The molecular mechanisms underlying the synthesis of large cell wall polysaccharides in plant cells are not fully understood. Here, we report that 2 atypical endo-β-1,4-mannanases (MANs), which are not secreted and do not degrade glucomannan in the cell wall, play a role in glucomannan synthesis. Among the 6 MANs in Arabidopsis (Arabidopsis thaliana), AtMAN2 and AtMAN5 contain a transmembrane domain at their N-terminal region instead of a signal peptide. Subcellular localization using MAN protein fused to a fluorescent protein demonstrated that AtMAN2 localizes to the endomembrane system, including the Golgi apparatus, in xylem and interfascicular fiber cells. An Arabidopsis man2 man5 double mutant lost 65% of glucomannan in the cell walls of the inflorescence stem. Immunostaining and immunoelectron microscopic observation also revealed that the man2 man5 double mutant loses glucomannan in the cell walls to about the same extent as the csla2 csla9 double mutant, which lacks major glucomannan synthases. Gene complementation experiments showed that the enzymatic activities of AtMAN2 and AtMAN5 are important for the synthesis of cell wall glucomannan. Arabidopsis possesses another atypical MAN, AtMAN6, with an HDEL retention signal at its C-terminus. However, mutation of AtMAN6 did not affect glucomannan content in the cell walls, suggesting distinct functions for these MANs. This study has identified AtMAN2 and AtMAN5 as factors necessary for normal glucomannan synthesis in Arabidopsis, along with GDP-mannose-generating enzymes and CslAs, and suggests that glucomannan hydrolysis by these MANs contributes to maintaining glucomannan synthesis.

Brassinosteroids (BRs) are important regulators of plant growth and stress responses. To identify novel transcription factors that regulate BR responses, we screened plants generated using chimeric repressor gene-silencing technology (CRES-T) in which transcription factors were converted into chimeric repressors by the fusion of the SRDX plant-specific repression domain with brassinazole (Brz), an inhibitor of BR biosynthesis. We found that a chimeric repressor of the homeobox transcription factor BRASSINOSTEROID-RELATED-HOMEOBOX-1 (BHB1-sx) induced a Brz-insensitive phenotype with a long hypocotyl, a slender dwarf growth habit, and epinastic leaves. Furthermore, the expression of BR-biosynthesis and BR-inducible genes was enhanced in BHB1-sx. In contrast, ectopic expression of BHB1 (BHB1-ox) resulted in a Brz-hypersensitive phenotype with a short hypocotyl, a dwarf habit, and hyponastic leaves, and the expression of BR-associated genes was repressed. Transcriptome analysis revealed that the expression of auxin-related genes was down-regulated in BHB1-ox plants, whereas that of stress-related genes was up-regulated, indicating that BHB1 negatively regulates BR responses. The expression of BHB1 was induced by BR and stress conditions, and the BHB1-ox plants showed higher tolerance to salt stress compared with the wild type, whereas the bhb1 mutant showed lower tolerance. Our results indicate that BHB1 might function as a hub factor mediating the crosstalk between salt stress and BR signaling.

Oryza sativa BABY BOOM 1 (OsBBM1), a member of the AP2/ERF family of transcription factors, plays a crucial role in the initiation of the early development of rice zygotes; however, how OsBBM1 is involved in the conversion of cellular and developmental fates of egg cells to zygotes remains unclear. Egg cells isolated from transgenic rice plants expressing OsBBM1 under the egg cell-specific promoter showed nuclear and cellular divisions, suggesting that OsBBM1 functions in the conversion of the cell fate of egg cells from a quiescent to a proliferative state. Ectopically expressed OsBBM1 regulates the expression profiles of genes related to cell cycle and division and microtubule-related machinery. Moreover, although cell wall formation generally occurs around zygotes after gamete fusion, egg cells expressing OsBBM1 formed cell walls around the cells before fertilization. These suggest that OsBBM1 plays roles in egg cell-zygote conversion and the subsequent proliferative growth of zygotes.

Drought is one of the most devastating causes of yield losses in crops like maize, and the anticipated increases in severity and duration of drought spells due to climate change pose an imminent threat to agricultural productivity. To understand the drought response, phenotypic and molecular studies are typically performed at a given time point after drought onset, representing a steady-state adaptation response. Because growth is a dynamic process, we monitored the drought response with high temporal resolution and examined cellular and transcriptomic changes after rehydration at 4 and 6 days after leaf four appearance. These data showed that division zone activity is a determinant for full organ growth recovery upon rehydration. Moreover, a prolonged maintenance of cell division by the ectopic expression of PLASTOCHRON1 extends the ability to resume growth after rehydration. The transcriptome analysis indicated that GROWTH-REGULATING FACTORS (GRFs) affect leaf growth by impacting cell division duration, which was confirmed by a prolonged recovery potential of the GRF1-overexpression line after rehydration. Finally, we used a multiplex genome editing approach to evaluate the most promising differentially expressed genes from the transcriptome study and as such narrowed down the gene space from 40 to seven genes for future functional characterization.

