Secondary metabolism contributes to the adaptation of a plant to its environment. In wine grapes, fruit secondary metabolism largely determines wine quality. Climate change is predicted to exacerbate drought events in several viticultural areas, potentially affecting the wine quality. In red grapes, water deficit modulates flavonoid accumulation, leading to major quantitative and compositional changes in the profile of the anthocyanin pigments; in white grapes, the effect of water deficit on secondary metabolism is still largely unknown. In this study we investigated the impact of water deficit on the secondary metabolism of white grapes using a large scale metabolite and transcript profiling approach in a season characterized by prolonged drought. Irrigated grapevines were compared to non-irrigated grapevines that suffered from water deficit from early stages of berry development to harvest. A large effect of water deficit on fruit secondary metabolism was observed. Increased concentrations of phenylpropanoids, monoterpenes, and tocopherols were detected, while carotenoid and flavonoid accumulations were differentially modulated by water deficit according to the berry developmental stage. The RNA-sequencing analysis carried out on berries collected at three developmental stages—before, at the onset, and at late ripening—indicated that water deficit affected the expression of 4,889 genes. The Gene Ontology category secondary metabolic process was overrepresented within up-regulated genes at all the stages of fruit development considered, and within down-regulated genes before ripening. Eighteen phenylpropanoid, 16 flavonoid, 9 carotenoid, and 16 terpenoid structural genes were modulated by water deficit, indicating the transcriptional regulation of these metabolic pathways in fruit exposed to water deficit. An integrated network and promoter analyses identified a transcriptional regulatory module that encompasses terpenoid genes, transcription factors, and enriched drought-responsive elements in the promoter regions of those genes as part of the grapes response to drought. Our study reveals that grapevine berries respond to drought by modulating several secondary metabolic pathways, and particularly, by stimulating the production of phenylpropanoids, the carotenoid zeaxanthin, and of volatile organic compounds such as monoterpenes, with potential effects on grape and wine antioxidant potential, composition, and sensory features.

Grapes are one of the major fruit crops and they are cultivated in many dry environments. This study comprehensively characterizes the metabolic response of grape berries exposed to water deficit at different developmental stages. Increases of proline, branched-chain amino acids, phenylpropanoids, anthocyanins, and free volatile organic compounds have been previously observed in grape berries exposed to water deficit. Integrating RNA-sequencing analysis of the transcriptome with large-scale analysis of central and specialized metabolites, we reveal that these increases occur via a coordinated regulation of key structural pathway genes. Water deficit-induced up-regulation of flavonoid genes is also coordinated with the down-regulation of many stilbene synthases and a consistent decrease in stilbenoid concentration. Water deficit activated both ABA-dependent and ABA-independent signal transduction pathways by modulating the expression of several transcription factors. Gene-gene and gene-metabolite network analyses showed that water deficit-responsive transcription factors such as bZIPs, AP2/ERFs, MYBs, and NACs are implicated in the regulation of stress-responsive metabolites. Enrichment of known and novel cis-regulatory elements in the promoters of several ripening-specific/water deficit-induced modules further affirms the involvement of a transcription factor cross-talk in the berry response to water deficit. Together, our integrated approaches show that water deficit-regulated gene modules are strongly linked to key fruit-quality metabolites and multiple signal transduction pathways may be critical to achieve a balance between the regulation of the stress-response and the berry ripening program. This study constitutes an invaluable resource for future discoveries and comparative studies, in grapes and other fruits, centered on reproductive tissue metabolism under abiotic stress.

