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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.

The aim of this study was to contribute to the reduction of synthetic chemical fungicide application in viticulture by using cyclo(- l -Leu- l -Phe) (cLF) produced by Bacillus subtilis KS1, a candidate for biological control agent. cLF is a diketopiperazine and belongs to the family of 2,5-diketopiperazines. KS1 secreted micromolar levels of cLF into culture medium. Micromolar concentrations of cLF structure-dependently decreased by ∼90% both disease severity and lesion density of downy mildew in grapevine cultivated in a growth chamber. Microscopic observation demonstrated that cLF inhibited Plasmopara viticola haustorium formation by 80% but not zoospore germination on leas disks. Interestingly, millimolar concentrations of cLF induced plant defense response, such as expression of genes encoding chitinase and β-1,3-glucanase, in grapevine leaves through the salicylic acid and jasmonate signaling pathways. We demonstrated that cLF was a weapon against P. viticola infection. Further evaluation of cLF in field trials is required to uncover its inherent characteristics. [ABSTRACT FROM AUTHOR]

Climate change, particularly warmer temperature, has resulted in reduced anthocyanin accumulation in grape berry skin. Because anthocyanin is a crucial determinant of red wine quality, viticulturists need to devise a solution for mitigating the poor coloration of red/black grape berry skin under elevated temperature conditions. In this study, we investigated the effects of geraniol on anthocyanin accumulation in grape berry skins of field-grown grapevines and elucidated the molecular mechanisms of the geraniol-triggered anthocyanin accumulation. Geraniol-treated bunches showed enhanced anthocyanin accumulation in berry skins at harvest (50 days after treatment). Geraniol treatment upregulated the transcription of MybA1 and UFGT, which encode the key factors in anthocyanin biosynthesis, in berry skins. Geraniol treatment also improved anthocyanin accumulation in grape cultured cells. We isolated grape ATP-binding cassette transporter G family protein VvABCG40, encoding abscisic acid (ABA) membrane transporter, from geraniol-treated grape cultured cells. VvABCG40 transcription was upregulated in berry skins 40 days after treatment. Geraniol treatment also upregulated the transcription of VvPP2C24, which encodes ABA-responsible type 2C protein phosphatases, in berry skins, but not the transcription of VvNCED1, which encodes a key enzyme in ABA biosynthesis. Taken together, geraniol-triggered anthocyanin accumulation in berry skins is promoted by ABA membrane transport and not by ABA biosynthesis, and geraniol treatment of field-grown grape bunches may contribute to alleviating the poor coloration of berry skin as a novel technique in viticulture. [ABSTRACT FROM AUTHOR]

Vitis vinifera cv. Koshu is an indigenous grape cultivar that has been cultivated for more than a thousand years in Japan and one of the most important cultivars in white winemaking. To improve Koshu wine quality, it is necessary to identify the metabolites in Koshu berry. We conducted a comprehensive and comparative lipidome analysis of Koshu and Pinot Noir berries cultivated in the same location in Japan using GC-MS/MS for fatty acids and LC-MS for glycerolipids and glycerophospholipids. Koshu skins and juices contained 22 and 19 fatty acids, respectively, whereas 23 and 20 fatty acids were detected in Pinot Noir skins and juices. C22:6n3 and C24:0 contents in Koshu skins were two and three times higher than those in Pinot Noir skins. C24:0 content in Koshu juices was also higher than that in Pinot Noir juices. Forty-nine lipid components (six digalactosyldiacylglycerols, one monogalactosyldiacylglycerol, 10 phosphatidylcholines, 12 phosphatidylethanolamines, and 20 triglycerides) were detected in Pinot Noir and Koshu skins. Strong peaks were observed for MGDG 36:6, DGDG 36:6, PC 34:2, PC 36:5, TG 54:6, TG 54:7, and TG 54:8 in Koshu skins. The contents of 36 of the 49 lipid components were significantly higher in Pinot Noir skins than Koshu skins. Pinot Noir skins contained more lipids whose alkyl chains have more than 18 carbons than Koshu skins. Further analysis of both lipid profiles revealed that the number of double bonds in a fatty acid molecule in Pinot Noir skins and juices was significantly larger than that in Koshu skins and juices. A strong relationship exists between the heat requirement of grapevine cultivars and the level of fatty acid desaturation. C18-fatty acids were the major components in Koshu and Pinot Noir berries. The expression levels of C18-fatty acid desaturases regulated the accumulation of C18-unsaturated fatty acids in berry skins. The loss of C18:3 in Koshu berries at the end of ripening was observed. Koshu might effectively convert C18:3 into (Z)-hex-3-enal for the production of C6-aroma compounds. These findings by the lipidome analysis are expected to contribute to the improvement of Koshu wine aroma and breeding strategies of cold-tolerant Koshu grapevines. [ABSTRACT FROM AUTHOR]

