VvGOLS1 and VvHsfA2 are Involved in the Heat Stress Responses in Grapevine Berries

Among various environmental factors, temperature is a major regulator affecting plant growth, development and fruit composition. Grapevine is the most cultivated fruit plant throughout the world, and grapes are used for wine production and human consumption. The molecular mechanisms involved in grapevine tolerance to high temperature, especially at the fruit level, are poorly understood. To better characterize the sensitivity of berries to the microenvironment, high temperature conditions were locally applied to Vitis vinifera Cabernet Sauvignon clusters. Two genes, VvGOLS1 and VvHsfA2, up-regulated by this treatment, were identified and further characterized. The expression profile of VvGOLS1 correlated positively with galactinol accumulation in heat-stressed berries. However, no galactinol derivatives, such as raffinose and stachyose, accumulated upon heat stress. Heterologous expression of VvGOLS1 in Escherichia coli showed that it encodes a functional galactinol synthase. Transient expression assays showed that the heat stress factor VvHsfA2 transactivates the promoter of VvGOLS1 in a heat stress-dependent manner. Taken together, our results highlight the intrinsic capacity of grape berries to perceive heat stress and to initiate adaptive responses, suggesting that galactinol may play a signaling role in these response

Purification and functional characterization of protoplasts and intact vacuoles from grape cells

<p>Abstract</p> <p>Background</p> <p>During grape berry ripening, the vacuoles accumulate water, sugars and secondary metabolites, causing great impact in plant productivity and wine quality. However, the molecular basis of these compartmentation processes is still poorly understood. As in many species, the major bottleneck to study these aspects in grapevine is to obtain highly purified vacuoles with a good yield. The present paper describes an isolation method of protoplasts and intact vacuoles from grape berry cells and their functional characterization by transport and cytometric assays.</p> <p>Findings</p> <p>Protoplasts were prepared by enzymatic digestion of grape cells, and vacuoles were released and purified by a Ficoll step gradient centrifugation. The tonoplast stained strongly with the fluorescent dye FM1-43 and most vacuoles maintained an internal acidic pH, as assessed by Neutral Red. Flow cytometry analysis of vacuole samples incubated with the calcium-sensitive fluorescent probe Fluo-4 AM revealed a well-defined sub-population of intact vacuoles. As assessed by the pH-sensitive probe ACMA, intact vacuoles generated and maintained a pH gradient through the activity of V-ATPase and V-PPase and were able to transport Ca²⁺via a proton-dependent transport system.</p> <p>Conclusions</p> <p>Highly pure, intact and functional protoplast and vacuole populations from grape cells were obtained with the present method, which revealed to be fast and efficient. The capacity of the vacuole population to sequester protons and accumulate Ca²⁺strongly suggests the intactness and physiological integrity of these extremely fragile organelles. Grapevine protoplasts and vacuoles may be used as models for both basic research and biotechnological approaches, such as proteomics, solute uptake and compartmentation, toxicological assessments and breeding programs.</p

Zebularine, a DNA Methylation Inhibitor, Activates Anthocyanin Accumulation in Grapevine Cells

Through its role in the regulation of gene expression, DNA methylation can participate in the control of specialized metabolite production. We have investigated the link between DNA methylation and anthocyanin accumulation in grapevine using the hypomethylating drug, zebularine and Gamay Teinturier cell suspensions. In this model, zebularine increased anthocyanin accumulation in the light, and induced its production in the dark. To unravel the underlying mechanisms, cell transcriptome, metabolic content, and DNA methylation were analyzed. The up-regulation of stress-related genes, as well as a decrease in cell viability, revealed that zebularine affected cell integrity. Concomitantly, the global DNA methylation level was only slightly decreased in the light and not modified in the dark. However, locus-specific analyses demonstrated a decrease in DNA methylation at a few selected loci, including a CACTA DNA transposon and a small region upstream from the UFGT gene, coding for the UDP glucose:flavonoid-3-O-glucosyltransferase, known to be critical for anthocyanin biosynthesis. Moreover, this decrease was correlated with an increase in UFGT expression and in anthocyanin content. In conclusion, our data suggest that UFGT expression could be regulated through DNA methylation in Gamay Teinturier, although the functional link between changes in DNA methylation and UFGT transcription still needs to be demonstrated

