Grape Cell Cultures Research Articles

Exosome-like Nanoparticles, High in Trans-δ-Viniferin Derivatives, Produced from Grape Cell Cultures: Preparation, Characterization, and Anticancer Properties.

Background: Recent interest in plant-derived exosome-like nanoparticles (ENs) has surged due to their therapeutic potential, which includes antioxidant, anti-inflammatory, and anticancer activities. These properties are attributed to their cargo of bioactive metabolites and other endogenous molecules. However, the properties of ENs isolated from plant cell cultures remain less explored. Methods: In this investigation, grape callus-derived ENs (GCENs) were isolated using differential ultracentrifugation techniques. Structural analysis through electron microscopy, nanoparticle tracking analysis, and western blotting confirmed that GCENs qualify as exosome-like nanovesicles. Results: These GCENs contained significant amounts of microRNAs and proteins characteristic of plant-derived ENs, as well as trans-δ-viniferin, a notable stilbenoid known for its health-promoting properties. Functional assays revealed that the GCENs reduced the viability of the triple-negative breast cancer cell line MDA-MB-231 in a dose-dependent manner. Moreover, the GCENs exhibited negligible effects on the viability of normal human embryonic kidney (HEK) 293 cells, indicating selective cytotoxicity. Notably, treatment with these GCENs led to cell cycle arrest in the G1 phase and triggered apoptosis in the MDA-MB-231 cell line. Conclusions: Overall, this study underscores the potential of grape callus-derived nanovectors as natural carriers of stilbenoids and proposes their application as a novel and effective approach in the management of cancer.

Application of Synephrine to Grape Increases Anthocyanin via Production of Hydrogen Peroxide, Not Phytohormones.

Global warming has caused such problems as the poor coloration of grape skin and the decreased production of high-quality berries. We investigated the effect of synephrine (Syn) on anthocyanin accumulation. Anthocyanin accumulation in cultured grape cells treated with Syn at concentrations of 1 mM or higher showed no significant difference, indicating that the accumulation was concentration-independent. On the other hand, anthocyanin accumulation was dependent on the compound used for treatment. The sugar/acid ratio of the juice from berries treated with Syn did not differ from the control. The expression of anthocyanin-biosynthesis-related genes, but not phytohormones, was increased by the treatment with Syn at 24 h or later. The Syn treatment of cultured cells increased SOD3 expression and hydrogen peroxide (H2O2) production from 3 to 24 h after treatment. Subsequently, the expression of CAT and APX6 encoding H2O2-scavenging enzymes was also increased. Treatment of cultured cells with Syn and H2O2 increased the expression of the H2O2-responsive gene Chit4 and the anthocyanin-biosynthesis-related genes mybA1 and UFGT 4 days after the treatment and increased anthocyanin accumulation 7 days after the treatment. On the other hand, the treatment of berries with Syn and H2O2 increased anthocyanin accumulation after 9 days. These results suggest that Syn increases anthocyanin accumulation through H2O2 production without changing phytohormone biosynthesis. Syn is expected to improve grape skin coloration and contribute to high-quality berry production.

Over 1000-Fold Synergistic Boost in Viniferin Levels by Elicitation of Vitis vinifera cv. Gamay Red Cell Cultures over Accumulating Phenylalanine.

Elicitation treatments of grape cell cultures with methyl jasmonate (MeJA), ultraviolet-C (UV-C) irradiation, and sucrose induce mild production of stilbenes and flavonoids due to limited substrate availability. However, these treatments cause a synergistic boost of stilbenes production when applied to two phenylalanine (Phe)-enriched transgenic grape cell lines, AroG* + STS and AroG* + FLS. The combined treatment of UV-C elicitation on the Phe-fed AroG* + STS line resulted in the highest content of stilbenes (37.8-fold increase, 17.39 mg/g dry weight (DW)) mainly due to resveratrol (64-fold, 3.23 mg/g DW) and viniferin (1343-fold, 13.43 mg/g DW). The synergistic increase following either UV-C or MeJA elicitation was due to the induction of stilbene-related genes, while sucrose treatment had no effect on gene expression levels and served as an additional carbon source for phenylpropanoids. The combined strategy presented may enable future usage of grape cell cultures for the production of stilbenes and in particular viniferin.