Ultrafine bubbles (UFBs) have gained attention for their potential to improve plant growth, but their effects at the cellular level in roots are not fully understood. This study investigates the impact of UFB-enriched water on the root growth as well as root cell number and size of rice (Oryza sativa) seedlings. Our results demonstrate that UFB treatment significantly enhanced both primary and crown root lengths. The elongation rate of primary roots treated with UFB increased more rapidly than that of untreated primary roots. Conditions in which elongation rates were altered were accompanied by cell density of root tip. The findings suggest that UFBs promote root development by maintaining and enhancing the activity of the root tip, providing a novel approach to improving crop productivity through targeted cellular growth mechanisms.

Endosperm-embryo development in flowering plants is regulated coordinately by signal exchange during seed development. However, such a reciprocal control mechanism has not been clearly identified. In this study, we identified an endosperm-specific gene, LBD35, expressed in an embryonic development-dependent manner, by a comparative transcriptome and cytological analyses of double-fertilized and single-fertilized seeds prepared by using the kokopelli mutant, which frequently induces single fertilization events. Transcriptome analysis using LBD35 as a marker of the central cell fertilization event identified that 141 genes, including 31 genes for small cysteine-rich peptides, are expressed in a double fertilization-dependent manner. Our results reveal possible embryonic signals that regulate endosperm gene expression and provide a practicable method to identify genes involved in the communication during endosperm-embryo development.

Apomixis, defined as the transfer of maternal germplasm to offspring without fertilization, enables the fixation of F1-useful traits, providing advantages in crop breeding. However, most apomictic plants require pollination to produce the endosperm. The endosperm is essential for embryogenesis, and its development is suppressed until fertilization. We show that the expression of a chimeric repressor of the Elongation of Siliques without Pollination 3 (ESP3) gene (Pro35S:ESP3-SRDX) induces ovule enlargement without fertilization in Arabidopsis thaliana. The ESP3 gene encodes a protein similar to the flowering Wageningen homeodomain transcription factor containing a StAR-related lipid transfer domain. However, ESP3 lacks the homeobox-encoding region. Genes related to the cell cycle and sugar metabolism were upregulated in unfertilized Pro35S:ESP3-SRDX ovules similar to those in fertilized seeds, while those related to autophagy were downregulated similar to those in fertilized seeds. Unfertilized Pro35S:ESP3-SRDX ovules partially nourished embryos when only the egg was fertilized, accumulating hexoses without central cell proliferation. ESP3 may regulate nutrient flow during seed development, and ESP3-SRDX could be a useful tool for complete apomixis that does not require pseudo-fertilization.

Brassinosteroid (BR) is a phytohormone that acts as important regulator of plant growth. To identify novel transcription factors that may be involved in unknown mechanisms of BR signaling, we screened the chimeric repressor expressing plants (CRES-T), in which transcription factors were converted into chimeric repressors by the fusion of SRDX plant-specific repression domain, to identify those that affect the expression of BR inducible genes. Here, we identified a homeobox-leucine zipper type transcription factor, BRASSINOSTEROID-RELATED-HOMEOBOX 3 (BHB3), of which a chimeric repressor expressing plants (BHB3-sx) significantly downregulated the expression of BAS1 and SAUR-AC1 that are BR inducible genes. Interestingly, ectopic expression of BHB3 (BHB3-ox) also repressed the BR inducible genes and shorten hypocotyl that would be similar to a BR-deficient phenotype. Interestingly, both BHB3-sx and BHB3-ox showed pale green phenotype, in which the expression of genes related photosynthesis and chlorophyll contents were significantly decreased. We found that BHB3 contains three motifs similar to the conserved EAR-repression domain, suggesting that BHB3 may act as a transcriptional repressor. These results indicate that BHB3 might play an important role not only to the BR signaling but also the regulation of greenings.

