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      The karrikin receptor KAI2 promotes drought resistance in Arabidopsis thaliana

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          Abstract

          Drought causes substantial reductions in crop yields worldwide. Therefore, we set out to identify new chemical and genetic factors that regulate drought resistance in Arabidopsis thaliana. Karrikins (KARs) are a class of butenolide compounds found in smoke that promote seed germination, and have been reported to improve seedling vigor under stressful growth conditions. Here, we discovered that mutations in KARRIKIN INSENSITIVE2 ( KAI2), encoding the proposed karrikin receptor, result in hypersensitivity to water deprivation. We performed transcriptomic, physiological and biochemical analyses of kai2 plants to understand the basis for KAI2-regulated drought resistance. We found that kai2 mutants have increased rates of water loss and drought-induced cell membrane damage, enlarged stomatal apertures, and higher cuticular permeability. In addition, kai2 plants have reduced anthocyanin biosynthesis during drought, and are hyposensitive to abscisic acid (ABA) in stomatal closure and cotyledon opening assays. We identified genes that are likely associated with the observed physiological and biochemical changes through a genome-wide transcriptome analysis of kai2 under both well-watered and dehydration conditions. These data provide evidence for crosstalk between ABA- and KAI2-dependent signaling pathways in regulating plant responses to drought. A comparison of the strigolactone receptor mutant d14 ( DWARF14) to kai2 indicated that strigolactones also contributes to plant drought adaptation, although not by affecting cuticle development. Our findings suggest that chemical or genetic manipulation of KAI2 and D14 signaling may provide novel ways to improve drought resistance.

          Author summary

          MORE AXILLARY GROWTH2 (MAX2) is a central regulator of both strigolactone and karrikin (KAR) signaling pathways, which control many aspects of plant growth and development. MAX2 promotes plant resistance to drought. Application of a strigolactone analog enhances plant adaptation to drought, implying that strigolactone signaling regulates this trait; however, the potential contributions of the KAR signaling pathway to plant drought responses have not been explored. In this study, the functions of the KAR receptor KARRIKIN INSENSITIVE2 (KAI2) were analyzed to elucidate the role of KAI2-mediated signaling in plant drought adaptation. Our results indicate that the KAI2 signaling positively regulates drought resistance of Arabidopsis plants by both drought avoidance and drought tolerance strategies, such as through enhancing cuticle formation, stomatal closure, cell membrane integrity and anthocyanin biosynthesis. Our findings provide potential for manipulation of KAR- and/or strigolactone-mediated signaling pathways for improvement of drought resistance.

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          The transcriptional regulatory network in the drought response and its crosstalk in abiotic stress responses including drought, cold, and heat

          Drought negatively impacts plant growth and the productivity of crops around the world. Understanding the molecular mechanisms in the drought response is important for improvement of drought tolerance using molecular techniques. In plants, abscisic acid (ABA) is accumulated under osmotic stress conditions caused by drought, and has a key role in stress responses and tolerance. Comprehensive molecular analyses have shown that ABA regulates the expression of many genes under osmotic stress conditions, and the ABA-responsive element (ABRE) is the major cis-element for ABA-responsive gene expression. Transcription factors (TFs) are master regulators of gene expression. ABRE-binding protein and ABRE-binding factor TFs control gene expression in an ABA-dependent manner. SNF1-related protein kinases 2, group A 2C-type protein phosphatases, and ABA receptors were shown to control the ABA signaling pathway. ABA-independent signaling pathways such as dehydration-responsive element-binding protein TFs and NAC TFs are also involved in stress responses including drought, heat, and cold. Recent studies have suggested that there are interactions between the major ABA signaling pathway and other signaling factors in stress responses. The important roles of these TFs in crosstalk among abiotic stress responses will be discussed. Control of ABA or stress signaling factor expression can improve tolerance to environmental stresses. Recent studies using crops have shown that stress-specific overexpression of TFs improves drought tolerance and grain yield compared with controls in the field.
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            Analysis of cytokinin mutants and regulation of cytokinin metabolic genes reveals important regulatory roles of cytokinins in drought, salt and abscisic acid responses, and abscisic acid biosynthesis.

            Cytokinins (CKs) regulate plant growth and development via a complex network of CK signaling. Here, we perform functional analyses with CK-deficient plants to provide direct evidence that CKs negatively regulate salt and drought stress signaling. All CK-deficient plants with reduced levels of various CKs exhibited a strong stress-tolerant phenotype that was associated with increased cell membrane integrity and abscisic acid (ABA) hypersensitivity rather than stomatal density and ABA-mediated stomatal closure. Expression of the Arabidopsis thaliana ISOPENTENYL-TRANSFERASE genes involved in the biosynthesis of bioactive CKs and the majority of the Arabidopsis CYTOKININ OXIDASES/DEHYDROGENASES genes was repressed by stress and ABA treatments, leading to a decrease in biologically active CK contents. These results demonstrate a novel mechanism for survival under abiotic stress conditions via the homeostatic regulation of steady state CK levels. Additionally, under normal conditions, although CK deficiency increased the sensitivity of plants to exogenous ABA, it caused a downregulation of key ABA biosynthetic genes, leading to a significant reduction in endogenous ABA levels in CK-deficient plants relative to the wild type. Taken together, this study provides direct evidence that mutual regulation mechanisms exist between the CK and ABA metabolism and signals underlying different processes regulating plant adaptation to stressors as well as plant growth and development.
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              The SHINE clade of AP2 domain transcription factors activates wax biosynthesis, alters cuticle properties, and confers drought tolerance when overexpressed in Arabidopsis.