The article studies the nested polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method to identify fungicide-resistant Botrytia cinerea. When the nested PCR-RFLP method was carried out, it successfully detected that benzimidazole-, phenylcarbarnate- and dicarboximide-resistant Botrytis cinerea isolates at an early growth stage of grapes. It was suggested that the early diagnosis of Botrytis cinerea would contribute to further improvement of integrated pest management against Botrytis cinerea in grapevine fields.

Global warming has resulted in the loss of anthocyanin accumulation in berry skin. Sound stimulation can be used as a potential method for enhancing fruit color development since many plants recognize sound vibration as an external stimulus and alter their physiological status in response to it. Sound stimulation (sine wave sound at 1000 Hz) enhanced anthocyanin accumulation in grape cultured cells and berry skins in field-grown grapevines at the early stage of ripening. The transcription of UFGT and ACO2, which encode the key enzymes in anthocyanin and ethylene biosynthesis, respectively, was upregulated in grape cultured cells exposed to sound stimulation. In contrast, the transcription of MybA1 and NCED1, which encode a transcription factor for UFGT and a key enzyme in abscisic acid biosynthesis, respectively, was not affected by the sound stimulation. A treatment with an ethylene biosynthesis inhibitor, aminoethoxyvinyl glycine hydrochloride, revered the enhancement of anthocyanin accumulation by sound stimulation. As the promoter assay using a GUS reporter gene demonstrated that UFGT promoter was directly activated by the ethylene-releasing compound ethephon, which enhanced anthocyanin accumulation in grape cultured cells, we conclude that sound stimulation enhanced anthocyanin accumulation through the direct upregulation of UFGT by ethylene biosynthesis. Our findings suggest that sound stimulation contributes to alleviating poor coloration in berry skin as a novel and innovative practical technique in viticulture. [ABSTRACT FROM AUTHOR]

Concern over environmental pollution generated by chemical fungicides has led to the introduction of alternative pest management strategies to chemical fungicide application. One of those strategies is the induction of plant defense response by an abiotic elicitor. In the present study, field-grown grapevines were subjected to electrical stimulation using a solar panel from two weeks before flowering to harvest in the 2016 and 2020 growing seasons. In both years, electrical stimulation decreased the incidence of gray mold and/or ripe rot on bunches and downy mildew on leaves of the field-grown grapevine. Transcription of a gene encoding β-1,3-glucanase but not class IV chitinase in leaves of potted grapevine seedlings was upregulated 20 days after electrical stimulation, suggesting that electrical stimulation acts as an abiotic elicitor of plant defense response to fungal diseases. The gene expression of PR1 but not PDF1.2 was upregulated in Arabidopsis plants subjected to electrical stimulation. On the other hand, PR1 gene expression was not induced in salicylic acid (SA)-insensitive Arabidopsis mutant npr1-5 subjected to electrical stimulation. Taken together, electrical stimulation is responsible for plant defense response through the SA-dependent defense pathway. These findings would help us develop a novel and innovative practical technique that uses electrical stimulation in integrated pest management. [ABSTRACT FROM AUTHOR]