Enhancement of Stilbene Biosynthesis in Grape-Cell Cultures by Natural Products Based on Endophytes of the Wild Grape Species Vitis amurensis RUPR.

Enhancement of Stilbene Biosynthesis in Grape-Cell Cultures by Natural Products Based on Endophytes of the Wild Grape Species Vitis amurensis RUPR.

RNA inhibition of the JAZ9 gene increases the production of resveratrol in grape cell cultures

RNA inhibition of the JAZ9 gene increases the production of resveratrol in grape cell cultures

A Synchronized Increase of Stilbenes and Flavonoids in Metabolically Engineered Vitis vinifera cv. Gamay Red Cell Culture.

Stilbenes and flavonoids are two major health-promoting phenylpropanoid groups in grapes. Attempts to promote the accumulation of one group usually resulted in a decrease in the other. This study presents a unique strategy for simultaneously increasing metabolites in both groups in V. vinifera cv. Gamay Red grape cell culture, by overexpression of flavonol synthase (FLS) and increasing Phe availability. Increased Phe availability was achieved by transforming the cell culture with a second gene, the feedback-insensitive E. coli DAHP synthase (AroG*), and feeding them with Phe. A combined metabolomic and transcriptomic analysis reveals that the increase in both phenylpropanoid groups is accompanied by an induction of many of the flavonoid biosynthetic genes and no change in the expression levels of stilbene synthase. Furthermore, FLS overexpression with increased Phe availability resulted in higher anthocyanin levels, mainly those derived from delphinidin, due to the induction of F3'5'H. These insights may contribute to the development of grape berries with increased health benefits.

The Influence of the Grapevine Bacterial and Fungal Endophytes on Biomass Accumulation and Stilbene Production by the In Vitro Cultivated Cells of Vitis amurensis Rupr.

Plant endophytes are known to alter the profile of secondary metabolites in plant hosts. In this study, we identified the main bacterial and fungal representatives of the wild grape Vitis amurensis Rupr. microbiome and investigated a cocultivation effect of the 14 endophytes and the V. amurensis cell suspension on biomass accumulation and stilbene biosynthesis. The cocultivation of the V. amurensis cell culture with the bacteria Agrobacterium sp., Bacillus sp., and Curtobacterium sp. for 2 weeks did not significantly affect the accumulation of cell culture fresh biomass. However, it was significantly inhibited by the bacteria Erwinia sp., Pantoea sp., Pseudomonas sp., and Xanthomonas sp. and fungi Alternaria sp., Biscogniauxia sp., Cladosporium sp., Didymella sp. 2, and Fusarium sp. Cocultivation of the grapevine cell suspension with the fungi Didymella sp. 1 and Trichoderma sp. resulted in cell death. The addition of endophytic bacteria increased the total stilbene content by 2.2–5.3 times, while the addition of endophytic fungi was more effective in inducing stilbene accumulation by 2.6–16.3 times. The highest content of stilbenes in the grapevine cells cocultured with endophytic fungi was 13.63 and 13.76 mg/g of the cell dry weight (DW) after cultivation with Biscogniauxia sp. and Didymella sp. 2, respectively. The highest content of stilbenes in the grapevine cells cocultured with endophytic bacteria was 4.49 mg/g DW after cultivation with Xanthomonas sp. The increase in stilbene production was due to a significant activation of phenylalanine ammonia lyase (PAL) and stilbene synthase (STS) gene expression. We also analyzed the sensitivity of the selected endophytes to eight antibiotics, fluconazole, and trans-resveratrol. The endophytic bacteria were sensitive to gentamicin and kanamycin, while all selected fungal strains were resistant to fluconazole with the exception of Cladosporium sp. All endophytes were tolerant of trans-resveratrol. This study showed that grape endophytes stimulate the production of stilbenes in grape cell suspension, which could further contribute to the generation of a new stimulator of stilbene biosynthesis in grapevine or grape cell cultures.

