Selenium Biofortification Impacts the Tomato Fruit Metabolome and Transcriptional Profile at Ripening.

In the present work, the effects of enriching tomatoes with selenium were studied in terms of physiological, metabolic, and molecular processes in the last stages of fruit development, particularly during ripening. A selenium concentration of 10 mg L-1 with sodium selenate and selenium nanoparticles was used in the spray treatments on the whole plants. No significant effects of selenium enrichment were detected in terms of ethylene production or color changes in the ripening fruit. However, selenium enrichment had an influence on both the primary and secondary metabolic processes and thus the biochemical composition of ripe tomatoes. Selenium decreased the amount of ß-carotene, increased the accumulation of naringenin and chlorogenic acid, and decreased the coumaric acid level. Selenium also affected the volatile organic compound profile, with changes in the level of specific apocarotenoid compounds, such as ß-ionone. These metabolomic changes may, to some extent, be due to the impact of selenium treatment on the transcription of genes involved in the metabolism of these compounds. RNA-seq analysis showed that the selenium application mostly impacted the expression of the genes involved in hormonal signaling, secondary metabolism, flavonoid biosynthesis, and glycosaminoglycan degradation.

Reducing crop losses by gene-editing control of organ developmental physiology.

Some physiological processes in reproductive organs, if not controlled, can lead to crop loss even in the absence of environmental stress. These processes may occur pre- or post- harvest, and in diverse species and include abscission processes in cereal grain, e.g., shattering and in immature fruit, e.g., preharvest drop, preharvest sprouting of cereals, and postharvest senescence in fruit. Some of the molecular mechanisms and genetic determinants underlying these processes are now better detailed, making it possible to refine them by gene editing. Here, we discuss using advanced genomics to identify genetic determinants underlying crop physiological traits. Examples of improved phenotypes developed for preharvest problems are provided, and suggestions for reducing postharvest fruit losses by gene and promoter editing were made.

Ozone-Induced Biochemical and Molecular Changes in Vitis vinifera Leaves and Responses to Botrytis cinerea Infections.

To investigate how plants cope with multi-stress conditions, we analyzed the biochemical and molecular changes of Vitis vinifera leaves subjected to single or sequential double stresses (infection by Botrytis cinerea (Bc) and ozone (O3, 100 ppb for 3 h) treatment). In Bc+/O3- leaves, the hydrogen peroxide (H2O2) induction (observed at 12 and 24 h from the end of treatment (FET)) triggered a production of ethylene (Et; +35% compared with Bc-/O3- leaves), which was preceded by an increase of salicylic acid (SA; +45%). This result confirms a crosstalk between SA- and Et-related signaling pathways in lesion spread. The ozone induced an early synthesis of Et followed by jasmonic acid (JA) and SA production (about 2-fold higher), where Et and SA signaling triggered reactive oxygen species production by establishing a feedback loop, and JA attenuated this cycle by reducing Et biosynthesis. In Bc+ + O3+ leaves, Et peaked at 6 and 12 h FET, before SA confirmed a crosstalk between Et- and SA-related signaling pathways in lesion propagation. In O3+ + Bc+ leaves, the H2O2 induction triggered an accumulation of JA and Et, demonstrating a synergistic action in the regulation of defence reactions. The divergence in these profiles suggests a rather complex network of events in the transcriptional regulation of genes involved in the systemic acquired resistance.

Short-Term CO2 Treatment of Harvested Grapes (Vitis vinifera L., cv. Trebbiano) before Partial Dehydration Affects Berry Secondary Metabolism and the Aromatic Profile of the Resulting Wine.