The brassinosteroid (BR) phytohormone is an important regulator of plant growth. To identify novel transcription factors that regulate BR responses, we screened chimeric repressor gene silencing technology (CRES-T) plants, in which transcription factors were converted into chimeric repressors by the fusion of SRDX plant-specific repression domain, with brassinazole (Brz), an inhibitor of BR biosynthesis. We identified that a line that expressed the chimeric repressor for zinc finger homeobox transcription factor, BRASSINOSTEORID-RELATED-HOMEOBOX-2 (BHB2-sx), exhibited Brz-hypersensitive phenotype with shorter hypocotyl under dark, dwarf and round and dark green leaves similar to BR-deficient phenotype. Similar to BHB2-sx plants, bhb2 knockout mutant also exhibited Brz hypersensitive phenotype. In contrast, ectopic expression of BHB2 (BHB2-ox) showed hypocotyl elongation phenotype (BR excessive), showing decrease to Brz sensitivity. The expression of the DWF4 and CPD BR biosynthesis genes was repressed in BHB2-sx plants, whereas it was enhanced in BHB2-ox plants. The BR deficient-like phenotype of BHB2-sx plants was partially restored by treatment with brassinolide (BL), indicating that the BR deficient phenotype of BHB2-sx plant may be due to suppression of BR biosynthesis. Our results indicate that BHB2 is a positive regulator of BR response may be due to the promotion of BR biosynthesis genes.

Soil waterlogging limits plant growth and has a major impact on agricultural productivity. Here, we tested the hypothesis that the overexpression of the NCED gene improves waterlogging and reoxygenation tolerance in soybean plants. This was tested using a transgenic soybean line overexpressing the NCED gene (2Ha11) and the conventional cultivar BRS184 (wild-type; WT), which is the genetic background of the 2Ha11 line. Plants at vegetative (V6) and reproductive (R2) stages were exposed to waterlogging for 60 h and posterior reoxygenation for an additional 60 h. Overall, the NCED overexpression increased the levels of lipid peroxidation in plants exposed to waterlogging and posterior reoxygenation while decreased the antioxidant activity and alcohol dehydrogenase activity. In addition, NCED overexpression decreased seed weight and grain yield in the 2Ha11 line exposed to waterlogging at the reproductive stage. Taken together, these results suggest that the overexpression of NCED triggers an increased waterlogging sensitivity in soybean plants especially when they are exposed to waterlogging at the reproductive stage.

Adaptation to the environment is essential for sessile organisms,such as plants,as they cannot move away from adverse condi-tions.Heat stress(HS)is one of the most serious environmental stresses in terms of its effect on crop growth and yield,especially in this era of global warming.Therefore,research on heat re-sponses and acclimation in plants is becoming increasingly important.Plants have evolved complex systems to respond to HS for their survival.In general,heat shock transcription factors(Hsfs)are involved in the acquisition of thermotolerance,through regulation of the expression of genes encoding the heat shock proteins(HSPs)that function as molecular chaperones protecting cells from HS(von Koskull-D?ring et al.,2007;Richter et al.,2010).

