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Background Xyloglucan endotransglucosylase/hydrolase (XTH) family plays an important role in cell wall reconstruction and stress resistance in plants. However, the detailed characteristics of XTH family genes and their expression pattern under salt stress have not been reported in poplar. Results In this study, a total of 43 PtrXTH genes were identified from Populus simonii × Populus nigra, and most of them contain two conserved structures (Glyco_hydro_16 and XET_C domain). The promoters of the PtrXTH genes contain mutiple cis-acting elements related to growth and development and stress responses. Collinearity analysis revealed that the XTH genes from poplar has an evolutionary relationship with other six species, including Eucalyptus robusta, Solanum lycopersicum, Glycine max, Arabidopsis, Zea mays and Oryza sativa. Based on RNA-Seq analysis, the PtrXTH genes have different expression patterns in the roots, stems and leaves, and many of them are highly expressed in the roots. In addition, there are11 differentially expressed PtrXTH genes in the roots, 9 in the stems, and 7 in the leaves under salt stress. In addition, the accuracy of RNA-Seq results was verified by RT-qPCR. Conclusion All the results indicated that XTH family genes may play an important role in tissue specificity and salt stress response. This study will lay a theoretical foundation for further study on molecular function of XTH genes in poplar.

Global warming has led to severe drought conditions. The selection of plant varieties that can withstand drought and produce increased yields are of utmost importance. In the current study, secondary metabolites, seed trait and fiber characteristic of cottonseeds (Gossypium hirsutum) exposed to double and third water stress exposure was investigated. Total phenol and tannin content in W1S33 increased significantly after third water stress exposure. Accumulation of wax was enhanced in seeds of W3S33 and W3S34 that were subjected to third water stress. Fiber quality parameters decreased when cottonseeds were rainfed. High irrigation resulted in fragile and delicate fiber. Seeds grown under 66% FC irrigation saved water and produced seeds that had the potential of producing high quality fibers. In silico analysis was performed on cellulose synthase A (CesA) and xyloglucan endotransglycosylase (XET) enzymes present in Gossypium hirsutum. The intracellular locations of the CesA and XET1 enzymes are the plasma membrane and cell wall, respectively. Proline is conserved in the C-terminal of the CesA enzyme and plays an important role in enzyme functionality. This study provides a better understanding as to the mechanisms by which the plant can tolerate and combat water stress conditions as well as reduce water consumption. In order to grow cotton seeds with desirable morphometric characteristics and optimal fibers under water stress exposure and in dry areas, it is better to use seeds that are irrigated under optimal irrigation conditions, ie 66% FC.

All land plants encode large multigene families of xyloglucan endotransglucosylase/hydrolases (XTHs), plant-specific enzymes that cleave and reconnect plant cell-wall polysaccharides. Despite the ubiquity of these enzymes, considerable uncertainty remains regarding the evolutionary history of the XTH family. Phylogenomic and comparative analyses in this study traced the non-plant origins of the XTH family to Alphaproteobacteria ExoKs, bacterial enzymes involved in loosening biofilms, rather than Firmicutes licheninases, plant biomass digesting enzymes, as previously supposed. The relevant horizontal gene transfer (HGT) event was mapped to the divergence of non-swimming charophycean algae in the Cryogenian geological period. This HGT event was the likely origin of charophycean EG16-2s, which are putative intermediates between ExoKs and XTHs. Another HGT event in the Cryogenian may have led from EG16-2s or ExoKs to fungal Congo Red Hypersensitive proteins (CRHs) to fungal CRHs, enzymes that cleave and reconnect chitin and glucans in fungal cell walls. This successive transfer of enzyme-encoding genes may have supported the adaptation of plants and fungi to the ancient icy environment by facilitating their sessile lifestyles. Furthermore, several protein evolutionary steps, including coevolution of substrate-interacting residues and putative intra-family gene fusion, occurred in the land plant lineage and drove diversification of the XTH family. At least some of those events correlated with the evolutionary gain of broader substrate specificities, which may have underpinned the expansion of the XTH family by enhancing duplicated gene survival. Together, this study highlights the Precambrian evolution of life and the mode of multigene family expansion in the evolutionary history of the XTH family.

Flower opening is an important phenomenon in plant that indicates the readiness of the flower for pollination leading to petal expansion and pigmentation. This phenomenon has great impact on crop yield, which makes researches of its mechanism attractive for both plant physiology study and agriculture. Gene promoters directing the expression in petal during the petal cell wall modification and expansion when flower opens could be a convenient tool to analyze or monitor gene expression targeting this event. However, there are no reports of isolated gene promoters that can direct gene expression in petal or petal limb during the rapid cell wall dynamics when the flower opens. Xyloglucan endotransglucosylase/hydrolase 7 (XTH7), a cell wall modifying enzyme, was reported having up-regulated gene expression in the petal of Arabidopsis thaliana and Petunia hybrida. In this study, we fused a 1,904 bp length P. hybrida XTH7 promoter with a gene encoding a bright bioluminescent protein (Green enhanced Nano-lantern) to report gene expression and observed petal up-regulated bioluminescence activity by means of a consumer-grade camera. More importantly, this novel promoter demonstrated up-regulated activity in the petal limb of P. hybrida matured flower during flower opening. P. hybrida XTH7 promoter would be a useful tool for flowering study, especially for petal expansion research during flower opening.

Summary The xyloglucan endotransglucosylase/hydrolases (XTHs) are enzymes involved in cell wall assembly and growth regulation, cleaving and re‐joining hemicellulose chains in the xyloglucan–cellulose network. Here, in a homologous system, we compare the secretion patterns of XTH11, XTH33 and XTH29, three members of the Arabidopsis thaliana XTH family, selected for the presence (XTH11 and XTH33) or absence (XTH29) of a signal peptide, and the presence of a transmembrane domain (XTH33). We show that XTH11 and XTH33 reached, respectively, the cell wall and plasma membrane through a conventional protein secretion (CPS) pathway, whereas XTH29 moves towards the apoplast following an unconventional protein secretion (UPS) mediated by exocyst‐positive organelles (EXPOs). All XTHs share a common C‐terminal functional domain (XET‐C) that, for XTH29 and a restricted number of other XTHs (27, 28 and 30), continues with an extraterminal region (ETR) of 45 amino acids. We suggest that this region is necessary for the correct cell wall targeting of XTH29, as the ETR‐truncated protein never reaches its final destination and is not recruited by EXPOs. Furthermore, quantitative real‐time polymerase chain reaction analyses performed on 4‐week‐old Arabidopsis seedlings exposed to drought and heat stress suggest a different involvement of the three XTHs in cell wall remodeling under abiotic stress, evidencing stress‐, organ‐ and time‐dependent variations in the expression levels. Significantly, XTH29, codifying the only XTH that follows a UPS pathway, is highly upregulated with respect to XTH11 and XTH33, which code for CPS‐secreted proteins., Significance Statement The cellular distribution of three xyloglucan endotransglucosylase/hydrolases (XTHs) was monitored comparatively while expressing respective fluorescent constructs in Arabidopsis cotyledons. We show that XTH11 and XTH33 follow conventional protein secretion towards cell wall and plasma membrane, respectively, whereas XTH29, the only XTH predicted to lack a signal peptide, is secreted into the wall via exocyst‐positive organelle‐mediated unconventional protein secretion (UPS). Significantly, XTH29, poorly expressed in physiological conditions, demonstrates upregulation under drought and high temperature conditions, suggesting a specific possible role for UPS‐secreted XTH29 in stress responses.