High CO2 concentrations applied to harvested horticultural products can modify primary and secondary metabolism. This work reports the metabolic responses to short-term CO2 treatments of white-skinned grapes (cv Trebbiano) undergoing postharvest partial dehydration. The influence of CO2 treatments on the aroma profile of the derived sweet wine was also assessed. Harvested grapes were treated with gaseous CO2 (30%) or air (control) for 24 h and then dehydrated (about 45% of weight loss) before vinification. Lipophilic and phenolic compounds of grape skin and the wine aroma profile were analyzed. In CO2-treated berries, the lipophilic and phenolic compounds decreased at a reduced and faster rate, respectively, during dehydration. Aroma profile of wine from CO2-treated grapes showed a slight but significantly higher content of glycosylated C13 and terpene compounds, and a decrease/absence of free acids, vanillin derivates and other phenol volatiles. The higher content of volatile alcohols in wine from treated berries suggests that the alcoholic fermentation was triggered. CO2 application before the withering process of Trebbiano grapes affects the aroma profile of the resulting wine by altering the free:glycosylated volatiles ratio. This study provides information on the possible use of CO2 as metabolic elicitor to modulate the aroma profile of the resulting wines obtained after grape dehydration.

Pre-processing Cooling of Harvested Grapes Induces Changes in Berry Composition and Metabolism, and Affects Quality and Aroma Traits of the Resulting Wine.

Due to the greenhouse gas increase, grapes are often exposed to high temperatures in several growing areas especially during the final developmental stages, and this is particularly true when early ripening cultivars are harvested. This may cause undesirable effects on berry metabolism and composition and wine quality, particularly concerning the aroma profile. Harvesting at night or keeping the harvested grapes in cold rooms before vinification are empirical protocols applied in specific viticultural areas. To study the effects of decreasing berry temperature after harvest, white-skinned berries (cv Vermentino) were maintained at 4 or 10°C for 24 or 48 h before processing (pre-cooling). Control grapes were kept at 22°C. Grapes cooled at 10°C for 24 and 48 h resulted richer in polyphenols and showed a significant up-regulation of genes involved in polyphenols biosynthesis (i.e., VvPAL, VvSTS2, and VvFLS1). Similar behavior was observed in samples kept at 4°C for 48 h. Pre-cooling induced specific changes in the volatile organic compound (VOC) profiles. In particular, higher amounts of a specific subcategory of terpenes, namely sesquiterpenes, were detected in cooled samples. The induction of the expression of key genes involved in terpenoids biosynthesis (VvHDR, VvDX3, VvTER, VvGT14) was detected in cooled grapes, with variable effects depending on temperature and treatment duration. In both cooled samples, the evolution of alcoholic fermentation followed a regular trend but ended earlier. Higher phenolic content was detected in wines obtained from the 10°C-treated grapes. Higher residual concentration of malic acid at the end of fermentation was detected in wine samples from grapes pre-cooled at 4°C. Sesquiterpenes also showed a general increase in wines from cooled grapes, especially after pre-cooling at 10°C for 48 h. Different sensory profiles characterized the wine samples, with the best scores in terms of general pleasantness obtained by the wine produced from grapes pre-cooled at 4°C for 24 h. These results demonstrate that pre-cooling harvested grapes induces specific effect on the VOC profile and other quality parameters of Vermentino wine, and this appears to be the result of specific metabolic and compositional changes occurring in the berries.

Potential Mitigation of Smoke Taint in Wines by Post-Harvest Ozone Treatment of Grapes.

When bushfires occur near grape growing regions, vineyards can be exposed to smoke, and depending on the timing and duration of grapevine smoke exposure, fruit can become tainted. Smoke-derived volatile compounds, including volatile phenols, can impart unpleasant smoky, ashy characters to wines made from smoke-affected grapes, leading to substantial revenue losses where wines are perceivably tainted. This study investigated the potential for post-harvest ozone treatment of smoke-affected grapes to mitigate the intensity of smoke taint in wine. Merlot grapevines were exposed to smoke at ~7 days post-veraison and at harvest grapes were treated with 1 or 3 ppm of gaseous ozone (for 24 or 12 h, respectively), prior to winemaking. The concentrations of smoke taint marker compounds (i.e., free and glycosylated volatile phenols) were measured in grapes and wines to determine to what extent ozonation could mitigate the effects of grapevine exposure to smoke. The 24 h 1 ppm ozone treatment not only gave significantly lower volatile phenol and volatile phenol glycoside concentrations but also diminished the sensory perception of smoke taint in wine. Post-harvest smoke and ozone treatment of grapes suggests that ozone works more effectively when smoke-derived volatile phenols are in their free (aglycone) form, rather than glycosylated forms. Nevertheless, the collective results demonstrate the efficacy of post-harvest ozone treatment as a strategy for mitigation of smoke taint in wine.

