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      Transcriptional regulation of abscisic acid biosynthesis and signal transduction, and anthocyanin biosynthesis in ‘Bluecrop’ highbush blueberry fruit during ripening

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          Abstract

          Highbush blueberry ( Vaccinium corymbosum) fruit accumulate high levels of anthocyanins during ripening, which might be controlled by abscisic acid (ABA), a signal molecule in non-climacteric fruits. For an integrated view of the ripening process from ABA to anthocyanin biosynthesis, we analyzed the transcriptomes of ‘Bluecrop’ highbush blueberry fruit using RNA-Seq at three ripening stages, categorized based on fruit skin coloration: pale green at ca. 30 days after full bloom (DAFB), reddish purple at ca. 40 DAFB, and dark purple at ca. 50 DAFB. Mapping the trimmed reads against the reference sequences yielded 25,766 transcripts. Of these, 143 transcripts were annotated to encode five ABA biosynthesis enzymes, four ABA signal transduction regulators, four ABA-responsive transcription factors, and 12 anthocyanin biosynthesis enzymes. The analysis of differentially expressed genes between the ripening stages revealed that 11 transcripts, including those encoding nine- cis-epoxycarotenoid dioxygenase, SQUAMOSA-class MADS box transcription factor, and flavonoid 3′,5′-hydroxylase, were significantly up-regulated throughout the entire ripening stages. In fruit treated with 1 g L −1 ABA, at least nine transcripts of these 11 transcripts as well as one transcript encoding flavonoid 3′-hydroxylase were up-regulated, presumably promoting anthocyanin accumulation and fruit skin coloration. These results will provide fundamental information demonstrating that ABA biosynthesis and signal transduction, and anthocyanin biosynthesis are closely associated with anthocyanin accumulation and skin coloration in highbush blueberry fruit during ripening.

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          Abscisic acid inhibits type 2C protein phosphatases via the PYR/PYL family of START proteins.

          Type 2C protein phosphatases (PP2Cs) are vitally involved in abscisic acid (ABA) signaling. Here, we show that a synthetic growth inhibitor called pyrabactin functions as a selective ABA agonist. Pyrabactin acts through PYRABACTIN RESISTANCE 1 (PYR1), the founding member of a family of START proteins called PYR/PYLs, which are necessary for both pyrabactin and ABA signaling in vivo. We show that ABA binds to PYR1, which in turn binds to and inhibits PP2Cs. We conclude that PYR/PYLs are ABA receptors functioning at the apex of a negative regulatory pathway that controls ABA signaling by inhibiting PP2Cs. Our results illustrate the power of the chemical genetic approach for sidestepping genetic redundancy.
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            Abscisic acid biosynthesis and catabolism.

            The level of abscisic acid (ABA) in any particular tissue in a plant is determined by the rate of biosynthesis and catabolism of the hormone. Therefore, identifying all the genes involved in the metabolism is essential for a complete understanding of how this hormone directs plant growth and development. To date, almost all the biosynthetic genes have been identified through the isolation of auxotrophic mutants. On the other hand, among several ABA catabolic pathways, current genomic approaches revealed that Arabidopsis CYP707A genes encode ABA 8'-hydroxylases, which catalyze the first committed step in the predominant ABA catabolic pathway. Identification of ABA metabolic genes has revealed that multiple metabolic steps are differentially regulated to fine-tune the ABA level at both transcriptional and post-transcriptional levels. Furthermore, recent ongoing studies have given new insights into the regulation and site of ABA metabolism in relation to its physiological roles.
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              Abscisic acid plays an important role in the regulation of strawberry fruit ripening.