SUMMARY Certain transglucanases can covalently graft cellulose and mixed‐linkage β‐glucan (MLG) as donor substrates onto xyloglucan as acceptor substrate and thus exhibit cellulose:xyloglucan endotransglucosylase (CXE) and MLG:xyloglucan endotransglucosylase (MXE) activities in vivo and in vitro. However, missing information on factors that stimulate or inhibit these hetero‐transglucosylation reactions limits our insight into their biological functions. To explore factors that influence hetero‐transglucosylation, we studied Equisetum fluviatile hetero‐trans‐β‐glucanase (EfHTG), which exhibits both CXE and MXE activity, exceeding its xyloglucan:xyloglucan homo‐transglucosylation (XET) activity. Enzyme assays employed radiolabelled and fluorescently labelled oligomeric acceptor substrates, and were conducted in vitro and in cell walls (in situ). With whole denatured Equisetum cell walls as donor substrate, exogenous EfHTG (extracted from Equisetum or produced in Pichia) exhibited all three activities (CXE, MXE, XET) in competition with each other. Acting on pure cellulose as donor substrate, the CXE action of Pichia‐produced EfHTG was up to approximately 300% increased by addition of methanol‐boiled Equisetum extracts; there was no similar effect when the same enzyme acted on soluble donors (MLG or xyloglucan). The methanol‐stable factor is proposed to be expansin‐like, a suggestion supported by observations of pH dependence. Screening numerous low‐molecular‐weight compounds for hetero‐transglucanase inhibition showed that cellobiose was highly effective, inhibiting the abundant endogenous CXE and MXE (but not XET) action in Equisetum internodes. Furthermore, cellobiose retarded Equisetum stem elongation, potentially owing to its effect on hetero‐transglucosylation reactions. This work provides insight and tools to further study the role of cellulose hetero‐transglucosylation in planta by identifying factors that govern this reaction., Significance Statement The enzyme HTG can graft segments of cellulose molecules onto xyloglucan (a hemicellulose), thereby re‐structuring the cell wall via hetero‐transglycosylation. In native walls, two endogenous hemicelluloses competed with cellulose (and with each other) as substrate. HTG more readily selected cellulose as substrate if an expansin‐enriched preparation was added. Hetero‐transglycosylation was inhibited by cellobiose – a potential tool for exploring HTG’s biological functions. Interestingly, cellobiose retarded Equisetum stem elongation, suggesting a role for HTG in growth.

Freezing triggers extracellular ice formation leading to cell dehydration and deformation during a freeze-thaw cycle. Many plant species increase their freezing tolerance during exposure to low, non-freezing temperatures, a process termed cold acclimation. In addition, exposure to mild freezing temperatures after cold acclimation evokes a further increase in freezing tolerance (sub-zero acclimation). Previous transcriptome and proteome analyses indicate that cell wall remodelling may be particularly important for sub-zero acclimation. In the present study, we used a combination of immunohistochemical, chemical and spectroscopic analyses to characterize the cell walls of Arabidopsis thaliana and characterized a mutant in the XTH19 gene, encoding a xyloglucan endotransglucosylase/hydrolase (XTH). The mutant showed reduced freezing tolerance after both cold and sub-zero acclimation, compared to the Col-0 wild type, which was associated with differences in cell wall composition and structure. Most strikingly, immunohistochemistry in combination with 3D reconstruction of centres of rosette indicated that epitopes of the xyloglucan-specific antibody LM25 were highly abundant in the vasculature of Col-0 plants after sub-zero acclimation but absent in the XTH19 mutant. Taken together, our data shed new light on the potential roles of cell wall remodelling for the increased freezing tolerance observed after low temperature acclimation.

Background Fruit softening is a major determinant of commercial value and shelf life. A transcriptomic analysis of 'Golden Delicious' and 'Golden Del. Reinders' (a bud mutation of 'Golden Delicious' that readily softens) apple fruit was conducted during storage. Results A comparative analysis of the obtained expression profiles of fruit between two cultivars identified 1345 upregulated and 3475 downregulated differentially expressed genes (DEGs). The DEGs identified were associated with cellular processes and carbohydrate metabolism and were especially enriched in cell-wall-related genes. Among the cell-wall-related genes, the xyloglucan endotransglucosylase/hydrolases (XTH) gene MdXTHB was significantly upregulated and exhibited high expression levels in 'Golden Del. Reinders' fruit, which had a lower level of firmness relative to 'Golden Delicious'. Overexpression of MdXTHB in both 'Golden Delicious' and 'Fuji', which typically maintain high levels of firmness in storage, exhibited faster rates of softening and an earlier peak of ethylene production than empty-vector-infiltrated fruit did. Conclusion The results of this study indicate that MdXTHB potentially promotes apple fruit softening by degrading the fruit cell wall. This result is also useful to designing further experiments on the molecular regulation of fruit softening in apple. © 2020 Society of Chemical Industry.

Lead (Pb) is a heavy metal with high toxicity to plants. Root is the major organ to respond to Pb stress. However, little is known about how plant roots perceive Pb stress signaling. Here, we describe the transcriptome of Arabidopsis root tips under Pb stress using the RNA-seq assay. A total of 703 differentially expressed genes (DEGs) were identified and expressed at every time points. Some early-responsive DEGs (1 h) were predicted to be negatively involved in cell elongation and cell expansion, while some late-responsive DEGs (24 h) positively participated in defense of oxidative stress. Hydrogen peroxide (H2O2) and superoxide (O2−) were increased significantly in root tips under Pb stress. Cell wall extension related gene XYLOGLUCAN ENDOTRANSGLUCOSYLASE/HYDROLASE 18 (XTH18) was induced in root tips, and xth18 showed reduced root growth inhibition by Pb stress. Our results revealed the potential mechanism of root growth inhibition by Pb stress and shed light for the further study.

Little is known about the physiological and molecular mechanisms underlying magnesium (Mg)-deficiency-induced enlargement, cracking and lignification of midribs and main lateral veins of Citrus leaves. Citrus sinensis (L.) Osbeck seedlings were irrigated with nutrient solution at a concentration of 0 (Mg-deficiency) or 2 (Mg-sufficiency) mM Mg(NO3)2 for 16 weeks. Enlargement, cracking and lignification of veins occurred only in lower leaves, but not in upper leaves. Total soluble sugars (glucose + fructose + sucrose), starch and cellulose concentrations were less in Mg-deficiency veins of lower leaves (MDVLL) than those in Mg-sufficiency veins of lower leaves (MSVLL), but lignin concentration was higher in MDVLL than that in MSVLL. However, all four parameters were similar between Mg-deficiency veins of upper leaves (MDVUL) and Mg-sufficiency veins of upper leaves (MSVUL). Using label-free, liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis, we identified 1229 and 492 differentially abundant proteins (DAPs) in MDVLL vs MSVLL and MDVUL vs MSVUL, respectively. Magnesium-deficiency-induced alterations of Mg, nonstructural carbohydrates, cell wall components, and protein profiles were greater in veins of lower leaves than those in veins of upper leaves. The increased concentration of lignin in MDVLL vs MSVLL might be caused by the following factors: (i) repression of cellulose and starch accumulation promoted lignin biosynthesis; (ii) abundances of proteins involved in phenylpropanoid biosynthesis pathway, hormone biosynthesis and glutathione metabolism were increased; and (iii) the abundances of the other DAPs [viz., copper/zinc-superoxide dismutase, ascorbate oxidase (AO) and ABC transporters] involved in lignin biosynthesis were elevated. Also, the abundances of several proteins involved in cell wall metabolism (viz., expansins, Rho GTPase-activating protein gacA, AO, monocopper oxidase-like protein and xyloglucan endotransglucosylase/hydrolase) were increased in MDVLL vs MSVLL, which might be responsible for the enlargement and cracking of leaf veins.