Changes in volatile organic composition of olive oil extracted from cv. 'Leccino' fruit subjected to ethylene treatments at different ripening stages.

BACKGROUND: Olive, as a non-climacteric fruit, is presumed to be ethylene independent with regard to ripening triggering/coordination. Nevertheless, studies have demonstrated that postharvest ethylene treatments induce changes in composition and properties also of non-climacteric fruits, including aroma profiles, a key quality parameter of extra virgin olive oils. Olive fruit of cv. 'Leccino' harvested at two distinct ripening stages (less advanced ripening, LAR; and more advanced ripening, MAR, with Jaén index of 4.58 and 5.10, respectively) were subjected to ethylene (1000 ppm in air) treatment for 24 h before oil extraction. RESULTS: Based on multivariate analysis of volatile organic compound (VOCs), the effect of ethylene treatment appeared to be more pronounced in MAR samples. However, differences in organoleptic analysis were also detected in ethylene-treated LAR olive oils. Ethylene seems to selectively affect linolenic/linoleic acid metabolism, particularly concerning the C5 pathway, and reduce specific defect-associated compounds. CONCLUSION: Exogenous ethylene applied to cv. 'Leccino' olives before processing was effective in inducing specific changes in the VOC profiles of the resulting oil. The effect was different depending on the ripening stage of the harvested olives. The lipoxygenase pathway (including the production of C5 compounds) and fermentative-related compounds appeared to be affected by the treatment. © 2020 Society of Chemical Industry.

Primary Metabolism in Fresh Fruits During Storage.

The extension of commercial life and the reduction of postharvest losses of perishable fruits is mainly based on storage at low temperatures alone or in combination with modified atmospheres (MAs) and controlled atmospheres (CAs), directed primarily at reducing their overall metabolism thus delaying ripening and senescence. Fruits react to postharvest conditions with desirable changes if appropriate protocols are applied, but otherwise can develop negative and unacceptable traits due to the onset of physiological disorders. Extended cold storage periods and/or inappropriate temperatures can result in development of chilling injuries (CIs). The etiology, incidence, and severity of such symptoms vary even within cultivars of the same species, indicating the genotype significance. Carbohydrates and amino acids have protective/regulating roles in CI development. MA/CA storage protocols involve storage under hypoxic conditions and high carbon dioxide concentrations that can maximize quality over extended storage periods but are also affected by the cultivar, exposure time, and storage temperatures. Pyruvate metabolism is highly reactive to changes in oxygen concentration and is greatly affected by the shift from aerobic to anaerobic metabolism. Ethylene-induced changes in fruits can also have deleterious effects under cold storage and MA/CA conditions, affecting susceptibility to chilling and carbon dioxide injuries. The availability of the inhibitor of ethylene perception 1-methylcyclopropene (1-MCP) has not only resulted in development of a new technology but has also been used to increase understanding of the role of ethylene in ripening of both non-climacteric and climacteric fruits. Temperature, MA/CA, and 1-MCP alter fruit physiology and biochemistry, resulting in compositional changes in carbon- and nitrogen-related metabolisms and compounds. Successful application of these storage technologies to fruits must consider their effects on the metabolism of carbohydrates, organic acids, amino acids and lipids.

The inner temperature of the olives (cv. Leccino) before processing affects the volatile profile and the composition of the oil.