              The interface between plants and the environment plays a dual role as a protective barrier as well as a medium for the exchange of gases, water, and nutrients. The primary aerial plant surfaces are covered by a cuticle, acting as the essential permeability barrier toward the atmosphere. It is a heterogeneous layer composed mainly of lipids, namely cutin and intracuticular wax with epicuticular waxes deposited on the surface. We identified an Arabidopsis thaliana activation tag gain-of-function mutant shine (shn) that displayed a brilliant, shiny green leaf surface with increased cuticular wax compared with the leaves of wild-type plants. The gene responsible for the phenotype encodes one member of a clade of three proteins of undisclosed function, belonging to the plant-specific family of AP2/EREBP transcription factors. Overexpression of all three SHN clade genes conferred a phenotype similar to that of the original shn mutant. Biochemically, such plants were altered in wax composition (very long fatty acid derivatives). Total cuticular wax levels were increased sixfold in shn compared with the wild type, mainly because of a ninefold increase in alkanes that comprised approximately half of the total waxes in the mutant. Chlorophyll leaching assays and fresh weight loss experiments indicated that overexpression of the SHN genes increased cuticle permeability, probably because of changes in its ultrastructure. Likewise, SHN gene overexpression altered leaf and petal epidermal cell structure, trichome number, and branching as well as the stomatal index. Interestingly, SHN overexpressors displayed significant drought tolerance and recovery, probably related to the reduced stomatal density. Expression analysis using promoter-beta-glucuronidase fusions of the SHN genes provides evidence for the role of the SHN clade in plant protective layers, such as those formed during abscission, dehiscence, wounding, tissue strengthening, and the cuticle. We propose that these diverse functions are mediated by regulating metabolism of lipid and/or cell wall components.
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                Author and article information

                Contributors
                Role: Data curationRole: Formal analysisRole: InvestigationRole: Writing – original draftRole: Writing – review & editing
                Role: Data curationRole: Formal analysisRole: Investigation
                Role: Data curationRole: Formal analysisRole: Investigation
                Role: Data curationRole: Formal analysisRole: Investigation
                Role: Data curationRole: Investigation
                Role: Data curationRole: Formal analysisRole: Investigation
                Role: Formal analysis
                Role: Data curationRole: Investigation
                Role: Data curationRole: InvestigationRole: Resources
                Role: Investigation
                Role: Data curationRole: Investigation
                Role: Formal analysis
                Role: Funding acquisitionRole: Resources
                Role: Funding acquisitionRole: Resources
                Role: Resources
                Role: Funding acquisitionRole: Writing – review & editing
                Role: Writing – review & editing
                Role: Writing – review & editing
                Role: ConceptualizationRole: Funding acquisitionRole: Project administrationRole: Writing – original draftRole: Writing – review & editing
                Role: Editor
                Journal
                PLoS Genet
                PLoS Genet
                plos
                plosgen
                PLoS Genetics
                Public Library of Science (San Francisco, CA USA )
                1553-7390
                1553-7404
                13 November 2017
                November 2017
                : 13
                : 11
                : e1007076
                Affiliations
                [1 ] Signaling Pathway Research Unit, RIKEN Center for Sustainable Resource Science, Yokohama, Japan
                [2 ] Faculty of Bioscience and Bioindustry, Tokushima University, Tokushima, Japan
                [3 ] Deutsche Forschungsgemeinschaft Center for Regenerative Therapies, Technische Universität Dresden, Fetscherstraße 105, Germany
                [4 ] Mass Spectrometry and Microscopy Unit, RIKEN Center for Sustainable Resource Science, Yokohama, Japan
                [5 ] Department of Genetics, University of Georgia, Athens, Georgia, United States of America
                [6 ] Plant Genomic Network Research Team, RIKEN Center for Sustainable Resource Science, Yokohama, Japan
                [7 ] Dormancy and Adaptation Research Unit, RIKEN Center for Sustainable Resource Science, Yokohama, Japan
                [8 ] Department of Biomolecular Sciences, Graduate School of Life Sciences, Tohoku University, Sendai, Miyagi, Japan
                [9 ] Department of Botany & Plant Sciences, University of California, Riverside, Riverside, California, United States of America
                [10 ] Key Laboratory of Mollisols Agroecology, Northeast Institute of Geography and Agroecology, Chinese Academy of Sciences, Changchun, People's Republic of China
                [11 ] Laboratorio Nacional de Genómica para la Biodiversidad (Langebio)/Unidad de Genómica Avanzada, Centro de Investigación y Estudios Avanzados del Instituto Politécnico Nacional, Irapuato, Guanajuato, Mexico
                University of California, San Diego, UNITED STATES
                Author notes

                The authors have declared that no competing interests exist.