Calcium and methyl jasmonate cross-talk in the secondary metabolism of grape cells

In grape cell cultures cv. Gamay Fréaux var. Teinturier, Ca was shown to decrease cell pigmentation through the inhibition of anthocyanin biosynthesis, while stimulating stilbenoids accumulation. Because methyl jasmonate (MeJA) is a well-known inducer of secondary metabolism in grape cells, and Ca antagonizes its stimulatory effect over several enzymes of core metabolic branches, in the present study we hypothesized that Ca and MeJA signaling pathways interact to regulate specific secondary metabolism routes. Grape cultured cells were elicited with MeJA or with MeJA + Ca and an UPLC-MS-based targeted metabolomic method was implemented to characterize their polyphenolic profiles. Results were compared with the profile of cells elicited with Ca only, previously reported. Data was complemented with gene expression analysis, allowing the assembly of a metabolic map that unraveled routes specifically regulated by both elicitors. MeJA + Ca specifically boosted E-resveratrol and E-ε-viniferin levels by 180% and 140%, respectively, in comparison to cells treated with MeJA only, while the stimulatory effect of MeJA over flavonoid synthesis was inhibited by Ca. In parallel, Ca downregulated most flavonoid pathway genes, including LAR1, ANS, BAN and ANR. Ca was able to mimic or potentiate the effect of MeJA on the expression of JA signaling genes, including JAR1, PIN and PR10. Transcript/metabolite correlation networks exposed the central influence of FLS1,STS,CDPK17 and COI1 in polyphenolic biosynthetic routes. This study highlights the potential of the MeJA-Ca combination for diverting polyphenolic metabolism towards the production of specific metabolites of interest, highly relevant in a biotechnological perspective.

Metabolic Engineering Strategy Enables a Hundred-Fold Increase in Viniferin Levels in Vitis vinifera cv. Gamay Red Cell Culture.

Stilbenes are phytoalexins with health-promoting benefits for humans. Here, we boost stilbenes' production, and in particular the resveratrol dehydrodimer viniferin, with significant pharmacological properties, by overexpressing stilbene synthase (STS) under unlimited phenylalanine (Phe) supply. Vitis vinifera cell cultures were co-transformed with a feedback-insensitive E. coli DAHP synthase (AroG*) and STS genes, under constitutive promoters. All transgenic lines had increased levels of Phe and stilbenes (74-fold higher viniferin reaching 0.74 mg/g DW). External Phe feeding of AroG* + STS lines caused a synergistic effect on resveratrol and viniferin accumulation, achieving a 26-fold (1.33 mg/g DW) increase in resveratrol and a 620-fold increase (6.2 mg/g DW) in viniferin, which to date is the highest viniferin accumulation reported in plant cultures. We suggest that this strategy of combining higher Phe availability and STS expression generates grape cell cultures as potential factories for sustainable production of stilbenes with a minor effect on the levels of flavonoids.

Intracellular Bacteria in Plants: Elucidation of Abundant and Diverse Cytoplasmic Bacteria in Healthy Plant Cells Using In Vitro Cell and Callus Cultures.

This study was initiated to assess whether the supposedly axenic plant cell cultures harbored any cultivation-recalcitrant endophytic bacteria (CREB). Adopting live-cell imaging with bright-field, fluorescent and confocal microscopy and bacterial 16S-rRNA gene taxonomic profiling, we report the cytoplasmic association of abundant and diverse CREBs in long-term actively maintained callus and cell suspension cultures of different plant species. Preliminary bright-field live-cell imaging on grape cell cultures showed abundant intracellular motile micro-particles resembling bacteria, which proved uncultivable on enriched media. Bacterial probing employing DNA stains, transmission electron microscopy, and Eubacterial FISH indicated abundant and diverse cytoplasmic bacteria. Observations on long-term maintained/freshly established callus stocks of different plant species—grapevine, barley, tobacco, Arabidopsis, and medicinal species—indicated intracellular bacteria as a common phenomenon apparently originating from field shoot tissues.Cultivation-independent 16S rRNA gene V3/V3–V4 amplicon profiling on 40-year-old grape cell/callus tissues revealed a high bacterial diversity (>250 genera), predominantly Proteobacteria, succeeded by Firmicutes, Actinobacteria, Bacteriodetes, Planctomycetes, and 20 other phyla, including several candidate phyla. PICRUSt analysis revealed diverse functional roles for the bacterial microbiome, majorly metabolic pathways. Thus, we unearth the widespread association of cultivation-recalcitrant intracellular bacteria “Cytobacts” inhabiting healthy plant cells, sharing a dynamic mutualistic association with cell hosts.