The effects of pre-processing decreasing temperature (19, 15 and 10 °C) of olive fruit (cv. Leccino) harvested at three developmental stages (semi-ripe, ripe, advanced ripening) have been evaluated on oil in terms of basic quality parameters, composition, organoleptic traits, and aroma profiles. A total of 40 metabolites (volatiles and non-volatiles) were identified by 1H NMR and GC/MS analyses. Multivariate statistical analysis showed that samples obtained from ripe and advanced ripe olives cooled at 10 and 15 °C better correlated with C6 aldehydes, mainly associated with herbal/green olfactory traits. Compounds responsible for sweet/fruity traits were more abundantly present in oil extracted from 19 °C olive samples. Decreasing pulp temperature before crushing also resulted in reduced presence of 1-penten-3-ol, 1-penten-3-one, acetic acid and ethyl alcohol, associated with specific defects of the oil. Results indicate that slightly lowering fruit temperature just before crushing modulates oil composition by reducing oil off flavours while enhancing green and fresh attributes in particular when ripe olives are processed.

Development of a Sensor Node for Remote Monitoring of Plants.

The appraisal of stress in plants is of great relevance in agriculture and any time the transport of living plants is involved. Wireless sensor networks (WSNs) are an optimal solution to simultaneously monitor a large number of plants in a mostly automatic way. A number of sensors are readily available to monitor indicators that are likely related to stress. The most common of them include the levels of total volatile compounds and CO2 together with common physical parameters such as temperature, relative humidity, and illumination, which are known to affect plants' behavior. Recent progress in microsensors and communication technologies, such as the LoRa protocol, makes it possible to design sensor nodes of high sensitivity where power consumption, transmitting distances, and costs are optimized. In this paper, the design of a WSN dedicated to plant stress monitoring is described. The nodes have been tested on European privet (Ligustrum Jonandrum) kept in completely different conditions in order to induce opposite level of stress. The results confirmed the relationship between the release of total Volatile Organic Compounds (VOCs) and the environmental conditions. A machine learning model based on recursive neural networks demonstrates that total VOCs can be estimated from the measure of the environmental parameters.

Short-Term Responses of Apple Fruit to Partial Reoxygenation during Extreme Hypoxic Storage Conditions.

The short-term (24 h) responses of apple fruit (cv. 'Granny Smith') to a shift in the oxygen concentration from 0.4 to 0.8 kPa, a protocol applied in the dynamic controlled atmosphere (DCA) storage technique, have been studied. Metabolomics and transcriptomics analyses of cortex tissue showed an immediate down-regulation of fermentative metabolism and of the GABA shunt in parallel with the activation of several 2-oxoglutarate-dependent dioxygenase genes. Down-regulation of the free phenylpropanoid pathway genes and the diversion of propanoid synthesis toward the methyl-erythritol phosphate route were also observed. Partial reoxygenation induced increases of glyceric, palmitic, and stearic acids and of several phosphatidylcholines and phosphatidylethanolamines and decreases of specific amino acids (valine, methionine, glycine, phenylalanine, and GABA), organic acids (arachidic and citric acids), and secondary metabolites (catechin and epicatechin). The oxygen shift also resulted in transcriptional rewiring of several components of IAA and ABA regulation and signaling. These results provide novel insights on the complexity of the short-term physiological responses of apple fruit to partial reoxygenation applied during DCA storage.

Metabolic Responses to Low Temperature of Three Peach Fruit Cultivars Differently Sensitive to Cold Storage.