Background: Most approaches to grape physiology accept that the berry and the future harvest should display identical developmental features, which obviously requires synchronised fruits.Aims: Rejecting this assumption compels to revisit the kinetic and metabolic bases of berry ripening.Methods and Results: Two to three thousand berries were individually analysed for sugar, malate and weight. The huge heterogeneity in sugar and malic acid concentrations among fruits was mostly explained by time lags in the onset of sugar storage, which proved nearly as long as the second growth phase. Individual berries from different cultivars displayed similar kinetics following the normalisation of their maximal volume. Phloem sucrose unloading started at its maximum speed at softening, but growth resumed one week later. Four hexoses accumulated per malic acid, which was oxidised during the first two weeks of ripening, and then malate breakdown stopped without affecting sugar accumulation. Sugar and water accumulation were simultaneously arrested four weeks after softening, at 0.9 M hexose, at which point sugar concentration continued through water losses.Conclusions: The accepted sequential random sampling methods representative of average fruit and future wine compositions have led to a scrambled vision of grape developmental biology, presenting serious kinetic and composition biases. Single berry composition provides first quantitative evidence for the induction of a dominant H+/sucrose exchange on the tonoplast, which is first electro-neutralised by malate breakdown, then by ATP demanding H+ recirculation, in line with functional and molecular studies.Significance of the Study: The kinetics of single berry ripening are presented for the first time. A more reliable and reproducible model of berry growth, sugar import and malate breakdown is shown here, which have definitively been improved from a quantitative point of view. It illustrates that the temporal structure of a berry population may largely contribute to future wine quality, in addition to metabolic plasticity, thereby providing another target for the impact of GxE interaction. In this respect, addressing the structure of berry cohorts may provide a new approach regarding the developmental biology/terroir nexus.

Variegation is a rare type of mosaicism not fully studied in plants, especially fruits. We examined red and white sections of grape (Vitis vinifera cv. ‘Béquignol’) variegated berries and found that accumulation of products from branches of the phenylpropanoid and isoprenoid pathways showed an opposite tendency. Light-responsive flavonol and monoterpene levels increased in anthocyanin-depleted areas in correlation with increasing MYB24 expression. Cistrome analysis suggested that MYB24 binds to the promoters of 22 terpene synthase (TPS) genes, as well as 32 photosynthesis/light-related genes, including carotenoid pathway members, the flavonol-regulator HY5 HOMOLOGUE (HYH), and other radiation response genes. Indeed, TPS35, TPS09, the carotenoid isomerase gene CRTISO2 and HYH were activated in the presence of MYB24 and MYC2. We suggest that MYB24 modulates ultraviolet and high-intensity visible light stress responses that include terpene and flavonol synthesis and potentially affect carotenoids. The MYB24 regulatory network is developmentally triggered after the onset of berry ripening, while the absence of anthocyanin sunscreens accelerates its activation, likely in a dose-dependent manner due to increased radiation exposure. Anthocyanins and flavonols in variegated berry skins act as effective sunscreens but for different wavelength ranges. The expression patterns of stress marker genes in red and white sections of ‘Béquignol’ berries strongly suggest that MYB24 promotes light-stress amelioration but only partly succeeds during late ripening.

Transcriptomic changes at the cessation of sugar accumulation in the pericarp ofVitis viniferawere addressed on single berries re-synchronised according to their individual growth patterns. The net rates of water, sugars and K⁺accumulation inferred from individual growth and solute concentration confirmed that these inflows stopped simultaneously in the ripe berry, while the small amount of malic acid remaining at this stage was still being oxidised at low rate. Re-synchronised individual berries displayed negligible variations in gene expression among triplicates. RNA-seq studies revealed sharp reprogramming of cell-wall enzymes and structural proteins at the stop of phloem unloading, associated with an 80% repression of multiple sugar transporters and aquaporins on the plasma or tonoplast membranes, with the noticeable exception of H⁺/sugar symporters, which were rather weakly and constitutively expressed. This was verified in three genotypes placed in contrasted thermo-hydric conditions. The prevalence of SWEET suggests that electrogenic transporters would play a minor role on the plasma membranes of SE/CC complex and the one of the flesh, while sucrose/H⁺exchangers dominate on its tonoplast.Cis-regulatory elements present in their promoters allowed to sort these transporters in different groups, also including specific TIPs and PIPs paralogs, and cohorts of cell wall-related genes. Together with simple thermodynamic considerations, these results lead to propose that H⁺/sugar exchangers at the tonoplast, associated with a considerably acidic vacuolar pH, may exhaust cytosolic sugars in the flesh and alleviate the need for supplementary energisation of sugar transport at the plasma membrane.