Exogenous calcium deflects grape berry metabolism towards the production of more stilbenoids and less anthocyanins

Calcium supplements have increasingly been used at pre- and post-harvest stages for improving fruit firmness, but elevated calcium levels in grape cells were shown to reduce total anthocyanin content. In this study, we hypothesized that exogenous calcium influences specific polyphenolic compounds, and performed targeted UPLC-MS analysis in fruits collected from vines cv. “Vinhão” sprayed with 2% (w/v) CaCl2 throughout the fruiting season, in two consecutive vintages, and in grape cell cultures elicited with calcium. Results showed that anthocyanin content is reduced upon calcium treatment, while stilbenoid synthesis is generally stimulated, in line with UFGT and STS expression patterns. The main metabolites involved in this response were malvidin-3-O-glucoside, E-piceid, E-ε-viniferin and E-resveratrol. The accumulation of phenolic acids, catechin and some quercetin derivatives was also favored by calcium, while other flavonols and flavan-3-ols were affected according to the vintage and berry developmental stage. In cell cultures, the entire flavonoid pathway was repressed.

Exogenous isoleucine and phenylalanine interact with abscisic acid-mediated anthocyanin accumulation in grape

Abstract Berry skin colour is a crucial determinant of red/black grape berry quality. We investigated the effects of combination treatments with amino acids and a low concentration of ABA on anthocyanin accumulation in grapes. Among the amino acids tested, isoleucine and phenylalanine resulted in high anthocyanin contents in grape cell cultures. The combination treatments with isoleucine or phenylalanine, and a low concentration of ABA enhanced anthocyanin accumulation in grape cells and detached grape berries. The combination treatment with isoleucine, but not with phenylalanine, and ABA upregulated MybA1 expression. Field-grown grapevines received combination treatments with isoleucine or phenylalanine, and ABA in two growing seasons. In the 2015 growing season, the combination treatments with isoleucine or phenylalanine, and a low concentration of ABA accelerated anthocyanin accumulation in grape berry skins of field-grown grapevines on days 10 and 31 post treatment. The effects on anthocyanin accumulation became negligible at harvest. The effect of the combination treatment with phenylalanine and a low concentration of ABA on anthocyanin accumulation was masked in the 2017 growing season due to the unexpected stimulation of anthocyanin accumulation by the low concentration of ABA, although the combination treatment accelerated anthocyanin accumulation on days 3 and 10 post treatment. Taken together, the results suggested that exogenous isoleucine and phenylalanine interacted with ABA-mediated anthocyanin accumulation in grape berry skins of field-grown grapevines when the activity of ABA used to treat the grapevines was inadequate.

Validation of reference genes for gene expression analysis of response to anthocyanin induction in cell cultures of Vitis davidii (Rom. Caill.) Foëx

Real-time quantitative polymerase chain reaction (RT-qPCR) is an effective method for detecting changes of gene expression in plant cell metabolic regulation. A set of 15 reference gene candidates were selected for the present study of anthocyanin biosynthesis regulation, and stability. The suitability of their expression was evaluated in eight different experimental treatments in spine grape (Vitis davidii [Rom. Caill.] Foex.) cell cultures. The results indicated that SAND family protein (SAND) and V-type proton ATPase subunit G (VAG) were the most stable reference genes for culture duration, tubulin alpha-3/alpha-5 chain (α-tubulin) and tubulin beta-1 chain (β-tubulin) for illumination conditions, ubiquitin-conjugating enzyme E2-17 kDa (UBQ) and VAG for UVB treatment, VAG and 60S ribosomal protein L18-2 (60SRP) for temperature treatment, AP47, clathrin adaptor complex subunit mu (AP-2) and 60SRP for cinnamic acid treatment, α-tubulin and UBQ for chitosan treatment, actin and alcohol dehydrogenase 2 (ADH2) for kinetin treatment, and β-tubulin and elongation factor 1-α (EF1-α) for cell line. Finally, the reliability of the selected reference genes was confirmed by investigating the expression profiles of the target gene dihydroflavonol 4-reductase (DFR) in spine grape cell cultures. The results of the present study offer the most robust platform for the most precise and broad application of RT-qPCR to investigate gene expression associated with anthocyanin biosynthesis in spine grape cell cultures.

