A Deep Proteomics Perspective Into Grape Berry Quality Traits During Ripening

Discovery‐based proteomics studies have an important role in the understanding of the biochemical processes that occur during grape berry ripening. The ripening process is relevant in determining grape berry quality. For a proteome analysis of grape berry ripening, Kambiranda et al. (2018) applied a label‐free mass spectrometry–based quantitative approach. The authors reported the identification of proteins associated with the production flavor, aroma and ethylene production. Despite the valuable contribution of discovery‐based proteomics studies, the picture is still incomplete. Future efforts in gaining proteome coverage would benefit the identification of proteins associated with grape berry quality traits.     

Cloning of 9-cis-epoxycarotenoid dioxygenase (NCED) gene and the role of ABA on fruit ripening

In order to understand more details about the role of abscisic acid (ABA) in fruit ripening and senescence, six 740 bp cDNAs (LeNCED1, LeNCED2, PpNCED1, VVNCED1, DKNCED1 and CMNCED1) which encode 9-cis-epoxycarotenoid dioxygenase (NCED) as a key enzyme in ABA biosynthesis, were cloned from fruits of tomato, peach, grape, persimmon and melon using an RT-PCR approach. A Blast homology search revealed a similarity of amino acid 85.76% between the NCEDs. A relationship between ABA and ethylene during ripening was also investigated. At the mature green stage, exogenous ABA treatment increased ABA content in flesh, and promoting ethylene synthesis and fruit ripening, while treatment with nordihydroguaiaretic acid (NDGA), inhibited them, delayed fruit ripening and softening. However, ABA inhibited the ethylene synthesis obviously while NDGA promoted them when treated the immature fruit with these chemicals. At the breaker, NDGA treatment cannot block ABA accumulation and ethylene synthesis. Based on the results obtained in this study, it was concluded that ABA plays different role in ethylene synthesis system in different stages of tomato fruit ripening.Key words: tomato, NCED gene, ABA, ethylene, fruit ripening, peach, grape, persimmon, melon     

Berry and phenology-related traits in grapevine (<Emphasis Type="Italic">Vitis vinifera</Emphasis>L.): From Quantitative Trait Loci to underlying genes

Background  

The timing of grape ripening initiation, length of maturation period, berry size and seed content are target traits in viticulture. The availability of early and late ripening varieties is desirable for staggering harvest along growing season, expanding production towards periods when the fruit gets a higher value in the market and ensuring an optimal plant adaptation to climatic and geographic conditions. Berry size determines grape productivity; seedlessness is especially demanded in the table grape market and is negatively correlated to fruit size. These traits result from complex developmental processes modified by genetic, physiological and environmental factors. In order to elucidate their genetic determinism we carried out a quantitative analysis in a 163 individuals-F₁ segregating progeny obtained by crossing two table grape cultivars.     

Proteomic analysis of grape berry cell cultures reveals that developmentally regulated ripening related processes can be studied using cultured cells

Background

This work describes a proteomics profiling method, optimized and applied to berry cell suspensions to evaluate organ-specific cultures as a platform to study grape berry ripening. Variations in berry ripening within a cluster(s) on a vine and in a vineyard are a major impediment towards complete understanding of the functional processes that control ripening, specifically when a characterized and homogenous sample is required. Berry cell suspensions could overcome some of these problems, but their suitability as a model system for berry development and ripening needs to be established first.

Methodology/Principal Findings

In this study we report on the proteomic evaluation of the cytosolic proteins obtained from synchronized cell suspension cultures that were established from callus lines originating from green, véraison and ripe Vitis vinifera berry explants. The proteins were separated using liquid phase IEF in a Microrotofor cell and SDS PAGE. This method proved superior to gel-based 2DE. Principal component analysis confirmed that biological and technical repeats grouped tightly and importantly, showed that the proteomes of berry cultures originating from the different growth/ripening stages were distinct. A total of twenty six common bands were selected after band matching between different growth stages and twenty two of these bands were positively identified. Thirty two % of the identified proteins are currently annotated as hypothetical. The differential expression profile of the identified proteins, when compared with published literature on grape berry ripening, suggested common trends in terms of relative abundance in the different developmental stages between real berries and cell suspensions.

Conclusions

The advantages of having suspension cultures that accurately mimic specific developmental stages are profound and could significantly contribute to the study of the intricate regulatory and signaling networks responsible for berry development and ripening.     