Proteomic analysis of early-stage incompatible and compatible interactions between grapevine and P. viticola

Wild grapevines can show strong resistance to the downy mildew pathogen P. viticola, but the associated mechanisms are poorly described, especially at early stages of infection. Here, we performed comparative proteomic analyses of grapevine leaves from the resistant genotype V. davidii « LiuBa-8 » (LB) and susceptible V. vinifera « « Pinot Noir� (PN) 12 h after inoculation with P. viticola. By employing the iTRAQ technique, a total of 444 and 349 differentially expressed proteins (DEPs) were identified in LB and PN, respectively. The majority of these DEPs were related to photosynthesis, respiration, cell wall modification, protein metabolism, stress, and redox homeostasis. Compared with PN, LB showed fewer downregulated proteins associated with photosynthesis and more upregulated proteins associated with metabolism. At least a subset of PR proteins (PR10.2 and PR10.3) was upregulated upon inoculation in both genotypes, whereas HSP (HSP70.2 and HSP90.6) and cell wall-related XTH and BXL1 proteins were specifically upregulated in LB and PN, respectively. In the incompatible interaction, ROS signaling was evident by the accumulation of H2O2, and multiple APX and GST proteins were upregulated. These DEPs may play crucial roles in the grapevine response to downy mildew. Our results provide new insights into molecular events associated with downy mildew resistance in grapevine, which may be exploited to develop novel protection strategies against this disease

Grapevine responses to high temperature - towards adaptation to climate change

Invited presentation. Conference on Grapevine Breeding and Genetics, 2019 Sept. 26-28, Nanjing, China

Phosphoproteins Involved in the Signal Transduction of Cryptogein, an Elicitor of Defense Reactions in Tobacco

International audienc

A Sugar-Inducible Protein Kinase, VvSK1, Regulates Hexose Transport and Sugar Accumulation in Grapevine Cells

In grapevine (Vitis vinifera), as in many crops, soluble sugar content is a major component of yield and economical value. This paper identifies and characterizes a Glycogen Synthase Kinase3 protein kinase, cloned from a cDNA library of grape Cabernet Sauvignon berries harvested at the ripening stage. This gene, called VvSK1, was mainly expressed in flowers, berries, and roots. In the berries, it was strongly expressed at postvéraison, when the berries accumulate glucose, fructose, and abscisic acid. In grapevine cell suspensions, VvSK1 transcript abundance is increased by sugars and abscisic acid. In transgenic grapevine cells overexpressing VvSK1, the expression of four monosaccharide transporters (VvHT3, VvHT4, VvHT5, and VvHT6) was up-regulated, the rate of glucose uptake was increased 3- to 5-fold, and the amount of glucose and sucrose accumulation was more than doubled, while the starch amount was not affected. This work provides, to our knowledge, the first example of the control of sugar uptake and accumulation by a sugar-inducible protein kinase

Penser et adapter la vigne pour demain

Vidéo de ‘2:30L'Umr Ecophysiologie et génomique fonctionnelle de la vigne cherche à identifier des réseaux de gènes et des modes de régulation permettant de contrôler la balance sucre-acide dans les baies de raisin à maturité. Ces travaux sont menés dans un contexte de changement climatique

The state-of-the-art of grapevine biotechnology and new breeding technologies (NBTS)

Context of the review: The manipulation of the genetic basis controlling grapevine adaptation and phenotypic plasticity can be performed either by classical genetics or biotechnologies. In the last 15 years, considerable knowledge has accumulated about t