Calcium- and hormone-driven regulation of secondary metabolism and cell wall enzymes in grape berry cells

The efficacy of calcium sprays for improving fleshy fruit resistance to abiotic/biotic stress and enhancement of fruit shelf life has increasingly been explored. However, because calcium is a powerful secondary messenger in many signaling pathways, including those driven by abscisic acid (ABA) and jasmonates, it may interfere with the biosynthesis of specialized metabolites highly important for fruit and wine quality, such as phenolic compounds. In this study, a combination of biochemical and molecular biology approaches were applied to grape cell cultures and detached grape berries, in order to investigate the effect of calcium in the modulation of enzymes involved in the biosynthesis of phenolic compounds and in cell wall organization. Concentrations up to 10 mM CaCl2 did not affect cell growth, size or viability, but triggered modifications in total phenolics content, particularly in anthocyanin levels in grape cell suspensions. The effects of calcium applied alone or in combination with ABA or methyl jasmonate (MeJA) were visible in several branches of specialized metabolic pathways, confirming that the calcium-hormone interplay regulates the expression of phenylalanine ammonia lyase (PAL), stilbene synthase (STS), dihydroflavonol reductase (DFR) and UDP-glucose:flavonoid 3-O-glucosyltransferase (UFGT). The activity of PAL and UFGT enzymes was also specifically modulated by calcium, ABA and MeJA. These results closely correlated to the modifications observed in the expression of VvAM1 and VvABCC1 encoding vacuolar anthocyanin transporters. Modulation of the expression and activity of pectin methyl esterases (PME) and polygalacturonases (PG) by calcium was also evident, confirming an important role of calcium in cell wall organization via the regulation of enzyme activity, besides its well-known role in the formation of cross links between pectin molecules. Overall, this study uncovers important biochemical mechanisms induced by calcium and stress hormones on grape berries, and highlights the need to consider the consequences of calcium treatments and stress for fruit quality.

Antidiabetic effects of novel cell culture established from grapevine, Vitis vinifera cv. Koshu.

Vitis vinifera cv. Koshu is an indigenous cultivar in Japan and has several characteristics that distinguish it from European V. vinifera. In Japan, Koshu is the most popular cultivar for wine making. We report herein a cell culture established from Koshu for use as a system for the production of resveratrol and its derivatives. Grape cell culture YU-1 was developed from the apex tissues of Koshu. YU-1 growth was favorably compared with BY-2 growth, a standard cell line in plant cell biology. Stilbene production and stilbene synthesis gene expression in YU-1 were upregulated by UV-C irradiation. YU-1 irradiated with UV-C decreased hemolymph sugar levels in model animals. Taken together, this study suggests that YU-1 may be used as a source of valuable medicinal components in plant cell bioreactor systems.

Improvement of Phenolic Antioxidant-linked Cancer Cell Cytotoxicity of Grape Cell Culture Elicited by Chitosan and Chemical Treatments