Refrigerated storage is widely applied in order to maintain peach quality but it can also induce chilling injuries (CIs) such as flesh browning and bleeding, and mealiness. Peach fruit from three cultivars ('Red Haven', RH, 'Regina di Londa', RL, and 'Flaminia', FL) were stored for 4 weeks under low temperatures (0.5 and 5.5°C). GC-MS was employed to study changes in both metabolome and volatilome induced by cold storage in the mesocarp. CIs were assessed both at the end of each week of storage and after subsequent shelf-life (SL) at 20°C. Flesh browning and mealiness appeared to be more related to 5.5°C storage, while flesh bleeding revealed high incidence following 0.5°C storage. Compared to RL and FL, RH showed a marked lower incidence of CIs. Multivariate statistical analyses indicate that RH peaches indeed differ from RL and FL in particular when considering data from samples collected at the end of the cold storage. Common and divergent responses have been identified in terms of metabolic responses to the applied low temperatures. In all three cultivars raffinose, glucose-6P, fucose, xylose, sorbitol, GABA, epicatechin, catechin, and putrescine markedly increased during cold storage, while citramalic, glucuronic, mucic and shikimic acids decreased. Among volatile organic compounds (VOCs), aldehydes and alcohols generally accumulated more under low temperature conditions while esters and lactones evolved during subsequent SL. The main cultivar differences developed after cold storage during SL although some common responses (e.g., an increased production of ethyl acetate) were observed. The lower levels of flesh browning and bleeding displayed by RH peaches were related to compounds with antioxidant activity, or acting as osmotic protectants and membrane stabilizer. Indeed, RH showed higher levels of amino acids and urea, together with a marked increase in putrescine, sorbitol, maltitol, myoinositol and sucrose detected during storage and SL.

Extreme Hypoxic Conditions Induce Selective Molecular Responses and Metabolic Reset in Detached Apple Fruit.

The ripening physiology of detached fruit is altered by low oxygen conditions with profound effects on quality parameters. To study hypoxia-related processes and regulatory mechanisms, apple (Malus domestica, cv Granny Smith) fruit, harvested at commercial ripening, were kept at 1°C under normoxic (control) and hypoxic (0.4 and 0.8 kPa oxygen) conditions for up to 60 days. NMR analyses of cortex tissue identified eight metabolites showing significantly different accumulations between samples, with ethanol and alanine displaying the most pronounced difference between hypoxic and normoxic treatments. A rapid up-regulation of alcohol dehydrogenase and pyruvate-related metabolism (lactate dehydrogenase, pyruvate decarboxylase, alanine aminotransferase) gene expression was detected under both hypoxic conditions with a more pronounced effect induced by the lowest (0.4 kPa) oxygen concentration. Both hypoxic conditions negatively affected ACC synthase and ACC oxidase transcript accumulation. Analysis of RNA-seq data of samples collected after 24 days of hypoxic treatment identified more than 1000 genes differentially expressed when comparing 0.4 vs. 0.8 kPa oxygen concentration samples. Genes involved in cell-wall, minor and major CHO, amino acid and secondary metabolisms, fermentation and glycolysis as well as genes involved in transport, defense responses, and oxidation-reduction appeared to be selectively affected by treatments. The lowest oxygen concentration induced a higher expression of transcription factors belonging to AUX/IAA, WRKY, HB, Zinc-finger families, while MADS box family genes were more expressed when apples were kept under 0.8 kPa oxygen. Out of the eight group VII ERF members present in apple genome, two genes showed a rapid up-regulation under hypoxia, and western blot analysis showed that apple MdRAP2.12 proteins were differentially accumulated in normoxic and hypoxic samples, with the highest level reached under 0.4 kPa oxygen. These data suggest that ripe apple tissues finely and specifically modulate sensing and regulatory mechanisms in response to different hypoxic stress conditions.

Berry ripening, pre-processing and thermal treatments affect the phenolic composition and antioxidant capacity of grape (Vitis vinifera L.) juice.