New PhytologistVolume 231, Issue 2 p. 512-515 LettersFree Access FIBexDB: a new online transcriptome platform to analyze development of plant cellulosic fibers Natalia Mokshina, Kazan Institute of Biochemistry and Biophysics, FRC Kazan Scientific Center of RAS, Lobachevsky Str 2/31, Kazan, 420111 RussiaSearch for more papers by this authorOleg Gorshkov, orcid.org/0000-0002-7787-4262 Kazan Institute of Biochemistry and Biophysics, FRC Kazan Scientific Center of RAS, Lobachevsky Str 2/31, Kazan, 420111 RussiaSearch for more papers by this authorHironori Takasaki, Graduate School of Science and Engineering, Saitama University, Saitama, 338-8570 JapanSearch for more papers by this authorHitomi Onodera, Bioproduction Research Institute, Global Zero Emission Research Center, National Institute of Advanced Industrial Science and Technology (AIST), Tsukuba, Ibaraki, 305-8566 JapanSearch for more papers by this authorShingo Sakamoto, orcid.org/0000-0001-6019-1732 Bioproduction Research Institute, Global Zero Emission Research Center, National Institute of Advanced Industrial Science and Technology (AIST), Tsukuba, Ibaraki, 305-8566 JapanSearch for more papers by this authorTatyana Gorshkova, Corresponding Author gorshkova@kibb.knc.ru orcid.org/0000-0003-0342-8195 Kazan Institute of Biochemistry and Biophysics, FRC Kazan Scientific Center of RAS, Lobachevsky Str 2/31, Kazan, 420111 Russia Authors for correspondence: emails: gorshkova@kibb.knc.ru (TG); nobutaka.mitsuda@aist.go.jp (NM)Search for more papers by this authorNobutaka Mitsuda, Corresponding Author nobutaka.mitsuda@aist.go.jp orcid.org/0000-0001-5689-3678 Bioproduction Research Institute, Global Zero Emission Research Center, National Institute of Advanced Industrial Science and Technology (AIST), Tsukuba, Ibaraki, 305-8566 Japan Authors for correspondence: emails: gorshkova@kibb.knc.ru (TG); nobutaka.mitsuda@aist.go.jp (NM)Search for more papers by this author Natalia Mokshina, Kazan Institute of Biochemistry and Biophysics, FRC Kazan Scientific Center of RAS, Lobachevsky Str 2/31, Kazan, 420111 RussiaSearch for more papers by this authorOleg Gorshkov, orcid.org/0000-0002-7787-4262 Kazan Institute of Biochemistry and Biophysics, FRC Kazan Scientific Center of RAS, Lobachevsky Str 2/31, Kazan, 420111 RussiaSearch for more papers by this authorHironori Takasaki, Graduate School of Science and Engineering, Saitama University, Saitama, 338-8570 JapanSearch for more papers by this authorHitomi Onodera, Bioproduction Research Institute, Global Zero Emission Research Center, National Institute of Advanced Industrial Science and Technology (AIST), Tsukuba, Ibaraki, 305-8566 JapanSearch for more papers by this authorShingo Sakamoto, orcid.org/0000-0001-6019-1732 Bioproduction Research Institute, Global Zero Emission Research Center, National Institute of Advanced Industrial Science and Technology (AIST), Tsukuba, Ibaraki, 305-8566 JapanSearch for more papers by this authorTatyana Gorshkova, Corresponding Author gorshkova@kibb.knc.ru orcid.org/0000-0003-0342-8195 Kazan Institute of Biochemistry and Biophysics, FRC Kazan Scientific Center of RAS, Lobachevsky Str 2/31, Kazan, 420111 Russia Authors for correspondence: emails: gorshkova@kibb.knc.ru (TG); nobutaka.mitsuda@aist.go.jp (NM)Search for more papers by this authorNobutaka Mitsuda, Corresponding Author nobutaka.mitsuda@aist.go.jp orcid.org/0000-0001-5689-3678 Bioproduction Research Institute, Global Zero Emission Research Center, National Institute of Advanced Industrial Science and Technology (AIST), Tsukuba, Ibaraki, 305-8566 Japan Authors for correspondence: emails: gorshkova@kibb.knc.ru (TG); nobutaka.mitsuda@aist.go.jp (NM)Search for more papers by this author First published: 23 April 2021 https://doi.org/10.1111/nph.17405AboutSectionsPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onEmailFacebookTwitterLinked InRedditWechat Plant fibers constitute a major part of renewable biomass and are a valuable resource for various industries, composite material chemistry, and bioconversion processes. Plant fibers are specialized long cells with thick cell walls that are a convenient model for investigating fundamental issues such as individual cell biogenesis, cell wall formation, and cell specialization. Plant fibers usually accumulate secondary cell walls, which contain lignin, cellulose, and hemicelluloses like xylan, and are laid over thin primary cell walls. Some types of fibers like those found in the phloem of fiber crops and bending parts of hardwood tree species deposit cellulose-enriched tertiary cell walls onto the secondary cell walls (Gorshkova et al., 2018). The tertiary cell wall, also called gelatinous layer (G-layer), plays an important mechanical role; in the fibers of tension wood it serves to pull upward an inclined trunk or branch (Gardiner et al., 2014); in annual fiber crops, fibers with tertiary cell walls help to keep long and narrow stems in a vertical position. The tertiary cell wall does not contain lignin or xylan – an important advantage for plant bioconversion because these polymers hamper the saccharification efficiency of cellulose in lignocellulose. The biogenesis of specialized fibers in trees and annual fiber crops has been studied separately, despite potentially common underlying mechanisms. Thus, a comprehensive comparison of fibers from different sources is required to identify the basic common mechanisms, if present, and to determine possible differences. To tackle this issue, we chose two model plants, flax (Linum usitatissimum L.) and poplar (Populus spp.), because : (1) they are widely used for studying fiber biogenesis, and (2) even though both plants form tertiary cell walls in their fibers, the origins and mechanisms of induction are different (Gorshkova et al., 2018); (3) the poplar and flax genomes were fully sequenced (Tuskan et al., 2006; Wang et al., 2012). Primary phloem fibers in flax form tertiary cell walls constitutively during normal plant development, while xylary fibers in poplar deposit tertiary cell walls only when induced by environmental factors. Each model system has its advantages: flax fiber bundles can be easily isolated, giving the unique possibility to analyze the distinct single cell type at the certain developmental stage. Some transcriptomic studies have been conducted in fiber-containing samples of other fiber crops (Guerriero et al., 2017; Xie et al., 2020); however, datasets for purified fibers are not available for any of these species. In poplar tension wood, the development of tertiary cell walls occurs in fibers as a part of joint reactions of several cell types. Analogically, tertiary cell walls are also induced in flax xylem fibers located at the pulling stem side after plant inclination (Ibragimova et al., 2017), providing a system very similar to tension wood and permitting both interspecies and intraspecies comparisons of tertiary cell wall deposition and regulation. The emergence of NGS (next generation sequencing) technology boosted transcriptomic studies of various biological subjects, including tertiary cell wall formation, leading to the accumulation of a vast array of big data sets, most of which have been analyzed individually and have not been properly compared. To utilize these data more efficiently, a unified open database needs to be created. There are some databases containing information about gene expression in poplar: AspWood (Sundell et al., 2017) and PopGenIE (Sjödin et al., 2009). However, they do not include gene expression data during tension wood formation. By contrast, there are no available transcriptomic databases for fiber crops including flax at all. We therefore combined and normalized transcriptomic data for flax and poplar and created a new database, 'FIBexDB' (fiber expression database; https://ssl.cres-t.org/fibex/), storing our published and newly obtained data and extracted data from publicly available open resources. FIBexDB can be used to identify key common participants in the fibers of both plants during tertiary cell wall formation and to analyze the peculiarities inherent to each species. FIBexDB contains two connected but independent databases, one for each species. There are three entrances, namely (1) flax, (2) poplar, and (3) combined top pages (Fig. 1a). For flax, we collected RNA-sequencing (RNA-Seq) data for 61 samples from nine different projects, comprising transcriptomes for seven tissue types (shoot apical meristem, phloem fibers at different stages of development, core parenchyma, xylem tissue, hypocotyls, roots, and leaves), fibers from different flax cultivars and subspecies, as well as parts of mutant stems (reduced fibers) (Supporting Information Fig. S1; Tables S1, S2). For poplar, we collected RNA-Seq data for 88 samples from seven projects, comprising transcriptomes of roots, leaves, normal wood, tension wood of wild-type and transgenic poplars, and samples after treatments related to tension wood formation (Fig. S2; Tables S3, S4). To bridge the genes from the two species, information regarding homologous relationships between flax, poplar and Arabidopsis is stored. Fig. 1Open in figure viewerPowerPoint Overview of FIBexDB. (a) Top of the first page of FIBexDB. There are six links shown to the right. Furthermore, the first page has a query box (b), and three link buttons for Blast search, multiple query search, and differentially expressed gene (DEG) finder (c). There are two major ways to use FIBexDB: (1) browsing expression patterns of single or multiple genes and finding coexpressed genes by keyword (b; Keyword search), and (2) finding upregulated/downregulated genes in particular situations/tissues (c; DEG finder). Gene information contains information about gene symbol, Arabidopsis homologs, flax/poplar paralogs/homologs, PFAM, and sequence. Expression profile, coexpressed genes, and coexpression network are also shown (b). In DEG finder (c), users can set