Grape (Vitis vinifera cv. Pok Dum) is a rich source of health relevant phenolic antioxidants and can be targeted to mitigate chronic oxidative stress commonly associated with noncommunicable chronic diseases (NCDs), such as cancer. Furthermore, improving health relevant phenolic bioactives and associated antioxidant properties of fruits by using chemical elicitation strategy has significant merit. Based on this biochemical rationale, chitosan and other chemical elicitors potassium dihydrogen phosphate (KH2PO4), potassium nitrate (KNO3), sodium selenite (Na2SeO3), and aluminum sulphate [Al2(SO4)3] were targeted to improve phenolic bioactive profiles and associated antioxidant and anticancer properties of cultured grape cells grown for 28 days. After chemical elicitor treatments, phenolic content, resveratrol content, antioxidant activity, phenylalanine ammonia-lyase (PAL) enzyme activity, and cytotoxicity (cell inhibition) against cancer cell lines of cultured grape cells were investigated using in vitro assay models. Overall, stimulation of phenolic bioactives and improvement in associated cytotoxicity against cancer cell lines were found in cultured grape cells after chemical elicitation treatments. Chitosan and other chemical elicitors resulted in lower growth of cultured grape cells; however, they enhanced phenolic biosynthesis on a cell weight basis when compared with the control. Chemical elicitor treatments, such as Na2SeO3 (50 mg·L−1 and 100 mg·L−1) and Al2(SO4)3 (50 mg·L−1), resulted in enhanced phenolic content at the end of 14 days of culture (1.7, 1.4, and 1.0-fold increase, respectively). Higher accumulation of resveratrol and higher antioxidant activity with Al2(SO4)3 (50 mg·L−1) and Na2SeO3 (100 mg·L−1) elicitation treatments were also observed. Enhanced phenolic bioactives in cultured grape cells in response to chemical elicitation treatment, such as Na2SeO3, also resulted in higher cytotoxicity against different cancer cell lines. Therefore, this study indicates that chemical elicitors, such as Na2SeO3 and Al2(SO4)3, as well as chitosan in select doses can be targeted to improve phenolic bioactives and associated antioxidant and anticancer properties in cultured grape cells and such strategy has relevance for wider applications with other phenolic antioxidant-enriched fruits.

Stilbene content and expression of stilbene synthase genes in cell cultures of Vitis amurensis treated with cinnamic and caffeic acids

It has previously been shown that exogenous application of p-coumaric acid (CA), a precursor of phenolic compounds, improved stilbene production in cell cultures of Vitis amurensis. This study examines the effect of cinnamic (Cin) and caffeic (Caf) acids, which are also phenolic precursors, on stilbene biosynthesis in the cell cultures. Five stilbenes, t-resveratrol diglucoside, t-piceid (t-resveratrol glucoside), t-resveratrol, t-ε-viniferin, and t-δ-viniferin, were found in the treated and untreated cells. Cin acid increased the total stilbene production in the grape cell cultures 2.3-3.5 times in comparison with that in the untreated cells. Caf acid increased the total stilbene production by 1.8- to 1.9-fold, but this increase was not considerably different from stilbene production in the untreated cells. Cin acid affected the total stilbene production via a marked increase in the content of t-resveratrol diglucoside (up to 2.2 times), t-piceid (up to three times), t-resveratrol (up to 5.1 times), t-ε-viniferin (up to eight times), and t-δ-viniferin (up to 9.2 times). Transcription levels of VaSTS5, 6, 7, 8, and 10 genes considerably increased under 0.1, 0.25, and 0.5 mM Cin acid. These results indicate that Cin acid increased stilbene production in V.amurensis calli via a selective enhancement of STS gene expression.

An Overview of Stress-Induced Resveratrol Synthesis in Grapes: Perspectives for Resveratrol-Enriched Grape Products.

Resveratrol is the most important stilbene phytoalexin synthesized naturally or induced in plants, as a part of their defense mechanism. Grapes and their derivative products, including juice and wine, are the most important natural sources of resveratrol, consisting of notably higher amounts than other natural sources like peanuts. Consumption of red wine with its presence of resveratrol explained the “French Paradox”. Hence, the demand of resveratrol from grapes is increasing. Moreover, as a natural source of resveratrol, grapes became very important in the nutraceutical industry for their benefits to human health. The accumulation of resveratrol in grape skin, juice, and wine has been found to be induced by the external stimuli: microbial infection, ultrasonication (US) treatment, light-emitting diode (LED), ultra violet (UV) irradiation, elicitors or signaling compounds, macronutrients, and fungicides. Phenylalanine ammonia lyase, cinnamate-4-hydroxylase, coumaroyl-CoA ligase, and stilbene synthase play a key role in the synthesis of resveratrol. The up-regulation of those genes have the positive relationship with the elicited accumulation of resveratrol. In this review, we encapsulate the effect of different external stimuli (biotic and abiotic stresses or signaling compounds) in order to obtain the maximum accumulation of resveratrol in grape skin, leaves, juice, wine, and cell cultures.