Abscisic acid (ABA) is an essential phytohormone and plays a key role in root architecture and plant stress responses. However, the ABA signalling pathway and its regulatory network in sugar beet roots remain unclear. Here, we carried out a time course experiment and performed global transcriptome profiling via strand-specific RNA sequencing (ssRNA-seq) to evaluate the response of sugar beet plants to exogenous ABA. According to the expression patterns of 5625 differentially expressed transcription units (TUs), the ABA-responsive stages within 24 h were divided into the early (1 h), intermediate (6 h and 12 h) and late (24 h) stages. Gene Ontology (GO) analysis revealed that oxidation reduction (GO: 0055114) and cell wall organization (GO: 0071555) were enriched in all ABA-responsive stages. For oxidation reduction, genes encoding cytochromes, peroxidases (PODs) and 2-oxoglutarate and Fe(II)-dependent oxygenases (2OG-Fe(II)s) constituted the largest proportion. ABA-responsive xyloglucan endotransglucosylase/hydrolase (XTH), expansin A (EXPA), pectinesterase (PME), pectate lyase (PL) and cellulose synthase (CES) were also detected in terms of cell wall organization. By performing regulation prediction and co-expression analysis, we determined that three genes, one encoding an AP2 domain-containing transcription factor (TF) and two encoding Dof domain-containing TFs (BVRB_4g074790, BVRB_8g180860 and BVRB_9g211370, respectively) may play an important role in the regulation of oxidation reduction and cell wall organization. Our profiling of ABA-responsive genes provides valuable information for understanding the molecular functions of regulatory genes and will aid in the future molecular breeding of sugar beet.

Background Xyloglucan endotransglycosylase/hydrolases (XTH) can disrupt and reconnect the xyloglucan chains, modify the cellulose-xyloglucan complex structure in the cell wall to reconstruct the cell wall. Previous studies have reported that XTH plays a key role in the aluminum (Al) tolerance of tea plants (Camellia sinensis), which is a typical plant that accumulates Al and fluoride (F), but its role in F resistance has not been reported. Results Here, 14 CsXTH genes were identified from C. sinensis and named as CsXTH1–14. The phylogenetic analysis revealed that CsXTH members were divided into 3 subclasses, and conserved motif analysis showed that all these members included catalytic active region. Furthermore, the expressions of all CsXTH genes showed tissue-specific and were regulated by Al3+ and F− treatments. CsXTH1, CsXTH4, CsXTH6–8 and CsXTH11–14 were up-regulated under Al3+ treatments; CsXTH1–10 and CsXTH12–14 responded to different concentrations of F− treatments. The content of xyloglucan oligosaccharide determined by immunofluorescence labeling increased to the highest level at low concentrations of Al3+ or F− treatments (0.4 mM Al3+ or 8 mg/L F−), accompanying by the activity of XET (Xyloglucan endotransglucosylase) peaked. Conclusion In conclusion, CsXTH activities were regulated by Al or F via controlling the expressions of CsXTH genes and the content of xyloglucan oligosaccharide in C. sinensis roots was affected by Al or F, which might finally influence the elongation of roots and the growth of plants.

Transglycanases remodel cell-wall polymers, having critical impact on many physiological processes. Unlike xyloglucan endotransglucosylase (XET) activity, widely studied in land-plants, very little is known about charophyte wall-modifying enzymes - information that would promote our understanding of the 'primordial' wall, revealing how the wall matrix is re-modelled in the closest living algal relatives of land-plants, and what changed during terrestrialisation. We conducted various in-vitro assays for wall-remodelling transglycosylases, monitoring either (a) polysaccharide-to-[3 H]oligosaccharide transglycosylation or (b) non-radioactive oligosaccharide-to-oligosaccharide transglycosylation. We screened a wide collection of enzyme extracts from charophytes (and early-diverging land-plants for comparison) and discovered several homo- and hetero-transglycanase activities. In contrast to most land-plants, charophytes possess high trans-s-1,4-mannanase activity, suggesting that land-plants' algal ancestors prioritised mannan remodelling. Trans-s-1,4-xylanase activity was also found, most abundantly in Chara, Nitella and Klebsormidium. Exo-acting transglycosidase activities (trans-s-1,4-xylosidase and trans-s-1,4-mannosidase) were also detected. In addition, charophytes exhibited homo- and hetero-trans-s-glucanase activities [XET, mixed-linkage-glucan:xyloglucan endotransglucosylase, and cellulose:xyloglucan endotransglucosylase] despite the paucity or lack of land-plant-like xyloglucan and mixed-linkage-glucan (MLG) as potential donor substrates in their cell walls. However, trans-α-xylosidase activity (which remodels xyloglucan in angiosperms) was absent in charophytes and early-diverging land-plants. Transglycanase action was also found in situ, acting on endogenous algal polysaccharides as donor-substrates and fluorescent xyloglucan-oligosaccharides as acceptor-substrates. We conclude that trans-s-mannanase and trans-s-xylanase activities are present and thus may play key roles in charophyte walls (most of which possess little or no xyloglucan and MLG, but often contain abundant s-mannans and s-xylans), comparable to the roles of XET in xyloglucan-rich land-plants.

Mangalore melon (Cucumis melo ssp. agrestis var. acidulus) is a non-dessert melon, extensively grown in the coastal districts of South India, but hardly known to the rest of the World. Immature or mature fruits of Mangalore melon are used in preparation of delicious dishes such as vegetable stew, chutneys and curries. They are appreciated for nutritional values, long shelf life and biotic stress resistance. Seventy-nine accessions of Mangalore melon were collected from five states of South India and their genetic diversity was assessed using inter simple sequence repeat (ISSR) markers. Putative candidate genes of extended shelf life in Mangalore melon were studied by quantitative reverse transcription polymerase chain reaction in comparison with cantaloupe (Cucumis melo L.). Shelf life varied from 65 days to 300 days at room temperature. Six ISSR primers amplified 142 fragments ranging from 80 bp to 2380 bp with an average of 23.66 bands per marker on a high-resolution capillary electrophoresis system. Neighbor joining phylogenetic tree construction from the ISSR allele similarity based genetic distance revealed two major clusters with 46 and 33 accessions in each cluster. Expression of fruit ripening related genes of ethylene biosynthesis (1-aminocyclopropane-1-carboxylate synthase, 1-aminocyclopropane-1-carboxylate oxidase) and cell wall metabolism (polygalacturonase, xyloglucan endotransglucosylase/hydrolase and expansin) in Mangalore melons was significantly lower than the cantaloupe melon at 180 days after harvest. Mangalore melon is a promising genetic resource for enhancing the shelf life of melons and the putative candidate genes are useful in enhancing shelf life of cantaloupe following validation and conformation.

Among the various abiotic stresses, salinity is one of the major limitations for production and productivity of wheat. Kharchia Local is the most salt-tolerant wheat cultivar developed from farmers’ selection on salt affected areas of India. Here, to investigate the molecular response of Kharchia Local under salinity stress, transcriptome sequencing of root tissue samples at the anthesis stage was performed. Illumina sequencing generated a total of 82.84 million clean reads and were assembled into 1,18,200 unigenes. A set of 10,805 unigenes were differentially expressed in response to salinity stress. Around 8232 unigene-derived SSRs were mined from these DEGs that can be used as functional molecular markers. Expression pattern of salinity stress-responsive unigenes was validated using real time PCR and results were found to be consistent with that of transcriptome profiling. Functional annotation of DEGs against GO, KEGG, COG and BLASTX using nr protein database was performed. This revealed the upregulation of genes involved in various biological processes including ROS homeostasis, ion transport, signal transduction, ABA biosynthesis and osmoregulation. Genes encoding expansin, xyloglucan endotransglucosylase/hydrolase, dehydrins and peroxidases that take part in enhancement of root growth were found to be upregulated under salinity. This could be the reason for better root growth of Kharchia Local under long-term salinity stress as compared to its susceptible counterpart. The present investigation provides primary information on transcriptome profiling of Kharchia Local roots under long-term salinity stress at the anthesis stage. In conclusion, the data generated in this study provide useful insights in understanding the molecular mechanism of salinity stress tolerance and will also serve as a valuable genomic reservoir for functional characterization of salinity responsive genes to develop tolerant genotypes.