              The plant hormone abscisic acid (ABA) has been suggested to play a role in fruit development, but supporting genetic evidence has been lacking. Here, we report that ABA promotes strawberry (Fragaria ananassa) fruit ripening. Using a newly established Tobacco rattle virus-induced gene silencing technique in strawberry fruit, the expression of a 9-cis-epoxycarotenoid dioxygenase gene (FaNCED1), which is key to ABA biosynthesis, was down-regulated, resulting in a significant decrease in ABA levels and uncolored fruits. Interestingly, a similar uncolored phenotype was observed in the transgenic RNA interference (RNAi) fruits, in which the expression of a putative ABA receptor gene encoding the magnesium chelatase H subunit (FaCHLH/ABAR) was down-regulated by virus-induced gene silencing. More importantly, the uncolored phenotype of the FaNCED1-down-regulated RNAi fruits could be rescued by exogenous ABA, but the ABA treatment could not reverse the uncolored phenotype of the FaCHLH/ABAR-down-regulated RNAi fruits. We observed that down-regulation of the FaCHLH/ABAR gene in the RNAi fruit altered both ABA levels and sugar content as well as a set of ABA- and/or sugar-responsive genes. Additionally, we showed that exogenous sugars, particularly sucrose, can significantly promote ripening while stimulating ABA accumulation. These data provide evidence that ABA is a signal molecule that promotes strawberry ripening and that the putative ABA receptor, FaCHLH/ABAR, is a positive regulator of ripening in response to ABA.
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                Author and article information

                Contributors
                Role: ConceptualizationRole: Data curationRole: Formal analysisRole: MethodologyRole: ValidationRole: VisualizationRole: Writing – original draftRole: Writing – review & editing
                Role: ConceptualizationRole: Data curationRole: ValidationRole: Writing – review & editing
                Role: Methodology
                Role: Data curationRole: Methodology
                Role: Data curationRole: MethodologyRole: ResourcesRole: ValidationRole: Writing – review & editing
                Role: ConceptualizationRole: Data curationRole: Funding acquisitionRole: InvestigationRole: SupervisionRole: ValidationRole: Writing – review & editing
                Role: Editor
                Journal
                PLoS One
                PLoS ONE
                plos
                plosone
                PLoS ONE
                Public Library of Science (San Francisco, CA USA )
                1932-6203
                18 July 2019
                2019
                : 14
                : 7
                : e0220015
                Affiliations
                [1 ] Department of Plant Science, Seoul National University, Seoul, Republic of Korea
                [2 ] Research Institute of Agriculture and Life Sciences, Seoul National University, Seoul, Republic of Korea
                Zhejiang University, CHINA
                Author notes

                Competing Interests: The authors have declared that no competing interests exist.

                Author information
                http://orcid.org/0000-0002-9367-9280
                Article
                PONE-D-19-11637
                10.1371/journal.pone.0220015
                6638965
                31318958
                fcce1128-3ccb-493e-8579-9df7cd23bc98
                © 2019 Chung 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
                : 24 April 2019
                : 5 July 2019
                Page count
                Figures: 7, Tables: 3, Pages: 18
                Funding
                Funded by: the Basic Science Research Program of the National Research Foundation of Korea, the Ministry of Education
                Award ID: NRF-2013R1A1A2011925
                Award Recipient :
                This work was supported by the Basic Science Research Program of the National Research Foundation of Korea, funded by the Ministry of Education (NRF-2013R1A1A2011925 to HJL). The funder had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
                Categories
                Research Article
                Biology and Life Sciences
                Organisms
                Eukaryota
                Plants
                Fruits
                Biology and Life Sciences
                Organisms
                Eukaryota
                Plants
                Fruits
                Berries
                Blueberries
                Biology and Life Sciences
                Biochemistry
                Biosynthesis
                Biology and Life Sciences
                Cell Biology
                Signal Transduction
                Biology and life sciences
                Biochemistry
                Proteins
                DNA-binding proteins
                Transcription Factors
                Biology and Life Sciences
                Genetics
                Gene Expression
                Gene Regulation
                Transcription Factors
                Biology and Life Sciences
                Biochemistry
                Proteins
                Regulatory Proteins
                Transcription Factors
                Biology and Life Sciences
                Genetics
                Gene Expression
                Biology and Life Sciences
                Organisms
                Eukaryota
                Plants
                Fruits
                Grapes
                Biology and Life Sciences
                Computational Biology
                Genome Analysis
                Gene Ontologies
                Biology and Life Sciences
                Genetics
                Genomics
                Genome Analysis
                Gene Ontologies
                Custom metadata
                All relevant data are within the paper and its Supporting Information files.

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                Uncategorized

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