In grapes, the number of flowers per inflorescence determines the compactness of grape bunches. Grape cultivars with tight bunches and thin-skinned berries easily undergo berry splitting, especially in growing areas with heavy rainfall during the grapevine growing season, such as Japan. We report herein that grape cytokinin oxidase/dehydrogenase 5 (VvCKX5) determines the number of berries per inflorescence in grapes. The number of berries per bunch was inversely proportional to the VvCKX5 expression level in juvenile inflorescences among the cultivars tested. VvCKX5 overexpression drastically decreased the number of flower buds per inflorescence in Arabidopsis plants, suggesting that VvCKX5 might be one of the negative regulators of the number of flowers per inflorescence in grapes. Similarly, the overexpression of grape sister of ramose 3 (VvSRA), which encodes trehalose 6-phosphate phosphatase that catalyzes the conversion of trehalose-6-phosphate into trehalose, upregulated AtCKX7 expression in Arabidopsis plants, leading to a decrease in the number of flower buds per Arabidopsis inflorescence. VvCKX5 gene expression was upregulated in grapevine cultured cells and juvenile grape inflorescences treated with trehalose. Finally, injecting trehalose into swelling buds nearing bud break using a microsyringe decreased the number of berries per bunch by half. VvCKX5 overexpression in Arabidopsis plants had no effect on the number of secondary inflorescences from the main inflorescence, and similarly trehalose did not affect pedicel branching on grapevine inflorescences, suggesting that VvCKX5, as well as VvSRA-mediated trehalose metabolism, regulates flower formation but not inflorescence branching. These findings may provide new information on the crosstalk between VvSRA-mediated trehalose metabolism and VvCKX-mediated cytokinin degradation for determining the number of berries per bunch. Furthermore, this study is expected to contribute to the development of innovative cultivation techniques for loosening tight bunches. [ABSTRACT FROM AUTHOR]

Vine vigor or vegetative growth is an important factor related to berry quality and vinicultural training management, but brassinosteroid (BR)-induced molecular mechanisms underlying vine growth remain unclear. In this study, the hypothesis that the Vitis vinifera CYP90D1 gene VvCYP90D1 , one of the genes for BR biosynthesis, plays a critical role in shoot elongation was tested. RNA sequencing analysis of shoots collected from the vigorous cultivar Koshu (KO) and the reference cultivar Pinot Noir (PN) 7 days after bud break showed higher expression levels of various genes in the BR biosynthesis pathway in KO than in PN. The VvCYP90D1 expression level in KO was highest in meristems, followed by internodes and leaves. Cluster analysis of amino acid sequences including those in other plant species showed that the isolated gene belonged to the CYP90D1 group. The vegetative growth and the endogenous BR (brassinolide; BL) content were significantly higher in VvCYP90D1-overexpressing Arabidopsis than in wild type. VvCYP90D1-overexpressing Arabidopsis treated with brassinazole (Brz), a BR biosynthesis inhibitor, showed recovery of vegetative growth. These results indicate that VvCYP90D1 in grapevine has a vegetative growth promoting effect via BR biosynthesis. Our findings on the mechanism of BR-induced grape shoot growth will contribute to the development of new shoot control techniques for grapevine. • BR biosynthesis pathway genes were highly expressed in vigorous Koshu grapevine. • Overexpression of grape cytochrome P450 CYP90D1 increased endogenous BR content in Arabidopsis thaliana. • VvCYP90D1 increased vegetative growth in Arabidopsis thaliana. [ABSTRACT FROM AUTHOR]

Plants are known to utilize electrical signals under several physiological conditions and electrical stimulation from the outside induces physiological changes in plants. In this study, to improve grape berry composition in field-grown grapevines, electrical stimulation of grapevine using solar panels was undertaken as an abiotic stress generator in 2015 and 2016 growing seasons. Electrical stimulation had a notable effect on grapevine growth and development as well as photosynthetic performance. Berry weight and tartaric acid and total phenolic contents in berries of grapevines exposed to electrical stimulation were similar among the grapevines tested. Brix in berries of grapevines exposed to electrical stimulation and electrode-treated grapevines was higher than that in control grapevines in both years. Electrical stimulation increased anthocyanin and resveratrol contents in berries of grapevines in both years relative to those of control grapevines and electrode-treated grapevines. The alteration of Brix and anthocyanin and resveratrol contents in berries was supported by the results of microarray analysis demonstrating the transcriptional upregulation of genes related to sucrose metabolism, phenylpropanoid biosynthesis, flavonoid biosynthesis, stilbenoid biosynthesis, and anthocyanin biosynthesis in grape cells exposed to electrical stimulation. Taken together, the results suggested that electrical stimulation of grapevine enhanced anthocyanin and resveratrol biosynthesis by activating their biosynthetic pathways. [ABSTRACT FROM AUTHOR]