multiple search conditions even from both flax and poplar simultaneously. There are two major ways to use FIBexDB: (1) browsing expression patterns of single or multiple genes and finding coexpressed genes (gene search; Fig. 1b); and (2) finding upregulated/downregulated genes in particular situations/tissues (differentially expressed gene (DEG) finder; Fig. 1c). Gene expression can be searched by gene identifier (ID) like 'Lus10028848' for flax or 'Potri.002G080700' for poplar or by gene symbol. This gene search can also accept Arabidopsis gene symbols or locus identifiers, like 'BGAL12' or 'AT4G26140', and shows corresponding homologous genes in flax and/or poplar (Fig. 1b). The page for each flax and poplar gene (individual gene view) shows fundamental information such as coding sequence, gene symbol if present and homologous Arabidopsis and poplar or flax genes, as well as paralogous genes in each species (Fig. 1b), which are helpful to examine whether the homologous genes show similar expression patterns. The expression pattern in each tissue or condition is shown as a line graph (Fig. 1b). In addition, coexpressed genes calculated from the dataset employed in this database are also presented (Fig. 1b). The relationships between these genes are described as a coexpression network by Cytoscape (Shannon et al., 2003) using coexpression data and homology data for amino acid sequences to connect each gene (Fig. 1b). In this network view, genes with less extent of coexpression are omitted to make the number of genes in the network less than 50. Each gene in the network is labeled by its symbol or the gene symbol of the corresponding Arabidopsis gene if the flax/poplar gene does not have a symbol. If the corresponding Arabidopsis gene encodes a transcription factor (TF), it is shown as a rectangle in the network and the name of the TF family is indicated in the list of coexpressed genes. For further clarity, genes in the network are shown in several different colors representing subnetworks classified by the igraph software (Csardi & Nepusz, 2006). For example, the LusBGAL12 gene (Lus10028848) encoding β-galactosidase, which plays a significant role in tertiary cell wall development in flax fibers (Roach et al., 2011), shows specific expression in phloem fibers. However, in poplar, the closest gene Potri.006G144500 does not show high expression in tension wood. Instead, another β-galactosidase gene (Potri.002G080700, PtrBGAL16) shows specific upregulation in samples enriched with tension wood. Both Lus10028848 and Potri.002G080700 are coexpressed with genes encoding RG-I rhamnosyltransferase from the GT106 family and rhamnogalacturonan lyase family proteins involved in RG-I synthesis and modification; this indicates LusBGAL12 and PtrBGAL16 have similar functions in fibers of different origin. The sets of coexpressed TFs for LusBGAL12 and PtrBGAL16 (maximum 300 genes for each) include 19 and 9 genes, respectively, but they do not share common Arabidopsis closest genes (counterparts). The FIBexDB allows users to perform this kind of analysis easily. FIBexDB can be also used to list genes upregulated/downregulated in a particular tissue or condition via DEG finder (Fig. 1c). The DEGs in 87 and 153 comparisons and their combinations were calculated by DEseq2 package (Love et al., 2014) of R statistical software and can be extracted from flax and poplar, respectively, by simply clicking graphical icons in schematic pictures of flax and poplar (Fig. 1c). The list of extracted DEGs has information for the closest Arabidopsis gene. A unique feature of the DEG finder in FIBexDB is that users can set multiple search conditions simultaneously, even from both flax and poplar. In this case, Arabidopsis genes whose counterparts in flax and poplar meet each search condition are extracted and shown with the counterparts in flax and poplar. In this function, 'counterparts' of a particular Arabidopsis gene means flax and poplar genes that best-hit to the particular Arabidopsis gene in a Blast search. This unique feature allows users to find common important genes for tertiary cell wall formation in both species. Furthermore, the extracted genes can be classified by hierarchical or k-means clustering (de Hoon et al., 2004) by a few clicks (Fig. 1c). FIBexDB is largely aimed at analyzing flax and poplar gene expression profiles in tissues and conditions where tertiary cell walls are massively produced; this focus is not envisaged in any other database. With the collected set of samples and the system developed for data analysis, FIBexDB can provide new insights into the mechanisms of complex and important processes, such as cellulose formation, RG-I biosynthesis, and transcriptional regulatory network during tertiary cell wall formation and allows us to compare the development of distinct cell wall types namely primary, secondary, and tertiary cell walls. As tertiary cell walls develop in the fibers of many plant species under various physiological situations, the database can be further developed by adding relevant datasets to identify the important common molecular players as well as ones in specific samples. We welcome any collaboration in the plant fiber field aimed at improving and updating FIBexDB with new data for species containing fibers that form cellulose-enriched cell walls. Acknowledgements The authors thank Prof. M. Deyholos (University of British Columbia, Canada) who provided seeds of the rdf mutant and corresponding background cultivar that were used to analyze transcriptomes in a specific sample set from an unpublished study. The authors thank T. Yanagida, K. Hosono, Y. Naraki (Space-Time Inc., Sapporo, Japan) for the design and user interface of the database. The authors thank Robbie Lewis, MSc, from Edanz Group (https://en-author-services.edanz.com/ac) for editing a draft of this manuscript. This work was supported by grants from the Russian Science Foundation (grant no. 19-14-00361; RNA-Seq of flax samples from gravistimulated plants, grant no. 20-44-07005, RNA-Seq data analysis) and Strategic International Collaborative Research project (grant no. JPJ0088379) promoted by the Ministry of Agriculture, Forestry and Fisheries, Tokyo, Japan. The authors also acknowledge financial support from the government assignment for the FRC Kazan Scientific Center of RAS (N Mokshina, OG, TG data normalization and discussion). No conflict of interest is declared. Author contributions N Mitsuda developed the entire database system and HO and SS provided ideas for the user interface. OG, HT, and N Mokshina assisted with data analysis for data normalization. N Mokshina, N Mitsuda and TG designed and supervised the study. N Mokshina, N Mitsuda, and TG wrote the manuscript. All authors edited and/or made comments on the manuscript. Supporting Information Filename Description nph17405-sup-0001-FigsS1-S2.pdfPDF document, 2.9 MB Fig. S1 Schematic diagram of flax sample collection for RNA-Seq analysis. Information about flax samples is given in Tables S1 and S2. Fig. S2 Schematic diagram of poplar sample collection for RNA-Seq based on published data. Information about poplar samples is given in Tables S3 and S4 nph17405-sup-0002-TablesS1-S4.pdfPDF document, 162.5 KB Table S1 List of experiments for flax transcriptomic data. Table S2 List of samples for flax transcriptomic data. Table S3 List of experiments for poplar transcriptomic data. Table S4 List of samples for poplar transcriptomic data. Please note: Wiley Blackwell are not responsible for the content or functionality of any Supporting Information supplied by the authors. Any queries (other than missing material) should be directed to the New Phytologist Central Office. Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article. References Csardi G, Nepusz T. 2006. The igraph software package for complex network research. Inter J. Complex Systems 1695. [WWW document] URL https://igraph.org. 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SummaryMetabolites, phytohormones, and genes involved in dehydration responses/tolerance have been predicted in several plants. However, metabolite/phytohormone–gene regulatory networks in soybean organs under dehydration conditions remain unclear. Here, we analyzed the organ specificity of metabolites, phytohormones, and gene transcripts and revealed the characteristics of their regulatory networks in dehydration‐treated soybeans. Our metabolite/phytohormone analysis revealed the accumulation of raffinose, trehalose, and cis‐zeatin (cZ) specifically in dehydration‐treated roots. In dehydration‐treated soybeans, raffinose, and trehalose might have additional roles not directly involved in protecting the photosynthetic apparatus; cZ might contribute to root elongation for water uptake from the moisture region in soil. Our integration analysis of metabolites–genes indicated that galactinol, raffinose, and trehalose levels were correlated with transcript levels for key enzymes (galactinol synthase, raffinose synthase, trehalose 6‐phosphate synthase, trehalose 6‐phosphate phosphatase) at the level of individual plants but not at the organ level under dehydration. Genes encoding these key enzymes were expressed in mainly the aerial parts of dehydration‐treated soybeans. These results suggested that raffinose and trehalose are transported from aerial plant parts to the roots in dehydration‐treated soybeans. Our integration analysis of phytohormones–genes indicated that cZ and abscisic acid (ABA) levels were correlated with transcript levels for key enzymes (cytokinin nucleoside 5′‐monophosphate phosphoribohydrolase, cytokinin oxidases/dehydrogenases, 9‐cis‐epoxycarotenoid dioxygenase) at the level of individual plants but not at the organ level under dehydration conditions. Therefore, processes such as ABA and cZ transport, among others, are important for the organ specificity of ABA and cZ production under dehydration conditions.