                Author information
                http://orcid.org/0000-0003-0087-9994
                http://orcid.org/0000-0002-6414-5191
                http://orcid.org/0000-0003-2736-5424
                http://orcid.org/0000-0003-1792-6005
                http://orcid.org/0000-0001-9883-9768
                Article
                PGENETICS-D-17-00328
                10.1371/journal.pgen.1007076
                5703579
                29131815
                ad247de9-de94-45c0-a25d-9e4744b48be1
                © 2017 Li et al

                This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

                History
                : 15 February 2017
                : 15 October 2017
                Page count
                Figures: 8, Tables: 0, Pages: 23
                Funding
                Funded by: funder-id http://dx.doi.org/10.13039/501100003382, Core Research for Evolutional Science and Technology;
                Award Recipient :
                Funded by: funder-id http://dx.doi.org/10.13039/100000001, National Science Foundation;
                Award ID: IOS-1350561
                Award Recipient :
                Funded by: Japan Advanced Plant Science Research Network
                Award Recipient :
                Funded by: funder-id http://dx.doi.org/10.13039/501100001691, Japan Society for the Promotion of Science;
                Award ID: #17K07459
                Award Recipient :
                Funded by: funder-id http://dx.doi.org/10.13039/501100001691, Japan Society for the Promotion of Science;
                Award ID: None
                Award Recipient :
                This project was supported in part by the Japan Society for the Promotion of Science (#17K07459 to LSPT), the Japan Science and Technology Agency (JST), Core Research for Evolutionary Science and Technology (CREST to MoS), and the National Science Foundation (IOS-1737153 to DCN). The ABA measurements were supported by the Japan Advanced Plant Science Research Network. MGM acknowledges the postdoc fellowship from the Japan Society for the Promotion of Science. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
                Categories
                Research Article
                Medicine and Health Sciences
                Diagnostic Medicine
                Signs and Symptoms
                Dehydration (Medicine)
                Medicine and Health Sciences
                Pathology and Laboratory Medicine
                Signs and Symptoms
                Dehydration (Medicine)
                Biology and Life Sciences
                Plant Science
                Plant Anatomy
                Leaves
                Ecology and Environmental Sciences
                Natural Resources
                Water Resources
                Research and Analysis Methods
                Experimental Organism Systems
                Model Organisms
                Arabidopsis Thaliana
                Research and Analysis Methods
                Model Organisms
                Arabidopsis Thaliana
                Biology and Life Sciences
                Organisms
                Eukaryota
                Plants
                Brassica
                Arabidopsis Thaliana
                Research and Analysis Methods
                Experimental Organism Systems
                Plant and Algal Models
                Arabidopsis Thaliana
                Biology and Life Sciences
                Plant Science
                Plant Physiology
                Plant Defenses
                Plant Resistance to Abiotic Stress
                Drought Adaptation
                Biology and Life Sciences
                Plant Science
                Plant Pathology
                Plant Resistance to Abiotic Stress
                Drought Adaptation
                Biology and Life Sciences
                Ecology
                Plant Ecology
                Plant-Environment Interactions
                Plant Resistance to Abiotic Stress
                Drought Adaptation
                Ecology and Environmental Sciences
                Ecology
                Plant Ecology
                Plant-Environment Interactions
                Plant Resistance to Abiotic Stress
                Drought Adaptation
                Biology and Life Sciences
                Plant Science
                Plant Ecology
                Plant-Environment Interactions
                Plant Resistance to Abiotic Stress
                Drought Adaptation
                Biology and Life Sciences
                Plant Science
                Plant Physiology
                Plant Defenses
                Plant Resistance to Abiotic Stress
                Biology and Life Sciences
                Plant Science
                Plant Pathology
                Plant Resistance to Abiotic Stress
                Biology and Life Sciences
                Ecology
                Plant Ecology
                Plant-Environment Interactions
                Plant Resistance to Abiotic Stress
                Ecology and Environmental Sciences
                Ecology
                Plant Ecology
                Plant-Environment Interactions
                Plant Resistance to Abiotic Stress
                Biology and Life Sciences
                Plant Science
                Plant Ecology
                Plant-Environment Interactions
                Plant Resistance to Abiotic Stress
                Biology and Life Sciences
                Genetics
                Gene Expression
                Biology and Life Sciences
                Plant Science
                Plant Physiology
                Custom metadata
                vor-update-to-uncorrected-proof
                2017-11-27
                The raw microarray data are available from the Gene Expression Omnibus database, accession number GSE90622 ( https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE90622). All other data are within the paper and its Supporting Information files.

                Genetics
                Genetics

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