Role of ABA and Gibberellin A<Subscript>3</Subscript> on gene expression pattern of sugar transporters and invertases in <Emphasis Type="Italic">Vitis vinifera</Emphasis> cv. Malbec during berry ripening

Sugars are key constituents that affect quality of grape berries, and consequently the grape metabolic profile relevant to wine’s industry. However, enzymes and transporter genes expression involved in sugar transport at different phenological stages are scarcely studied. In addition, little is known about the role of the plant hormones ABA and Gibberellin (GA₃) as endogenous regulators, over the expression pattern of the sugars transporters genes in grapevine. The aim of this study was to analyze the expression pattern of the most relevant sugar transporters and invertases in leaves and berries of grapevine plants cv. Malbec during berry ripening stages and its shift after ABA and GA₃ sprays. In leaves, VvHT1 was the sugar transporter highly expressed, whereas VvHT6 was the most abundant in berries throughout berry ripening. Moreover, VvSUC12 and VvSUC27 were expressed at veraison greater in leaves than in berries, suggesting an active phloem loading at the onset of ripening. Applications of ABA and GA₃ enhanced the expression of VvSUC12 and VvSUC27 in pre-veraison leaves. Furthermore, hormones increased the expression of VvHT2, VvHT3 and VvHT6 in berries at different stages of ripening favoring sugar unloading from phloem. In conclusion, ABA and GA₃ are involved in the long-distance sugar transport from leaves to berries in Vitis vinifera L. cv. Malbec, and their exogenous application could be a suitable strategy to improve the process.     

Effects of ethylene and 2-chloroethylphosphonic Acid on the ripening of grapes

The effects of ethylene gas, 2-chloroethylphosphonic acid, and the auxin, benzothiazole-2-oxyacetic acid, on the ripening of grapes (Vitis vinifera L.) was investigated. Ethylene hastened the start of ripening of Doradillo grapes when it was aplied for 10 days starting midway through the slow growth phase. 2-Chloroethylphosphonic acid applied to Shiraz grapes showed the same effect, but when it was applied earlier, during the second half of the first rapid growth phase or at the start of the slow growth phase of berry development, it delayed ripening. 2-Chloroethylphosphonic acid and benzothiazole-2-oxyacetic acid delayed the ripening of Doradillo grapes, and ethylene partially reversed the effect of benzothiazole-2-oxyacetic acid. The results demonstrate the importance of the slow growth stage in grape berry development and suggest that an auxin-ethylene relationship may be involved in the regulation of grape ripening.     

ABA Initiates Anthocyanin Production in Grape Cell Cultures

Abscisic acid (ABA) has a well-known positive impact on grape ripening, especially color development, but its role in the initiation of anthocyanin synthesis remains unclear. To elucidate this point, ABA treatment was applied to a simple Vitis vinifera model, consisting of Cabernet Sauvignon cell suspensions that do not spontaneously produce anthocyanins under laboratory conditions. Endogenous ABA levels, the expression of some genes in the upstream part of the anthocyanin pathway, and anthocyanin content were determined. Exogenous ABA treatment sharply increased cell ABA content and induced both structural and regulatory genes involved in anthocyanin production. These changes were promptly detected, as early as 6 h after ABA treatment, whereas anthocyanin production was observed only after 4 days in culture. These results demonstrate that ABA promotes anthocyanin synthesis in grape cell culture.     

Transformation of a marker-free and vector-free antisense ACC oxidase gene cassette into melon via the pollen-tube pathway

Purpose of work  

Melons have short shelf-lives due to fruit ripening caused by ethylene production. The 1-aminocyclopropane-1-carboxylic acid (ACC) oxidase gene is essential for ethylene biosynthesis. As fruit ripening in other fruit crops can be deterred by down-regulation of ACC oxidase expression, we have carried out similar work to improve fruit quality and shelf-life of the melon Cucumis melo.     

Effect of Abscisic Acid on Banana Fruit Ripening in Relation to the Role of Ethylene

Abstract The role of abscisic acid (ABA) in banana fruit ripening was examined with the ethylene binding inhibitor, 1-methylcyclopropene (1-MCP). ABA (0, 10⁻⁵, 10⁻⁴, or 10⁻³ mol/L) was applied by vacuum infiltration into fruit. 1-MCP (1 μL/L) was applied by injecting a measured volume of stock gas into sealed glass jars containing fruit. Fruit ripening, as judged by ethylene evolution and respiration associated with color change and softening, was accelerated by 10⁻⁴ or 10⁻³ mol/L ABA. ABA at 10⁻⁵ mol/L had no effect. The acceleration of ripening by ABA was greater at 10⁻³ mol/L than at 10⁻⁴ mol/L. ABA-induced acceleration of banana fruit ripening was not observed in 1-MCP treated fruit, especially when ABA was applied after exposure to 1-MCP. Thus, ABA's promotion of ripening in intact banana fruit is at least partially mediated by ethylene. Exposure of ABA-treated fruit to 0.1 μL/L ethylene for 24 h resulted in increased ethylene production and respiration, and associated skin color change and fruit softening. Control fruit (no ABA) was unresponsive to similar ethylene treatments. The data suggest that ABA facilitates initiation and progress in the sequence of ethylene-mediated ripening events, possibly by enhancing the sensitivity to ethylene. Received 29 January 1999; accepted 16 January 2000     

Functional Analysis of <Emphasis Type="Italic">VvBG1</Emphasis> During Fruit Development and Ripening of Grape