A comparative transcriptome analysis between unripe and ripe berries of Vitis flexuosa, a Korean native wild grape, was conducted to identify differentially expressed genes (DEGs). About 12 billion nucleotides derived from unripe and ripe berries were sequenced and 180,503 contigs were obtained from the assembly results. Expression profiles of transcriptome data revealed that 5249 DEGs showed significant differences at transcript levels between unripe and ripe berries. The top five up-regulated genes in ripe berries against unripe berries were cupin family protein, cytochrome p450 79a2, HSP20-like chaperones superfamily protein, chalcone and stilbene synthase family protein, and beta-tonoplast intrinsic protein. The five most down-regulated genes in ripe berries against unripe berries were cellulose synthase family protein, laccase 17, proline-rich protein 2, subtilase 1.3, and laccase/diphenol oxidase family protein. Among DEGs, top 10 up- and down-regulated carbohydrate, organic substance, and phenylpropanoid metabolic process-related genes were selected from transcriptome analysis and their expression profiles were validated by real-time PCR. Real-time PCR analysis revealed higher expression level of DEGs encoding xyloglucan endotransglucosylase/hydrolase 32 and pectin lyase-like superfamily protein, which are related to starch and sucrose metabolism, cruciferin 3 and AGAMOUS-like 104, which are involved in organic acid metabolism, and MYB domain protein 113 and chalcone and stilbene synthase family protein, which play a role in the phenylalanine ammonia-lyase pathway in ripe than in unripe berries. Overall, we present an overview of transcriptome changes and carbohydrate, organic substance, and phenylpropanoid metabolic process-related expression patterns in unripe and ripe berries of V. flexuosa during berry ripening.

Plants have evolved various strategies to sense and respond to saline environments, which severely reduce plant growth and limit agricultural productivity. Alteration to the cell wall is one strategy that helps plants adapt to salt stress. However, the physiological mechanism of how the cell wall components respond to salt stress is not fully understood. Here, we show that expression of XTH30, encoding xyloglucan endotransglucosylase-hydrolase30, is strongly up-regulated in response to salt stress in Arabidopsis. Loss-of-function of XTH30 leads to increased salt tolerance and overexpression of XTH30 results in salt hypersensitivity. XTH30 is located in the plasma membrane and is highly expressed in the root, flower, stem, and etiolated hypocotyl. The NaCl-induced increase in xyloglucan (XyG)-derived oligosaccharide (XLFG) of the wild type is partly blocked in xth30 mutants. Loss-of-function of XTH30 slows down the decrease of crystalline cellulose content and the depolymerization of microtubules caused by salt stress. Moreover, lower Na+ accumulation in shoot and lower H2O2 content are found in xth30 mutants in response to salt stress. Taken together, these results indicate that XTH30 modulates XyG side chains, altered abundance of XLFG, cellulose synthesis, and cortical microtubule stability, and negatively affecting salt tolerance., Xyloglucan endotransglucosylase-hydrolase30 modulates xyloglucan side chains by altering abundance of xyloglucan-derived oligosaccharide, negatively affecting salt tolerance.

Maize, a moderately salt sensitive crop, first experiences osmotic stress that cause reduction in plant growth under salt stress. Fluctuation in cell wall elongation is one of the reasons of this reduction. Along with others, two important proteins expansins and xyloglucan endotransglucosylase are involved in regulation of cell wall elasticity, but the role of epigenetic mechanisms in regulating the cell wall related genes is still elusive. The present study was conducted with the aim of understanding the role of DNA methylation in regulating ZmEXPB2 and ZmXET1 genes. One salt sensitive and one salt tolerant maize cultivar was grown under hydroponic conditions at different levels of salt stress: T1 = 1 mM (control), T2 = 100 mM and T3 = 200 mM in three replicates. DNA and RNA were extracted from roots. After bisulfite treatment, Methyl Sensitive PCR was used for the DNA methylation analysis. It was revealed that fragment in promoter of ZmEXPB2 gene showed high level of DNA methylation under T1 in both varieties. Comparison of different stress treatments revealed decrease in DNA methylation with the increase in salt stress, significantly lower methylation appearing in T3. Similarly, the fragment in promoter of ZmXET1 gene also showed high levels of DNA methylation in T1. When different treatments were analysed, this gene significantly hypomethylated at T2 which continued to decrease in T3 in sensitive variety but remain stable in tolerant variety. Although, further in-depth analysis is required, our results demonstrate region-specific and genotype-specific methylation shift in the promoter of the ZmEXPB2 and ZmXET1 genes when subjected to the salt stress confirming the epigenetic regulation of these genes under stress conditions.

Chickpea (Cicer arietinum L.) is a pulse crop valued for its high protein content, grown in semi-arid tropics and Mediterranean regions. Its yield remains affected by biotic and abiotic stresses with drought alone being responsible for up to 50% annual loss of yield. Transcriptome analysis of a sensitive and a tolerant cultivar of chickpea has been done earlier to unravel the molecular basis for drought and salinity stress responses. In the present study, we performed transcriptome analysis of two drought-tolerant genotypes, BG-362 and P-256, under polyethylene glycol-simulated drought stress to decipher the genes and pathways that are commonly regulated in these genotypes. RNA-Seq using Illumina platform generated 152 million high-quality reads. Reference-guided assembly of genome yielded a total of 37,943 transcripts representing 22,701 genes. Among the 1624 genes that were observed to be differentially expressed under drought, 97 genes were common in both the genotypes. These included the upregulated genes, such as probable mannitol dehydrogenase, serine hydroxymethyltransferase 4-like, 17.5 kDa class I heat shock protein-like, cytochrome P450 81E8-like, and galactinol-sucrose galactosyltransferase-like, and downregulated genes, such as probable xyloglucan endotransglucosylase/hydrolase protein 23, abscisic acid 8′-hydroxylase 1-like, Calmodulin-like protein 11, and proline dehydrogenase 2 mitochondrial-like genes. A major finding was the involvement of transcription factors, including AP2-EREBP, bHLH, bZIP, C3H, MYB, NAC, WRKY, and MADS. The present study is the first comparative analysis of RNA-Seq data for two drought-tolerant chickpea genotypes. These findings would help in improving drought tolerance across chickpea genotypes.

Fleshy fruit ripening is governed by multiple external and internal cues and accompanied by changes in color, texture, volatiles, and nutritional quality traits. While extended shelf-life and increased phytonutrients are desired, delaying ripening via genetic or postharvest means can be accompanied by reduced nutritional value. Here we report that the high pigment 1 (hp1) mutation at the UV-DAMAGED DNA BINDING PROTEIN 1 (DDB1) locus, previously shown to influence carotenoid and additional phytonutrient accumulation via altered light signal transduction, also results in delayed ripening and firmer texture, resulting at least in part from decreased ethylene evolution. Transcriptome analysis revealed multiple ethylene biosynthesis and signaling-associated genes downregulated in hp1. Furthermore, the hp1 mutation impedes softening of the pericarp, placenta, columella as well as the whole fruit, in addition to reduced expression of the FRUITFUL2 (FUL2) MADS-box transcription factor and xyloglucan endotransglucosylase/hydrolase 5 (XTH5). These results indicate that DDB1 influences a broader range of fruit development and ripening processes than previously thought and present an additional genetic target for increasing fruit quality and shelf-life.

Buckwheat (Fagopyrum esculentum) exhibits high Al tolerance, but only a few genes have been functionally characterized. We previously characterized a half-type ABC transporter, Fagopyrum esculentum sensitive to Al rhizotoxicity1 (FeSTAR1), in buckwheat Al tolerance. This study aims to investigate whether and how another half-type ABC transporter (FeSTAR2) plays role in Al tolerance in buckwheat. The expression of FeSTAR2 and complementation test in Arabidopsis als3 mutant was examined. The interaction between FeSTAR1 and FeSTAR2 and subcellular location were analyzed by bimolecular fluorescence complementation (BiFC) and yeast two-hybrid (Y2H) assays. Al rapidly and specifically induced FeSTAR2 expression. Having transmembrane domains, FeSTAR2 localizes to membrane. BiFC and Y2H assays showed that FeSTAR2 could interact with FeSTAR1 which contains only nucleotide binding domain. Intriguingly, interaction between FeSTAR1 and FeSTAR2 altered their locations. Both heterologous expression of FeSTAR2 in als3 and exogenous UDP-glucose rescued its Al hypersensitivity of als3 mutant, suggesting that involvement of FeSTAR2 in Al tolerance requires UDP-glucose. Furthermore, Al-induced inhibition of xyloglucan endotransglucosylase (XET) activity in both atstar1 and als3 mutants could be restored by UDP-glucose. Our results indicate that FeSTAR2 interacts with FeSTAR1 to form an ABC transporter to regulate Al tolerance by vesicular transport of UDP-glucose which affects hemicellulose metabolism by regulating XET activity.