Water deficit is an important climatic problem that can impair agriculture yield and economy. Genetically modified soybean plants containing the AtNCED3 gene were obtained aiming drought-tolerance improvement. The NCED3 gene encodes a 9-cis-epoxycarotenoid dioxygenase (NCED, EC 1.13.11.51), an important enzyme in abscisic acid biosynthesis. ABA activates the expression of drought-responsive genes, in water-deficit conditions, targeting defense mechanisms and enabling plants to survive under low water availability. Results from greenhouse experiments showed that the transgene AtNCED3 and the endogenous genes GmAREB1, GmPP2C, GmSnRK2 and GmAAO3 presented higher expression under water deficit (WD) in the event 2Ha11 than in WT-plants. No significant correlation was observed between the plant materials and WD conditions for growth parameters; however, gas exchange measurements decreased in the GM event, which also showed 80% higher intrinsic water use when compared to WT plants. In crop season 2015/16, event 2Ha11 showed higher total number of pods, higher number of pods with seeds and yield than WT plants. ABA concentration was also higher in GM plants under WD. These results obtained in field screenings suggest that AtNCED3 soybean plants might outperform under drought, reducing economic and yield losses, thus being a good candidate line to be incorporated in the soybean-breeding program to develop drought-tolerant cultivars.

The plant hormone abscisic acid (ABA) is accumulated after drought stress and plays critical roles in the responses to drought stress in plants, such as gene regulation, stomatal closure, seed maturation, and dormancy. Although previous reports revealed detailed molecular roles of ABA in stress responses, the factors that contribute to the drought-stress responses-in particular, regulation of ABA accumulation-remain unclear. The enzyme NINE-CIS-EPOXYCAROTENOID DIOXYGENASE 3 (NCED3) is essential for ABA biosynthesis during drought stress, and the NCED3 gene is highly induced by drought stress. In the present study, we isolated NGATHAs (NGAs) as candidate transcriptional regulators of NCED3 through a screen of a plant library harboring the transcription factors fused to a chimeric repressor domain, SRDX. The NGA proteins were directly bound to a cis-element NGA-binding element (NBE) in the 5' untranslated region (5' UTR) of the NCED3 promoter and were suggested to be transcriptional activators of NCED3 Among the single-knockout mutants of four NGA family genes, we found that the NGATHA1 (NGA1) knockout mutant was drought-stress-sensitive with a decreased expression level of NCED3 during dehydration stress. These results suggested that NGA1 essentially functions as a transcriptional activator of NCED3 among the NGA family proteins. Moreover, the NGA1 protein was degraded under nonstressed conditions, and dehydration stress enhanced the accumulation of NGA1 proteins, even in ABA-deficient mutant plants, indicating that there should be ABA-independent posttranslational regulations. These findings emphasize the regulatory mechanisms of ABA biosynthesis during early drought stress.

