6 triple mutant showed reduced ABA-induced leaf senescence relative to the WT, whereas pRD29A::PYL9 transgenic plants showed enhanced ABA-induced leaf senescence. We found that leaf senescence may benefit drought resistance by helping to generate an osmotic potential gradient, which is increased in pRD29A::PYL9 transgenic plants and causes water to preferentially flow to developing tissues. Our results uncover the molecular mechanism of ABA-induced leaf senescence and suggest an important role of PYL9 and leaf senescence in promoting resistance to extreme drought stress.drought stress | abscisic acid | PYL | dormancy | Arabidopsis C ell and organ senescence causes programmed cell death to regulate the growth and development of organisms. In plants, leaf senescence increases the transfer of nutrients to developing and storage tissues. Recently, studies on transgenic tobacco showed that delayed leaf senescence increases plant resistance to drought stress (1). However, the senescence and abscission of older leaves and subsequent transfer of nutrients are known to increase plant survival under abiotic stresses, including drought, low or high temperatures, and darkness (2, 3). Senescence mainly develops in an age-dependent manner and is also triggered by environmental stresses and phytohormones, such as abscisic acid (ABA), ethylene, salicylic acid, and jasmonic acid, but delayed by cytokinin (4).Senescence-associated genes (SAGs) are induced by leaf senescence. The expression of SAGs is tightly controlled by several senescence-promoting, plant-specific NAC (NAM, ATAF1, and CUC2) transcription factors, such as Oresara 1 (ORE1) (5), Oresara 1 sister 1 (ORS1) (6), and AtNAP (7). Environmental stimuli and phytohormones may regulate leaf senescence through NACs. Phytochrome-interacting factor 4 (PIF4) and PIF5 transcription factors promote dark-induced senescence by activating ORE1 expression (8). The expression of ORE1, AtNAP, and OsNAP (ortholog of AtNAP) is up-regulated by ABA by an unknown molecular m...

Sucrose nonfermenting 1 (SNF1)-related protein kinase 2s (SnRK2s) are central components of abscisic acid (ABA) signaling pathways. The snrk2.2/2.3/2.6 triple-mutant plants are nearly completely insensitive to ABA, suggesting that most of the molecular actions of ABA are triggered by the SnRK2s-mediated phosphorylation of substrate proteins. Only a few substrate proteins of the SnRK2s are known. To identify additional substrate proteins of the SnRK2s and provide insight into the molecular actions of ABA, we used quantitative phosphoproteomics to compare the global changes in phosphopeptides in WT and snrk2.2/2.3/2.6 triple mutant seedlings in response to ABA treatment. Among the 5,386 unique phosphorylated peptides identified in this study, we found that ABA can increase the phosphorylation of 166 peptides and decrease the phosphorylation of 117 peptides in WT seedlings. In the snrk2.2/ 2.3/2.6 triple mutant, 84 of the 166 peptides, representing 58 proteins, could not be phosphorylated, or phosphorylation was not increased under ABA treatment. In vitro kinase assays suggest that most of the 58 proteins can serve as substrates of the SnRK2s. The SnRK2 substrates include proteins involved in flowering time regulation, RNA and DNA binding, miRNA and epigenetic regulation, signal transduction, chloroplast function, and many other cellular processes. Consistent with the SnRK2 phosphorylation of flowering time regulators, the snrk2.2/2.3/2.6 triple mutant flowered significantly earlier than WT. These results shed new light on the role of the SnRK2 protein kinases and on the downstream effectors of ABA action, and improve our understanding of plant responses to adverse environments. T he phytohormone abscisic acid (ABA) plays important roles in plant development and responses to stressful environments (1, 2). Recently, the discovery of the PYR1 (Pyrabactin Resistance 1)/ PYL (PYR1-Like)/RCAR (Regulatory Component of ABA Receptor) family of ABA receptors led to the elucidation of the core ABA signaling pathway. ABA binds to the PYLs, triggering the PYLs to interact with and inactivate clade A protein phosphatase 2Cs (PP2Cs). This releases Sucrose nonfermenting 1 (SNF1)-related protein kinase 2s (SnRK2s) from inhibition by the PP2Cs, allowing the kinases to phosphorylate downstream effectors of ABA responses (3-5).SnRK2s are a plant-specific protein kinase family related to the yeast SNF1 and animal AMP-dependent protein kinase (AMPK) (6), and the family has 10 members (SnRK2.1-2.10) in Arabidopsis. ABA treatment can quickly activate SnRK2.2, 2.3 and 2.6 (7), and the snrk2.2/2.3/2.6 triple-knockout mutant has a very strong ABAinsensitive phenotype and shows little response to even very high concentrations of ABA in seed germination, root growth, and stomatal movement (8). In contrast, mutations in the other seven SnRK2 family members do not cause significant ABA insensitivity (9). Notwithstanding the key role of SnRK2.2/2.3/2.6 in ABA signaling, some ABA responses are possibly independent of the SnRK2s, because the PYL r...