Organ abscission in plants requires the hydrolysis of cell wall components, mainly including celluloses, pectins, and xyloglucans. However, how the genes that encode those hydrolytic enzymes are regulated and their function in abscission remains unclear. Previously we revealed that two cellulase genes LcCEL2/8 and two polygalacturonase genes LcPG1/2 were responsible for the degradation of celluloses and pectins, respectively, during fruitlet abscission in litchi. Here, we further identified three xyloglucan endotransglucosylase/hydrolase genes (LcXTH4, LcXTH7, LcXTH19) that are also involved in this process. Nineteen LcXTHs, named LcXTH1-19, were identified in the litchi genome. Transcriptome data and qRT-PCR confirmed that LcXTH4/7/19 were significantly induced at the abscission zone (AZ) during fruitlet abscission in litchi. The GUS reporter driven by each promoter of LcXTH4/7/19 was specifically expressed at the floral abscission zone of Arabidopsis, and importantly ectopic expression of LcXTH19 in Arabidopsis resulted in precocious floral organ abscission. Moreover, electrophoretic mobility shift assay (EMSA) and dual-luciferase reporter analysis showed that the expression of LcXTH4/7/19 could be directly activated by two ETHYLENE INSENSITIVE 3-like (EIL) transcription factors LcEIL2/3. Collectively, we propose that LcXTH4/7/19 are involved in fruitlet abscission, and LcEIL2/3-mediated transcriptional regulation of diverse cell wall hydrolytic genes is responsible for this process in litchi.

Flesh firmness of watermelon is an important quality trait for commercial fruit values, including fruit storability, transportability, and shelf life. To date, knowledge of the gene networks underlying this trait is still limited. Herein, we used weighted genes co-expression network analysis (WGCNA) based on correlation and the association of phenotypic data (cell wall contents) with significantly differentially expressed genes between two materials, a near isogeneic line “HWF” (with high average flesh firmness) and inbred line “203Z” (with low average flesh firmness), to identify the gene networks responsible for changes in fruit flesh firmness. We identified three gene modules harboring 354 genes; these gene modules demonstrated significant correlation with water-soluble pectin, cellulose, hemicellulose, and protopectin. Based on intramodular significance, eight genes involved in cell wall biosynthesis and ethylene pathway are identified as hub genes within these modules. Among these genes, two genes, Cla012351 (Cellulose synthase) and Cla004251 (Pectinesterase), were significantly correlated with cellulose (r2 = 0.83) and protopectin (r2 = 0.81); three genes, Cla004120 (ERF1), Cla009966 (Cellulose synthase), and Cla006648 (Galactosyltransferase), had a significant correlation with water-soluble pectin (r2 = 0.91), cellulose (r2 = 0.9), and protopectin (r2 = 0.92); and three genes, Cla007092 (ERF2a), Cla004119 (probable glycosyltransferase), and Cla018816 (Xyloglucan endotransglucosylase/hydrolase), were correlated with hemicellulose (r2 = 0.85), cellulose (r2 = 0.8), and protopectin (r2 = 0.8). This study generated important insights of biosynthesis of a cell wall structure and ethylene signaling transduction pathway, the mechanism controlling the flesh firmness changes in watermelon, which provide a significant source to accelerate future functional analysis in watermelon to facilitate crop improvement.

Fruit cracking is a physiological disease that occurs during fruit development, which limits the quality and marketability of the fruit and causes great economic losses. Fruit cracking is affected by physiological, genetic and environmental factors. In this paper, the mechanism of fruit cracking was elaborated from cutin and cell wall, especially the gene families related to cell wall metabolism, including the polygalacturonase (PG) gene family, xylologlucan endotransglucosylase/hydrolase (XTH) gene family and expansin gene family. In addition, due to the advancement of high-throughput sequencing technology, an increasing number of horticultural plants have completed genome sequencing. This paper expounds the application of omics, including transcriptome, proteome, metabolomics and integrative omics in fruit cracking. The measures to reduce fruit cracking include using plastic rain covers and bagging, and spraying mineral and plant growth regulators. In this paper, the mechanisms of fruit cracking are reviewed at the molecular level, and the problems needing to be solved in fruit cracking research are put forward.

Frankincense tree (Boswellia sacra Fluek) has been poorly known on how it responds to tapping and wound-recovery process at molecular levels. Here, we used RNA-sequencing analysis to profile transcriptome of B. sacra after 30 min, 3 h and 6 h of post-tapping. Results showed 5525 differentially expressed genes (DEGs) that were related to terpenoid biosynthesis, phytohormonal regulation, cellular transport, and cell-wall synthesis. Plant-growth-regulators were applied exogenously which showed regulation of endogenous jasmonates and resulted in rapid recovery of cell-wall integrity by significantly up-regulated gene expression of terpenoid biosynthesis (germacrene-D synthase, B-amyrin synthase, and squalene epioxidase-1) and cell-wall synthesis (xyloglucan endotransglucosylase, cellulose synthase-A, and cell-wall hydrolase) compared to control. These findings suggest that tapping immediately activated several cell-developmental and regeneration processes, alongwith defense-induced terpenoid metabolism, to improve the healing process in epidermis. Exogenous growth regulators, especially jasmonic acid, can drastically help tree recovery from tissue degeneration and might help in tree conservation purposes.

The contamination of soils with cadmium (Cd) has become a serious environmental issue that needs to be addressed. Elucidating the mechanisms underlying Cd accumulation may facilitate the development of plants that accumulate both high and low amounts of Cd. In this study, a combination of phenotypic, physiological, and comparative transcriptomic analyses was performed to investigate the effects of different Cd concentrations (0, 5, 10, 30, 50 mg/kg) on Brassica juncea L. Our results suggest that B. juncea L. seedlings had a degree of tolerance to the 5 mg/kg Cd treatment, whereas higher Cd stress (10–50 mg/kg) could suppress the growth of B. juncea L. seedlings. The contents of soluble protein, as well as MDA (malondialdehyde), were increased, but the activities of CAT (catalase) enzymes and the contents of soluble sugar and chlorophyll were decreased, when B. juncea L. was under 30 and 50 mg/kg Cd treatment. Comparative transcriptomic analysis indicated that XTH18 (xyloglucan endotransglucosylase/hydrolase enzymes), XTH22, and XTH23 were down-regulated, but PME17 (pectin methylesterases) and PME14 were up-regulated, which might contribute to cell wall integrity maintenance. Moreover, the down-regulation of HMA3 (heavy metal ATPase 3) and up-regulation of Nramp3 (natural resistance associated macrophage proteins 3), HMA2 (heavy metal ATPase 2), and Nramp1 (natural resistance associated macrophage proteins 1) might also play roles in reducing Cd toxicity in roots. Taken together, the results of our study may help to elucidate the mechanisms underlying the response of B. juncea L. to various concentrations of Cd.

Low-temperature atmospheric pressure plasma technology has been used in agriculture and plant science by direct and indirect treatment of bio-samples. However, the cellular and molecular mechanisms affected by plasma-activated water (PAW) are largely unexplored. In this study, PAW generated from a surface dielectric barrier discharge (SDBD) device was used for plant development. Physicochemical analysis was performed to confirm the PAW properties that correlated with the plasma treatment time. Arabidopsis thaliana L. was utilized to study the effect of the PAW treatment in the early developmental stage. The plasma-activated water samples are denoted as PAW5 time in minutes (min), PAW7 min, PAW12 min, PAW19 min and PAW40 min with the plasma treatment time. Seedlings grown in the PAW5, PAW7 and PAW12 had increased root lengths while the root lengths were decreased in the PAW19 and PAW40. In the cellular level observation, the PAW treatment specifically increased the root hair numbers per unit of the root but suppressed the root hair length in the PAW, indicating that PAW mainly modulates the root hair cell density in the root. Furthermore, we found that the root hair density and length at PAW5 in maximal observed conditions were positively regulated by root developmental-related genes including COBRA-LIKE9 (COBL9), XYLOGLUCAN ENDOTRANSGLUCOSYLASE/HYDROLASE9 (XTH9), XTH17, AUXIN1 (AUX1) and LIKE-AUXIN (LAX3).