We investigated the effect of vanillylacetone (VA) on anthocyanin accumulation with aim of improving grape berry coloration. Spraying Vitis vinifera cv. Muscat Bailey A berries with VA at veraison increased sugar/acid ratio, an indicator of maturation and total anthocyanin accumulation. To elucidate the molecular mechanism underlying the effect of VA on anthocyanin accumulation, in vitro VA treatment of a grapevine cell culture was carried out. Endogenous abscisic acid (ABA) content was higher in the VA-treated cell cultures than in control at 3 h after treatment. Consistent with this, the relative expression levels of anthocyanin-synthesis-related genes, including DFR , LDOX , MybA1 and UFGT, in VA-treated cell cultures were much higher than those in control, and high total anthocyanin accumulation was noted in the VA-treated cell cultures as well. These results suggest that VA up-regulates the expression of genes leading to anthocyanin accumulation by inducing endogenous ABA. In addition, VA increased total anthocyanin content in a dose-dependent manner. Although VA treatment in combination with exogenous ABA did not exhibit any synergistic effect, treatment with VA alone showed an equivalent effect to that with exogenous ABA alone on total anthocyanin accumulation. These findings point to the possibility of using VA for improving grape berry coloration. [ABSTRACT FROM AUTHOR]

To identify apoplasmic proteins in grape cells, we investigated proteins secreted into grape cell culture medium and detected β-1,3-glucanase VvGHF17 in the medium. VvGHF17 is expressed constitutively in grape leaves and berry pulp and skin. Experiments using VvGHF17:GFP fusion protein demonstrated that VvGHF17 is localized at the apoplasmic space of cells, suggesting that VvGHF17 is apoplasmic β-1,3-glucanase. To understand how VvGHF17 protects grapevine against phytopathogens, we created Arabidopsis transgenic plants expressing VvGHF17. VvGHF17-expressing Arabidopsis plants exhibited disease resistance to both Botrytis cinerea and Colletotrichum higginsianum , but not to Pseudomonas syringae pv. tomato DC3000. The inhibitory effect of VvGHF17 to the infection by the fungi was promoted when VvGHF17 protein was expressed in large amounts in the plants. In conclusion, apoplasmic VvGHF17 may be a candidate for the transgene to create transgenic grapevines that are robust to multiple fungal attacks. [ABSTRACT FROM AUTHOR]

Abstract: We here report characterization of a grape (Vitis vinifera) acyl-CoA-binding protein (VvACBP). Expression of VvACBP was detected in grape leaves exposed to tunicamycin-induced endoplasmic reticulum (ER) stress as well as cold and heat shock treatments. In tendrils and peduncles, however, high-temperature treatment induced BiP (luminal binding protein) expression, a marker of ER stress in berry skin, but not VvACBP expression. We hypothesize that VvACBP may be sorted to the periphery of plant cells. Transgenic Arabidopsis plants, expressing VvACBP, exhibited slowed-down floral transition. The gene expression of proteins related to the photoperiodic pathway, CONSTANS, FLOWERING LOCUS T (FT), and SUPPRESSOR OF OVEREXPRESSION OF CONSTANS 1 (SOC1), was down-regulated in transgenic seedlings. These results underscore the possibility that VvACBP may affect the regulation of floral transition in Arabidopsis by suppressing the photoperiodic pathway. The transgenic Arabidopsis plants also exhibited morphological changes such as thicker inflorescences and rosette leaves. In addition, the rosette leaves of the transgenic plants had higher anthocyanin, total phenol, and chlorophyll contents than those of the control plants. Finally, the transgenic plants showed disease resistance to Pseudomonas syringae and Colletotrichum higginsianum, suggesting that VvACBP may also enhance disease resistance in grapevine. [Copyright &y& Elsevier]