The phytohormone abscisic acid (ABA) plays important roles in plant development and adaptation to environmental stress. ABA induces the production of nitric oxide (NO) in guard cells, but how NO regulates ABA signaling is not understood. Here, we show that NO negatively regulates ABA signaling in guard cells by inhibiting open stomata 1 (OST1)/sucrose nonfermenting 1 (SNF1)-related protein kinase 2.6 (SnRK2.6) through S-nitrosylation. We found that SnRK2.6 is S-nitrosylated at cysteine 137, a residue adjacent to the kinase catalytic site. Dysfunction in the S-nitrosoglutathione (GSNO) reductase (GSNOR) gene in the gsnor1-3 mutant causes NO overaccumulation in guard cells, constitutive S-nitrosylation of SnRK2.6, and impairment of ABA-induced stomatal closure. Introduction of the Cys137 to Ser mutated SnRK2.6 into the gsnor1-3/ ost1-3 double-mutant partially suppressed the effect of gsnor1-3 on ABA-induced stomatal closure. A cysteine residue corresponding to Cys137 of SnRK2.6 is present in several yeast and human protein kinases and can be S-nitrosylated, suggesting that the S-nitrosylation may be an evolutionarily conserved mechanism for protein kinase regulation.A bscisic acid (ABA) plays critical roles in seed dormancy and germination, plant growth, and adaptation to environmental challenges (1, 2). Stresses, such as drought and high salt conditions, increase ABA concentration in plants as a result of ABA biosynthesis or ABA release from its inactive, conjugated forms (3). In the presence of ABA, the ABA receptors in the PYR1 (Pyrabactin Resistance 1)/PYL (PYR1-Like)/RCAR (Regulatory Component of ABA receptor) protein family bind to and inhibit the activity of clade A protein phosphatase 2Cs (PP2Cs), which are considered as coreceptors and negative regulators of ABA signaling (4-6). This process then results in the release of sucrose nonfermenting 1 (SNF1)-related protein kinase 2s (SnRK2s) from suppression by the PP2Cs. As central components of the ABA signaling pathway, the activated SnRK2s phosphorylate dozens of downstream effectors to regulate various physiological processes, including stomatal closure, root growth and development, seed dormancy, seed germination, and flowering (7).As the gateway for photosynthetic CO 2 uptake and transpirational water loss, stomata are critical for plant growth and physiology (8). ABA regulates stomatal movement and mutations in ABA biosynthesis genes (9), or in the PYL or SnRK2.6 (also known as OST1) genes cause open-stomata phenotypes (10). On the other hand, dysfunction of the PP2Cs or overexpression of RCAR1/PYL9 causes stomatal closure (5). Among the three SnRK2s, SnRK2.2, -2.3, and -2.6, which are most important for ABA signaling, SnRK2.6 is preferentially expressed in guard cells and plays a critical role in stomatal regulation, whereas SnRK2.2 and -2.3 are mainly expressed in seeds and young seedlings and are thus more important for seed germination and seedling growth (4, 11). SnRK2.6 phosphorylates the slow (S-type) anion channel associated 1 and inward potass...

Nitric oxide (NO) is a bioactive molecule that functions in numerous physiological and developmental processes in plants, including lateral root development. In this study, we used biochemical and genetic approaches to analyze the function of Arabidopsis thaliana mitogen-activated protein kinase 6 (MPK6) in the regulation of NO synthesis in response to hydrogen peroxide (H 2 O 2 ) during lateral root development. In both mpk6 mutants studied, H 2 O 2 -induced NO synthesis and nitrate reductase (NR) activity were decreased dramatically. Furthermore, one NR isoform, NIA2, was required for the MPK6-mediated production of NO induced by H 2 O 2 . Notably, NIA2 interacted physically with MPK6 in vitro and in vivo and also served as a substrate of MPK6. Phosphorylation of NIA2 by MPK6 led to an increase in NR activity, and Ser-627 was identified as the putative phosphorylation site on NIA2. Phenotypical analysis revealed that mpk6-2 and mpk6-3 seedlings produce more and longer lateral roots than wild-type plants did after application of the NO donor sodium nitroprusside or H 2 O 2 . These data support strongly a function of MPK6 in modulating NO production and signal transduction in response to H 2 O 2 during Arabidopsis root development.