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Study of the molecular mechanisms involved in the beneficial effect of high CO2 levels to preserve table grape quality during the storage at low temperature

Final Report Summary - MOLCHARAVITIS (Study of the molecular mechanisms involved in the beneficial effect of high CO2 levels to preserve table grape quality during the storage at low temperature)

Elucidate the mechanisms by which grapes respond to low non-freezing temperatures, and how CO2 treatment is able to reduce the damage associated to the temperature shift to low temperature is of enormous interest for companies and breeders in order to improve the grape production and competitiveness. It is interesting to mention that according to the Food and Agriculture Organization (FAO), 75.866 km2 of the world are dedicated to grapes. Approximately 71% of world grape production is used for wine, 27% as fresh fruit, and 2% as dried fruit. Spain is the first country in terms of area dedicated to grapes for wine making with 11,750 km2; and it is the fifth country in terms of grape production with 6.1 million tons.
In relation to the postharvest technologies, the use of SO2 is being restricted in most countries due to environmental and health concerns, justifying the search for alternatives such as CO2 treatment. Thus, the host group has been studying the use of pretreatment with high CO2 levels during 3 days at 0ºC since many years ago searching for good alternatives to maintain the quality of different crops like cherimoya, strawberries and table grapes during storage at low temperature.
In order to improve the quality of table grapes during the postharvest period, the overall goal of this work was to obtain a comprehensive understanding of the molecular mechanisms involved in the differences observed in response to low-storage temperature between table grape bunches non-treated and treated with high levels of CO2 during 3 days at 0ºC. Likewise, it has been analyzed whether the CO2 treatment was effective in different table grapes cultivars. A transcriptomic study using the custom-made GrapeGen GeneChip® was developed and results obtained by the host group revealed that the effect of the high CO2 pretreatment maintaining table grapes quality seems to be an active process requiring the activation of transcription factors as well as protein kinases implicate in the regulation of protein function. Up to 20% of the transcription factors from the AP2/EREBP family (APETALA2 and Ethylene-Responsive Element Binding Proteins) which were present in the microarray, showed changes in their expression levels once the CO2 treatment was finished. Among the transcription factors of this family, there have been identified five subfamilies including ERFs (Ethylene-Responsive element-binding Factors) and CBFs (C-repeat Binding Factors) proteins.
As a first result, it has been proven for the first time the effectiveness of the CO2 treatment associated with low temperature for long periods of storage in the preservation of white table grape cultivars. Furthermore, results obtained in this project give clues of the genes related with the improvement observed after the CO2 treatment. Thus, it has been showed the in vitro functional characterization of a recombinant dehydrin class I which showed cryoprotective and slightly antifungal patterns. These findings reinforce the idea that these proteins are appealing to the industry and might be used as an ideal starting point for the creation of novel cryo-resistant enzymes. By contrast, in terms of regulation of dehydrin class I by CBFs in fruits treated with CO2 and stored at low temperatures, it has been shown for the first time that this regulation could be attributed to other different pathways than modulation via CBFs according to the results of gene expression observed in the grape tissues analyzed and the EMSA affinity assays. By other hand, since we have observed that some members of CBFs are able to regulate the DRE motif present in a dehydrin class II, it would be interesting in a short future to functional characterize this member of LEA proteins. However, some family members of ERFs seem to be involved in the modulation of PRs from grapes treated with high concentrations of CO2 and low temperatures.
New interesting vias, opened to the light of these results, would be to investigate new transcription factors according to the importance observed in the transcriptomic array. Moreover, the eventual confirmation of these results through reverse genetics in vivo would be very interesting for the investigation itself and also for the host group, since would implicate some knowledge transferred by Dra. Romero from their previous postdoctoral experience.
This investigation has led to 1 published paper, 3 submitted papers and at least 2 more papers that will be written in a short future. Results have been shown in 2 National Congresses and in an International COST. It is important to point out that the talk about ERFs arisen from the results of this project given in the Postharvest Congress in Valencia was awarded as “best talk” with an economic prize.
Some results derived from this project will be included in the Master Thesis of Andrea Eoli in June 2015.
Finally, during the time of this project Dra. Romero managed to get a permanent position as a Licentiate in the Host group where she will continue working, deeping in some pendent aspects of this proposal.