Abstract: VIGG is a putative endoplasmic reticulum (ER) resident protein induced by virus infection and ER stress, and is correlated with fruit quality in grapevine. The present study was undertaken to determine the biological function of VIGG in grapevine. Experiments using fluorescent protein-VIGG fusion protein demonstrated that VIGG is localized in ER and the ER targeting sequence is in the N-terminus. The overexpression of VIGG in Arabidopsis plant led to growth retardation. The rosette leaves of VIGG-overexpressing plants were smaller than those of the control plants and rolled at 42days after seeding. VIGG-overexpressing plants revealed robustness to ER stress as well as the low expression of ER stress marker proteins, such as the luminal binding proteins. These characteristics of VIGG-overexpressing plants were supported by a microarray experiment that demonstrated the disruption of genes related to ER stress response and flowering, as well as cation mobility, in the plants. Finally, cation homeostasis in the plants was disturbed by the overexpression of VIGG. Taken together, these results suggest that VIGG may disturb cation homeostasis in plant, which is correlated with the robustness to ER stress and growth retardation. [Copyright &y& Elsevier]

Abstract: We report the cloning and characterization of heat shock-induced genes in grapevine (Vitis vinifera L.). Using the cDNA subtraction method, four heat shock-induced genes were identified in heat shock-treated Pinot noir grapevine. The four genes were immediately induced and upregulated in leaves and berry skins by heat treatment at 45°C. The recovery treatment at 26°C reduced the upregulated transcription of the heat shock-induced genes to near basal levels. The predicted amino acid sequences of three genes, HSG4, HSG14, and HSG19, showed high homologies to those of known small heat shock proteins of other plants. The predicted amino acid sequence of the fourth gene, HSG1, has two conserved motifs, the IQ motif and the Bcl-2-associated athanogene (BAG) domain, suggesting that HSG1 may be a member of the BAG family in grapevine. Although no morphological changes were observed, the overexpression Arabidopsis lines of HSG1, HSG4, HSG14, or HSG19 exhibited faster growth including floral transition than the control line, suggesting that the constitutive expression of HSG1, HSG4, HSG14, or HSG19 protein resulted in increased growth rates without any detectable harm. The thermotolerance test demonstrated that all of the overexpression lines remained viable and noticeably healthy compared with the control line even after exposure to severe heat shock, suggesting that HSG1, HSG4, HSG14, or HSG19 protein might be related to the acquisition of thermotolerance in grapevine. [Copyright &y& Elsevier]

Anthocyanin accumulation in grape berry skin is a crucial determinant of red/black grape berry quality. Recently, viticulture has been hampered by the issue of global warming, which has led to loss of anthocyanin accumulation in berry skin. The objectives of this study were to investigate the effect of allantoin on anthocyanin accumulation in berry skins of field-grown grapevines and to elucidate the molecular mechanism of the allantoin-induced anthocyanin accumulation in the berry skins. Allantoin enhanced anthocyanin accumulation in grape cultured cells and berry skins from field-grown grapevines at the early stage of ripening. MybA1 and UFGT , which encode the key transcription factor and enzyme in anthocyanin biosynthesis, were upregulated in allantoin-treated berry skins. Allantoin seems to increase the contents of delphinidin-based anthocyanin 3-glucosides in berry skins through the upregulation of F3'5'H gene that drives the synthesis of delphinidin-based anthocyanin 3-glucosides, compared with control berry skins. Allantoin increased soluble sugar contents in berries 10 days after treatment and upregulated abscisic acid (ABA)-responsible HT6 gene that encodes a key hexose transporter in sugar accumulation during ripening, in berry skins. These results suggested that physiological changes in allantoin-treated grape berries occurred in an ABA-dependent manner. Allantoin produced bioactive ABA through the β-glucosidase-catalyzed hydrolysis of glucose-conjugated ABA but not through the ABA biosynthesis by NCED, a key enzyme in ABA biosynthesis, in berry skins. Allantoin application in viticulture under global warming conditions is expected to contribute to mitigating loss of red/black berry skin color. [ABSTRACT FROM AUTHOR]