Fruit ripening and abscission are the results of the cell wall modification concerning different components of the signaling network. However, molecular-genetic information on the cross-talk between ripe fruit and their abscission zone (AZ) remains limited. In this study, we investigated transcriptional and hormonal changes in olive (Olea europaea L. cv Picual) pericarp and AZ tissues of fruit at the last stage of ripening, when fruit abscission occurs, to establish distinct tissue-specific expression patterns related to cell-wall modification, plant-hormone, and vesicle trafficking in combination with data on hormonal content. In this case, transcriptome profiling reveals that gene encoding members of the &alpha, galactosidase and &beta, hexosaminidase families associated with up-regulation of RabB, RabD, and RabH classes of Rab-GTPases were exclusively transcribed in ripe fruit enriched in ABA, whereas genes of the arabinogalactan protein, laccase, lyase, endo-&beta, mannanase, ramnose synthase, and xyloglucan endotransglucosylase/hydrolase families associated with up-regulation of RabC, RabE, and RabG classes of Rab-GTPases were exclusively transcribed in AZ-enriched mainly in JA, which provide the first insights into the functional divergences among these protein families. The enrichment of these protein families in different tissues in combination with data on transcript abundance offer a tenable set of key genes of the regulatory network between olive fruit tissues in late development.

Transglycanases are enzymes that remodel the primary cell wall in plants, potentially loosening and/or strengthening it. Xyloglucan endotransglucosylase (XET; EC 2.4.1.207), ubiquitous in land plants, is a homo-transglucanase activity (donor, xyloglucan; acceptor, xyloglucan) exhibited by XTH (xyloglucan endotransglucosylase/hydrolase) proteins. By contrast, hetero-trans-β-glucanase (HTG) is the only known enzyme that is preferentially a hetero-transglucanase. Its two main hetero-transglucanase activities are MLG : xyloglucan endotransglucosylase (MXE) and cellulose : xyloglucan endotransglucosylase (CXE). HTG is highly acidic and found only in the evolutionarily isolated genus of fern-allies, Equisetum. We now report genes for three new highly acidic HTG related XTHs in E. fluviatile (EfXTH-A, EfXTH-H and EfXTH-I). We expressed them heterologously in Pichia and tested the encoded proteins’ enzymic activities to determine whether their acidity and/or their Equisetum-specific sequences might confer high hetero-transglucanase activity. Untransformed Pichia was found to secrete MLG-degrading enzyme(s), which had to be removed for reliable MXE assays. All three acidic EfXTHs exhibited very predominantly XET activity, although low but measurable hetero-transglucanase activities (MXEand CXE) were also detected in EfXTH-H and EfXTH-I. We conclude that the extremely high hetero-transglucanase activities of Equisetum HTG are not emulated by similarly acidic Equisetum XTHs that share up to 55.5% sequence identity with HTG.

Cadmium (Cd) is a toxic metal element and the mechanism(s) underlying Cd tolerance in plants are still unclear. Increasingly more studies have been conducted on Cd binding to plant cell walls (CW) but most of them have focused on Cd fixation by CW pectin, and few studies have examined Cd binding to cellulose and hemicellulose. Here we found that Cd binding to CW pectin, cellulose, and hemicellulose was significantly higher in Tor-1, a Cd tolerant A. thaliana ecotype, than in Ph2-23, a sensitive ecotype, as were the concentrations of pectin, cellulose, and hemicellulose. Transcriptome analysis revealed that the genes regulating CW pectin, cellulose, and hemicellulose polysaccharide concentrations in Tor-1 differed significantly from those in Ph2-23. The expressions of most genes such as pectin methyl esterase inhibitors (PMEIs), pectin lyases, xyloglucan endotransglucosylase/hydrolase, expansins (EXPAs), and cellulose hydrolase were higher in Ph2-23, while the expressions of cellulose synthase-like glycosyltransferase 3 (CSLG3) and pectin ethyl esterase 4 (PAE4) were higher in Tor-1. The candidate genes identified here seem to regulate CW Cd fixation by polysaccharides. In conclusion, an increase in pectin demethylation activity, the higher concentration of cellulose and hemicellulose, regulated by related genes, in Tor-1 than in Ph2-23 are likely involved in enhanced Cd CW retention and reduce Cd toxicity.

The notion that xyloglucans (XG) play a pivotal role in tethering cellulose microfibrils in the primary cell wall of plants can be traced back to the first molecular model of the cell wall proposed in 1973, which was reinforced in the 1990s by the identification of Xyloglucan Endotransglucosylase/Hydrolase (XTH) enzymes that cleave and reconnect xyloglucan crosslinks in the cell wall. However, this tethered network model has been seriously challenged since 2008 by the identification of the Arabidopsis thaliana xyloglucan-deficient mutant (xxt1 xxt2), which exhibits functional cell walls. Thus, the molecular mechanism underlying the physical integration of cellulose microfibrils into the cell wall remains controversial. To resolve this dilemma, we investigated the cell wall regeneration process using mesophyll protoplasts derived from xxt1 xxt2 mutant leaves. Imaging analysis revealed only a slight difference in the structure of cellulose microfibril network between xxt1 xxt2 and wild-type (WT) protoplasts. Additionally, exogenous xyloglucan application did not alter the cellulose deposition patterns or mechanical stability of xxt1 xxt2 mutant protoplasts. These results indicate that xyloglucan is not essential for the initial assembly of the cellulose network, and the cellulose network formed in the absence of xyloglucan provides sufficient tensile strength to the primary cell wall regenerated from protoplasts.

Faba bean (Vicia faba L.) is an important food legume crop. Salinity soils severely constrain the production of faba bean, however, the seed germination of faba bean, which is a vital plant growth stage, is sensitive to salinity. Planting improved varieties of faba bean, which exhibit salt tolerance in seed germination stage, is an optimal strategy for faba bean product. To investigate the genes dynamics during the seed germination stage under salinity, RNA-seq method was used to investigate genome-wide transcription profiles of two faba bean varieties with contrast salt-tolerance during the seed germination. A total of 4,486 differentially expressed genes (DEGs) were identified among the comparison of salt-tolerant variety Y134 and salt-sensitive variety Y078 treated with salinity or not. Of these, 1,410 candidate DEGs were identified as salt-stress response genes. Furthermore, 623 DEGs were identified as variety-specific response gene during seed germination at 16 h or 24 h with salt treatment. Based on the pathway enrichment according to the Kyoto Encyclopedia of Genes and Genomes database (KEGG), these DEGs involving in cell wall loosening (e.g., xyloglucan endotransglucosylase/hydrolase, chitinase, and expansin), hormone metabolism (e.g., LEA genes, genes associated with ABA or ethylene signal pathway), chromatin remodeling (e.g., chromatin structure proteins, LHP1), small interfering RNA pathway, etc., were significantly up-regulated in salt-tolerance variety with salt treatment, indicating that they play critical roles in regulation of seed germination. The results indicated that a clearer mechanism of gene regulation that regulates the seed germination responding to salinity in faba bean. These findings are helpful to increase the understanding of the salt tolerance mechanism of crops during seed germination, and provide valuable genetic resource for the breeding of salt-tolerant faba bean varieties in future.