β-glucosidase (BG) was believed to take part in abscisic acid (ABA) synthesis via hydrolysis of ABA glucose ester to release active ABA during plant growth and development. However, there is no genetic evidence available to indicate the role of genes during fruit ripening. Here, the expression patterns of three genes (VvBG1, VvBG2, and VvBG3) encoding β-glucosidase were analyzed during grape fruit development, and it was found that β-glucosidase activity increased in grape fruit in response to various stresses. Furthermore, to verify the function of β-glucosidase during fruit ripening, heterogeneous expression of the VvBG1 gene in strawberry fruit was validated, and the results showed that the VvBG1 over-expression increased β-glucosidase and promoted the fruit ripening process in strawberry. In addition, we found that ABA contents increased in the VvBG1 over-expression of strawberry fruit, which induced fruit anthocyanin, soluble solid accumulation, and fruit softening. Moreover, genes related to coloring (CHS, CHI, F3H, and UFGT), softening (PG1, PL1, and EXP1), and aroma (SAAT, and QR) were up-regulated. This work will elucidate the specific roles of VvBGs in the synthesis of ABA and provide some new insights into the ABA-controlled grape ripening mechanism.     

Abscisic Acid Sprays Significantly Increase Yield per Plant in Vineyard-Grown Wine Grape (<Emphasis Type="Italic">Vitis vinifera</Emphasis> L.) cv. Cabernet Sauvignon Through Increased Berry Set with No Negative Effects on Anthocyanin Content and Total Polyphenol Index of Both Juice and Wine

In many cultivars of Vitis vinifera periods of mild water stress during ripening are thought to increase grape quality for winemaking, even though yields may be negatively affected. Because abscisic acid (ABA) is involved in the signaling of water stress in plants, we examine the effects of the ABA signal being given without the concomitant water stress. ABA at 250 mg l⁻¹ was sprayed weekly or biweekly from bud-burst until harvest onto the leaves of vineyard-grown plants of cv. Cabernet Sauvignon. For ABA-treated plants berry yield per bunch and per plant was significantly increased (1.5- to 2.0-fold) across three consecutive harvests (2005 through 2007). Number of berries per bunch and per plant was the primary basis for the significant crop increases, although bunches per plant also tended to increase (1.1- to 1.3-fold) across all three harvests. Other parameters assessed included number of internodes, shoot length, leaf area, leaf water potential at midday, photosynthesis, and stomatal conductance. These parameters showed no significant change with ABA treatment, although shoot length tended to be reduced, as was leaf area relative to control plants. The significantly increased fruit yields were thus accomplished without accompanying increases in leaf photosynthesis and leaf areas. Juice at harvest had equal levels of sugars (Brix) and somewhat higher levels of anthocyanins and total polyphenols relative to control values. The two latter trends continued for the resultant wine across two vintage years. In conclusion, three seasons of experimental trials have demonstrated that ABA application can significantly enhance yield per plant in the field-grown grape (cv. Cabernet Sauvignon) by favoring increased berry set without diminishing the quality of the fruit for winemaking use.     

Pectic-β(1,4)-galactan,extensin and arabinogalactan–protein epitopes differentiate ripening stages in wine and table grape cell walls

Background and Aims

Cell wall changes in ripening grapes (Vitis vinifera) have been shown to involve re-modelling of pectin, xyloglucan and cellulose networks. Newer experimental techniques, such as molecular probes specific for cell wall epitopes, have yet to be extensively used in grape studies. Limited general information is available on the cell wall properties that contribute to texture differences between wine and table grapes. This study evaluates whether profiling tools can detect cell wall changes in ripening grapes from commercial vineyards.

Methods

Standard sugar analysis and infra-red spectroscopy were used to examine the ripening stages (green, véraison and ripe) in grapes collected from Cabernet Sauvignon and Crimson Seedless vineyards. Comprehensive microarray polymer profiling (CoMPP) analysis was performed on cyclohexanediaminetetraacetic acid (CDTA) and NaOH extracts of alcohol-insoluble residue sourced from each stage using sets of cell wall probes (mAbs and CBMs), and the datasets were analysed using multivariate software.

Key Results

The datasets obtained confirmed previous studies on cell wall changes known to occur during grape ripening. Probes for homogalacturonan (e.g. LM19) were enriched in the CDTA fractions of Crimson Seedless relative to Cabernet Sauvignon grapes. Probes for pectic-β-(1,4)-galactan (mAb LM5), extensin (mAb LM1) and arabinogalactan proteins (AGPs, mAb LM2) were strongly correlated with ripening. From green stage to véraison, a progressive reduction in pectic-β-(1,4)-galactan epitopes, present in both pectin-rich (CDTA) and hemicellulose-rich (NaOH) polymers, was observed. Ripening changes in AGP and extensin epitope abundance also were found during and after véraison.

Conclusions

Combinations of cell wall probes are able to define distinct ripening phases in grapes. Pectic-β-(1,4)-galactan epitopes decreased in abundance from green stage to véraison berries. From véraison there was an increase in abundance of significant extensin and AGP epitopes, which correlates with cell expansion events. This study provides new ripening biomarkers and changes that can be placed in the context of grape berry development.