In plants, cold temperatures trigger stress responses and long-term responses that result in cold tolerance. In Arabidopsis thaliana, three dehydration-responsive element (DRE) binding protein 1/C-repeat binding factors (DREB1/CBFs) act as master switches in cold-responsive gene expression. Induction of DREB1 genes triggers the cold stress-inducible transcriptional cascade, followed by the induction of numerous genes that function in the cold stress response and cold tolerance. Many regulatory factors involved in DREB1 induction have been identified, but how these factors orchestrate the cold stress-specific expression of DREB1s has not yet been clarified. Here, we revealed that plants recognize cold stress as two different signals, rapid and gradual temperature decreases, and induce expression of the DREB1 genes. CALMODULIN BINDING TRANSCRIPTION ACTIVATOR3 (CAMTA3) and CAMTA5 respond to a rapid decrease in temperature and induce the expression of DREB1s, but these proteins do not respond to a gradual decrease in temperature. Moreover, they function during the day and night, in contrast to some key circadian components, including CIRCADIAN CLOCK ASSOCIATED1 and LATE ELONGATED HYPOCOTYL, which regulate cold-responsive DREB1 expression as transcriptional activators only during the day. Thus, plants efficiently control the acquisition of freezing tolerance using two different signaling pathways in response to a gradual temperature decrease during seasonal changes and a sudden temperature drop during the night.