Current cell-wall models assume no covalent bonding between cellulose and hemicelluloses such as xyloglucan or mixed-linkage β-d-glucan (MLG). However, Equisetum hetero-trans-β-glucanase (HTG) grafts cellulose onto xyloglucan oligosaccharides (XGOs) – and, we now show, xyloglucan polysaccharide – in vitro, thus exhibiting CXE (cellulose:xyloglucan endotransglucosylase) activity. In addition, HTG also catalyzes MLG-to-XGO bonding (MXE activity). In this study, we explored the CXE action of HTG in native plant cell walls and tested whether expansin exposes cellulose to HTG by disrupting hydrogen bonds. To quantify and visualize CXE and MXE action, we assayed the sequential release of HTG products from cell walls pre-labeled with substrate mimics. We demonstrated covalent cellulose–xyloglucan bonding in plant cell walls and showed that CXE and MXE action was up to 15% and 60% of total transglucanase action, respectively, and peaked in aging, strengthening tissues: CXE in xylem and cells bordering intercellular canals and MXE in sclerenchyma. Recombinant bacterial expansin (EXLX1) strongly augmented CXE activity in vitro. CXE and MXE action in living Equisetum structural tissues potentially strengthens stems, while expansin might augment the HTG-catalyzed CXE reaction, thereby allowing efficient CXE action in muro. Our methods will enable surveys for comparable reactions throughout the plant kingdom. Furthermore, engineering similar hetero-polymer formation into angiosperm crop plants may improve certain agronomic traits such as lodging tolerance., Studying a hetero-transglucanase showed that cellulose and mixed-linkage glucan can be covalently linked to the hemicellulose xyloglucan in native plant cell walls. These reactions increase with tissue age and predominate in structural tissues and in cells surrounding cavities, suggesting a role in tissue strengthening. Expansin strongly augments cellulose hetero-transglucosylation but has no effect on mixed-linkage glucan.

The fruit physiology of banana cultivars other than Cavendish is poorly understood. To study the ripening process, samples were taken daily from plantain and Cavendish bananas and the ripening stages were determined. We present data from the green to the fully mature stage. By analyzing the protein abundances during ripening we provide some new insights into the ripening process and how plantains fruits are different. Multivariate analysis of the proteins was performed correlated to the starch dynamics. A drop in sucrose synthase and a rise of acid invertase during ripening indicated a change in the balance of the sucrose fate. During ripening, sugars may no longer be available for respiration since they are stored in the vacuoles, making citrate the preferred respiratory substrate. We found significant cultivar specific differences in granule-bound starch synthase, alpha- and beta amylases and cell wall invertase when comparing the protein content at the same ripening stage. This corroborates the difference in starch content/structure between both banana types. Differences in small heat shock proteins and in the cell wall-modifying enzyme xyloglucan endotransglucosylase/hydrolase support respectively the presumed higher carotenoid content and the firmer fruit structure of plantains. SIGNIFICANCE: We follow the proteome during ripening and correlate the proteins to the measured starch content. We discuss the changes in two contrasting genotypes. This gives us for the first time insight into the ripening of plantain and how this is different from the well-known Cavendish banana. This will revive the plantain breeding programs since for the first time we get insight into the plantain ripening. ispartof: Journal Of Proteomics vol:214 ispartof: location:Netherlands status: published

Accurate analysis of gene expression requires selection of appropriate reference genes. In this study, we report analysis of eight candidate reference genes (ACTIN, UBQ, EF-1&alpha, UBC, IF-4&alpha, TUB, PP2A, and HIS), which were screened from the genome and transcriptome data in Brassica juncea. Four statistical analysis softwares geNorm, NormFinder, BestKeeper, and RefFinder were used to test the reliability and stability of gene expression of the reference genes. To further validate the stability of reference genes, the expression levels of two CYCD3 genes (BjuB045330 and BjuA003219) were studied. In addition, all genes in the xyloglucan endotransglucosylase/hydrolase (XTH) family were identified in B. juncea and their patterns at different periods of stem enlargement were analyzed. Results indicated that UBC and TUB genes showed stable levels of expression and are recommended for future research. In addition, XTH genes were involved in regulation of stem enlargement expression. These results provide new insights for future research aiming at exploring important functional genes, their expression patterns and regulatory mechanisms for mustard development.

Recent studies have shown that histone acetylation, which is an important epigenetic modification, plays a key role in abiotic stress responses in plants. Programmed cell death and aerenchyma formation occur in cortical cells of wheat seminal roots under waterlogging stress. To explore the role of histone acetylation in aerenchyma formation of cortical cells under waterlogging, the seminal roots of two wheat cultivars, namely, Huamai 8 (waterlogging-tolerant) and Huamai 9 (waterlogging-sensitive) were investigated with waterlogging and simultaneous treatment with an acetylation inhibitor. In this study, the immunefluorescence technique and Western blotting were used to determine the histone acetylation levels in wheat seminal roots under waterlogging stress. Cell wall degradation-related enzymes (cellulase and pectinase) were observed using the method of ultracytochemical localization. We also tested the expression of related genes, such as histone acetyltransferase (HAT1), histone deacetylase (HD3), endoglucanase (CEL), polygalacturonase (PG), and xyloglucan endotransglucosylase (XET). The results indicated that histone acetylation is involved in eliciting responses to waterlogging stress as well as in the aerenchyma formation by affecting cell wall degradation of cortical cells in wheat seminal roots. We also present a model of waterlogging-induced aerenchyma formation in cortical cells of wheat seminal roots based on our experimental results and the findings of previous studies.

The fleshy taproot of radish is an important storage organ determining its yield and quality. Taproot thickening is a complex developmental process in radish. However, the molecular mechanisms governing this process remain unclear at the proteome level. In this study, a comparative proteomic analysis was performed to analyze the proteome changes at three developmental stages of taproot thickening using iTRAQ approach. In total, 1862 differentially expressed proteins (DEPs) were identified from 6342 high-confidence proteins, among which 256 up-regulated proteins displayed overlapped accumulation in S1 (pre-cortex splitting stage) vs. S2 (cortex splitting stage) and S1 vs. S3 (expanding stage) pairs, whereas 122 up-regulated proteins displayed overlapped accumulation in S1 vs. S3 and S2 vs. S3 pairs. Gene Ontology (GO) and pathway enrichment analysis showed that these DEPs were mainly involved in several processes such as “starch and sucrose metabolism”, “plant hormone signal transduction”, and “biosynthesis of secondary metabolites”. A high concordance existed between iTRAQ and RT-qPCR at the mRNA expression levels. Furthermore, association analysis showed that 187, 181, and 96 DEPs were matched with their corresponding differentially expressed genes (DEGs) in S1 vs. S2, S1 vs. S3, and S2 vs. S3 comparison, respectively. Notably, several functional proteins including cell division cycle 5-like protein (CDC5), expansin B1 (EXPB1), and xyloglucan endotransglucosylase/hydrolase protein 24 (XTH24) were responsible for cell division and expansion during radish taproot thickening process. These results could facilitate a better understanding of the molecular mechanism underlying taproot thickening, and provide valuable information for the identification of critical genes/proteins responsible for taproot thickening in root vegetable crops., Root vegetables: tapping into root thickening for improved yields An investigation into the proteins and genes responsible for taproot thickening in radish could help improve yields of root vegetables. The fleshy taproot of vegetables like radish are vital to food supplies worldwide, but the precise mechanisms behind healthy, high quality taproot formation, and thickening are unclear. Liwang Liu at Nanjing Agricultural University in Nanjing, China, and co-workers have identified several functional proteins and corresponding genes responsible for cell division in taproot growth and thickening. The team analyzed changes in protein expression at three different developmental stages of radish taproot. They identified 1862 differentially expressed proteins involved in several processes including sugar and starch metabolism, hormone signaling, and biosynthesis of metabolites. Their results provide fundamental insights into the regulation of taproot thickening in radish and could be expanded to other root vegetables.