The Grapevine Uncharacterized Intrinsic Protein 1 (VvXIP1) Is Regulated by Drought Stress and Transports Glycerol, Hydrogen Peroxide, Heavy Metals but Not Water.

A MIP (Major Intrinsic Protein) subfamily called Uncharacterized Intrinsic Proteins (XIP) was recently described in several fungi and eudicot plants. In this work, we cloned a XIP from grapevine, VvXIP1, and agrobacterium-mediated transformation studies in Nicotiana benthamiana revealed that the encoded aquaporin shows a preferential localization at the endoplasmic reticulum membrane. Stopped-flow spectrometry in vesicles from the aqy-null yeast strain YSH1172 overexpressing VvXIP1 showed that VvXIP1 is unable to transport water but is permeable to glycerol. Functional studies with the ROS sensitive probe CM-H2DCFDA in intact transformed yeasts showed that VvXIP1 is also able to permeate hydrogen peroxide (H2O2). Drop test growth assays showed that besides glycerol and H2O2, VvXIP1 also transports boric acid, copper, arsenic and nickel. Furthermore, we found that VvXIP1 transcripts were abundant in grapevine leaves from field grown plants and strongly repressed after the imposition of severe water-deficit conditions in potted vines. The observed downregulation of VvXIP1 expression in cultured grape cells in response to ABA and salt, together with the increased sensitivity to osmotic stress displayed by the aqy-null yeast overexpressing VvXIP1, corroborates the role of VvXIP1 in osmotic regulation besides its involvement in H2O2 transport and metal homeostasis.

Abscisic acid transcriptomic signaling varies with grapevine organ.

BackgroundAbscisic acid (ABA) regulates various developmental processes and stress responses over both short (i.e. hours or days) and longer (i.e. months or seasons) time frames. To elucidate the transcriptional regulation of early responses of grapevine (Vitis vinifera) responding to ABA, different organs of grape (berries, shoot tips, leaves, roots and cell cultures) were treated with 10 μM (S)-(+)-ABA for 2 h. NimbleGen whole genome microarrays of Vitis vinifera were used to determine the effects of ABA on organ-specific mRNA expression patterns.ResultsTranscriptomic analysis revealed 839 genes whose transcript abundances varied significantly in a specific organ in response to ABA treatment. No single gene exhibited the same changes in transcript abundance across all organs in response to ABA. The biochemical pathways affected by ABA were identified using the Cytoscape program with the BiNGO plug-in software. The results indicated that these 839 genes were involved in several biological processes such as flavonoid metabolism, response to reactive oxygen species, response to light, and response to temperature stimulus. ABA affected ion and water transporters, particularly in the root. The protein amino acid phosphorylation process was significantly overrepresented in shoot tips and roots treated with ABA. ABA affected mRNA abundance of genes (CYP707As, UGTs, and PP2Cs) associated with ABA degradation, conjugation, and the ABA signaling pathway. ABA also significantly affected the expression of several transcription factors (e.g. AP2/ERF, MYC/MYB, and bZIP/AREB). The greatest number of significantly differentially expressed genes was observed in the roots followed by cell cultures, leaves, berries, and shoot tips, respectively. Each organ had a unique set of gene responses to ABA.ConclusionsThis study examined the short-term effects of ABA on different organs of grapevine. The responses of each organ were unique indicating that ABA signaling varies with the organ. Understanding the ABA responses in an organ-specific manner is crucial to fully understand hormone action and plant responses to water deficit.Electronic supplementary materialThe online version of this article (doi:10.1186/s12870-016-0763-y) contains supplementary material, which is available to authorized users.