Water deficit may occur at any stage of crop development, affecting productivity and causing economic losses. In response to drought, raffinose family oligosaccharides (RFOs) are accumulated in plant tissues stabilizing and protecting cell membranes and keeping the vital functions. The enzyme galactinol synthase (GolS, EC 2.4.1.123) catalyzes the first step in the biosynthesis of RFOs. In our study, soybean events overexpressing 35S:AtGolS2 were molecularly, physiological, and agronomical characterized, under drought simulated in greenhouse and in field conditions during the crop season 2014/2015. The conventional soybean cultivar BRS 184 was transformed and five positive events were obtained. Four events transmitted the transgene to further generations and in the events 2Ia1 and 2Ia4, two to four copies of AtGols2 gene were observed. Results in greenhouse showed that the overexpression of AtGolS2 in genetically modified (GM) plants led to increased galactinol transcripts, probably resulting in changes in carbohydrate metabolism. Accumulation of these transcripts that may have acted as osmoprotectors, lead to higher drought tolerance and survival rate of 2Ia4 plants. In addition, in field conditions, higher yield was observed for 2Ia4 plants under irrigated (IRR) and non-irrigated (NIRR) treatments. This result can be due to the increased synthesis of RFOs even under well-watered conditions. This field screening showed promising results for drought tolerance, suggesting that 2Ia4 plants may be useful in a breeding program for the development of drought-tolerant plants. However, additional studies are needed in further crop seasons and other sites to better characterize how these plants may outperform the WT plants under water deficit.

Leaf senescence is the terminal phenotype of plant leaf development, and ethylene is a major plant hormone inducing leaf senescence. Recent studies have shown that abscisic acid (ABA) also induces leaf senescence. However, the detailed mechanisms of ABA-induced leaf senescence remain unclear. We focused on the A subfamily of stress-responsive NAC (SNAC-A) transcription factors, the expression of which is induced by abiotic stresses, particularly ABA. Gene expression analysis revealed that seven SNAC-A genes including ANAC055, ANAC019, ANAC072/RD26, ANAC002/ATAF1, ANAC081/ATAF2, ANAC102 and ANAC032 were induced by long-term treatment with ABA and/or during age-dependent senescence. The SNAC-A septuple mutant clearly showed retardation of ABA-inducible leaf senescence. Microarray analysis indicated that SNAC-As induce ABA- and senescence-inducible genes. In addition, comparison of the expression profiles of the downstream genes of SNAC-As and ABA-responsive element (ABRE)-binding protein (AREB)/ABRE-binding factor (ABF) (AREB/ABFs) indicates that SNAC-As induce a different set of ABA-inducible genes from those mediated by AREB/ABFs. These results suggest that SNAC-As play crucial roles in ABA-induced leaf senescence signaling. We also discuss the function of SNAC-As in the transcriptional change of leaf senescence as well as in ABA response under abiotic stress conditions.