Fruit senescence is induced by ethylene in pear, but it is unclear which ethylene responsive factor(s) is involved in. In this study, the ripening fruits of cv. Housui were respectively treated by ethephon and 1-MCP, and the treated fruits were used for transcriptome sequencing to reveal the genes associated with ethylene-induced senescence. As expected, ethylene biosynthesis and signal pathway genes were detected and had remarkably higher levels of expression in all the pre-decayed fruits than in the ripening fruits, but only one novel ERF gene (Pbr022708.1) was induced by ethylene in post-harvested fruits during storage. Moreover, based on the changes of fruit firmness, two ethylene-induced genes that individually encode polygalacturonase (Pbr010853.1) and xyloglucan endotransglucosylase/hydrolase (Pbr040203.1) were isolated to be associated with fruit softening during post-harvest storage. In addition, auxin signal and stress tolerance were likely involved into fruit senescence. These result will be available for understanding gene regulation of post-harvested fruits during storage.

Recently, two rice genes, OsAPETALA2 (OsAP2) and OsWRKY24 have been reported to be positive regulators involved in increased lamina inclination and grain size through cell elongation. Here, we found that the two genes have tightly linked expression patterns and functional convergence in rice, and are also likely to play an opposite role in Arabidopsis. Overexpression of the two rice transcription factors in Arabidopsis caused smaller plant size with reduced cell size, and the expression of a series of genes encoding expansins and xyloglucan endotransglucosylase/hydrolases (XTHs) involved in cell elongation was reduced. However, transgenic Arabidopsis expressing OsWRKY24-SRDX as a synthetic chimeric repressor displayed indistinguishable phenotypes from wild-type plants. Moreover, the subcellular localization pattern of OsWRKY24 in Arabidopsis was different from that in rice. Thus, we demonstrate an example of transcription factors from one species playing distinct roles in different plant species.

The role of xyloglucan endotransglycosylases in the regulation and promotion of plant growth in response to such widespread stress factors as drought, salinization, and hypothermia remains poorly understood. The tXET-B2 (SlXTH10) gene encodes one of the xyloglucan endotransglycosylases of tomato, which is most closely related in the nucleotide sequence to the AtXTH15 and AtXTH16 genes of Arabidopsis thaliana. At present, the specific functions of the tXET-B2 gene, as well as of its homologs, AtXTH15 and AtXTH16, remain obscure. To study the role of tXET-B2 in the regulation of growth and adaptation to abiotic stress factors, transgenic tobacco plants with estradiol-inducible expression of the tomato tXET-B2 gene were generated. Overexpression of this gene promoted tobacco root growth in a medium containing 50 mM NaCl. Under drought conditions, exogenous treatment with estradiol resulted in a considerable increase in fresh and dry weight in many of the studied transgenic lines. Under normal conditions, as well as under salinization and hypothermia stress, such positive effect was detected only for some transgenic lines. The obtained data point to the possibility of using genetically engineered constructs of the tXET-B2 gene to correct growth parameters of transgenic plants under the influence of stress factors.

Aluminum (Al) toxicity is one of the primary factors limiting crop production in acid soils worldwide. The cell wall is the major target of Al toxicity owing to the presence of many Al binding sites. Previous studies have found that XTH, encoding xyloglucan endohydrolase (XEH) and xyloglucan endotransglucosylase (XET), could participate in cell wall extension and affect the binding ability of the cell wall to Al by impeding the activities of these two enzymes. In this study, we found that ZmXTH, an XTH gene in maize, was involved in Al detoxification. The Al-induced up-regulation of ZmXTH occurred in the roots, prominently in the root tips. Additionally, the expression of ZmXTH was specifically induced by Al3+ but no other divalent or trivalent cations. Compared with the wild-type Arabidopsis, ZmXTH overexpressing plants grew more healthy and had decreased Al content in their root and root cell wall after Al stress. Overall, the results suggest that ZmXTH could confer the Al tolerance of transgenic Arabidopsis plants by reducing the Al accumulation in their roots and cell walls.

Brassinosteroids (BRs) play numerous important roles in plant growth and development. Previous studies reported that BRs could promote stem growth by regulating the expression of xyloglucan endotransglucosylase/hydrolases (XTHs). However, the mechanism of XTHs involved in stem growth remains unclear. In this study, PcBRU1, which belonged to the XTH family, was upregulated by exogenous BL treatment in Pyrus communis. The expression of PcBRU1 was highest in stems and lowest in leaves. Subcellular localization analysis indicated that PcBRU1 was located in the plasma membrane. Furthermore, overexpressing PcBRU1 in tobaccos promoted the plant height and internode length. Electron microscopy and anatomical structure analysis showed that the cell wall was significantly thinner and the cells were slenderer in transgenic tobacco lines overexpressing PcBRU1 than in wild-type tobaccos. PcBRU1 promoted stem growth as it loosened the cell wall, leading to the change in cell morphology. In addition, overexpressing PcBRU1 altered the root development and leaf shape of transgenic tobaccos. Taken together, the results could provide a theoretical basis for the XTH family in regulating cell-wall elongation and stem growth.

Citrus, as one of the most economically important fruits worldwide, is adversely affected by salinity stress. However, its molecular mechanisms underlying salinity tolerance are still not clear. In this study, next-generation RNA-seq technology was applied to analyze the gene expression profiling of citrus roots at 3 time points over a 24-h period of salt treatment. A total of 1831 differentially expressed genes (DEGs) were identified. Among them, 1195 and 1090 DEGs were found at 4 and 24 h, of which 454 were overlapped. Based on functional annotation, the salt overly sensitive (SOS) and reactive oxygen species (ROS) signaling pathways were found to be involved. Meanwhile, we found that hormone metabolism and signaling played important roles in salt stress. In addition, a multitude of transcription factors (TFs) including WRKY, NAC, MYB, AP2/ERF, bZIP, GATA, bHLH, ZFP, SPL, CBF, and CAMTA were identified. The genes related to cell wall loosening and stiffening (xyloglucan endotransglucosylase/hydrolases, peroxidases) were also involved in salt stress. Our data not only provided a genetic resource for discovering salt tolerance-related genes, but also furthered our understanding of the molecular mechanisms underlying salt tolerance in citrus.

Background The mission of the BioEnergy Science Center (BESC) was to enable efficient lignocellulosic-based biofuel production. One BESC goal was to decrease poplar and switchgrass biomass recalcitrance to biofuel conversion while not affecting plant growth. A transformation pipeline (TP), to express transgenes or transgene fragments (constructs) in these feedstocks with the goal of understanding and decreasing recalcitrance, was considered essential for this goal. Centralized data storage for access by BESC members and later the public also was essential. Results A BESC committee was established to codify procedures to evaluate and accept genes into the TP. A laboratory information management system (LIMS) was organized to catalog constructs, plant lines and results from their analyses. One hundred twenty-eight constructs were accepted into the TP for expression in switchgrass in the first 5 years of BESC. Here we provide information on 53 of these constructs and the BESC TP process. Eleven of the constructs could not be cloned into an expression vector for transformation. Of the remaining constructs, 22 modified expression of the gene target. Transgenic lines representing some constructs displayed decreased recalcitrance in the field and publications describing these results are tabulated here. Transcript levels of target genes and detailed wall analyses from transgenic lines expressing six additional tabulated constructs aimed toward modifying expression of genes associated with wall structure (xyloglucan and lignin components) are provided. Altered expression of xyloglucan endotransglucosylase/hydrolases did not modify lignin content in transgenic plants. Simultaneous silencing of two hydroxycinnamoyl CoA:shikimate hydroxycinnamoyl transferases was necessary to decrease G and S lignin monomer and total lignin contents, but this reduced plant growth. Conclusions A TP to produce plants with decreased recalcitrance and a LIMS for data compilation from these plants were created. While many genes accepted into the TP resulted in transgenic switchgrass without modified lignin or biomass content, a group of genes with potential to improve lignocellulosic biofuel yields was identified. Results from transgenic lines targeting xyloglucan and lignin structure provide examples of the types of information available on switchgrass lines produced within BESC. This report supplies useful information when developing coordinated, large-scale, multi-institutional reverse genetic pipelines to improve crop traits. Electronic supplementary material The online version of this article (10.1186/s13068-017-0991-x) contains supplementary material, which is available to authorized users.