BACKGROUND: Grape juice is an important dietary source of health-promoting antioxidant molecules. Different factors may affect juice composition and nutraceutical properties. The effects of some of these factors (harvest time, pre-processing ethylene treatment of grapes and juice thermal pasteurization) were here evaluated, considering in particular the phenolic composition and antioxidant capacity. RESULTS: Grapes (Vitis vinifera L., red-skinned variety Sangiovese) were collected twice in relation to the technological harvest (TH) and 12 days before TH (early harvest, EH) and treated with gaseous ethylene (1000 ppm) or air for 48 h. Fresh and pasteurized (78 °C for 30 min) juices were produced using a water bath. Three-way analysis of variance showed that the harvest date had the strongest impact on total polyphenols, hydroxycinnamates, flavonols, and especially on total flavonoids. Pre-processing ethylene treatment significantly increased the proanthocyanidin, anthocyanin and flavan-3-ol content in the juices. Pasteurization induced a significant increase in anthocyanin concentration. Antioxidant capacity was enhanced by ethylene treatment and pasteurization in juices from both TH and EH grapes. CONCLUSION: These results suggest that an appropriate management of grape harvesting date, postharvest and processing may lead to an improvement in nutraceutical quality of juices. Further research is needed to study the effect of the investigated factors on juice organoleptic properties.

Cell wall metabolism of peaches and nectarines treated with UV-B radiation: a biochemical and molecular approach.

BACKGROUND: Ultra-violet B (UV-B) radiation has been shown to improve, at least in selected genotypes, both the health-promoting potential and the aesthetic properties of tomato and peach fruits during their post-harvest period. The effects of post-harvest UV-B treatment on the cell-wall metabolism of peaches and nectarines (Prunus persica L. Batsch) were assessed in this study. Three cultivars, Suncrest (melting flesh, MF) and Babygold 7 (non-melting flesh, NMF) peaches and Big Top (slow melting, SM) nectarine, differing in the characteristics of textural changes and softening during ripening, were analysed. RESULTS: The effects of UV-B differ in relation to the cultivar considered. In MF 'Suncrest' fruit, UV-B treatment significantly reduced the loss of flesh firmness despite the slight increase in the presence and activity of endo-polygalacturonase. The activity of exo-polygalacturonase increased as well, while endo-1,4-ß-D-glucanase/ß-D-glucosidase, ß-galactosidase and pectin methylesterase were substantially unaffected by the treatment. The UV-B-induced reduction of flesh softening was paralleled by the inhibition of PpExp gene transcription and expansin protein accumulation. The UV-B treatment did not induce differences in flesh firmness between control and UV-B-treated NMF 'Babygold 7' and SM 'Big Top' fruit. CONCLUSION: Based on these results, post-harvest UV-B treatment may be considered a promising tool to improve shelf-life and quality of peach fruit.

Post-harvest UV-B irradiation induces changes of phenol contents and corresponding biosynthetic gene expression in peaches and nectarines.

In the present study the possibility of enhancing phenolic compound contents in peaches and nectarines by post-harvest irradiation with UV-B was assessed. Fruits of 'Suncrest' and 'Babygold 7' peach and 'Big Top' nectarine cultivars were irradiated with UV-B for 12 h, 24 h and 36 h. Control fruits underwent the same conditions but UV-B lamps were screened by benzophenone-treated polyethylene film. The effectiveness of the UV-B treatment in modulating the concentration of phenolic compounds and the expression of the phenylpropanoid biosynthetic genes, was genotype-dependent. 'Big Top' and 'Suncrest' fruits were affected by increasing health-promoting phenolics whereas in 'Babygold 7' phenolics decreased after UV-B irradiation. A corresponding trend was exhibited by most of tested phenylpropanoid biosynthesis genes. Based on these results UV-B irradiation can be considered a promising technique to increase the health-promoting potential of peach fruits and indirectly to ameliorate the aesthetic value due to the higher anthocyanin content.

Postharvest treatments with ethylene on Vitis vinifera (cv Sangiovese) grapes affect berry metabolism and wine composition.

Grapes (Vitis vinifera, cv Sangiovese), harvested at standard commercial maturity, were treated for 36 h with ethylene (ET, 1000 ppm) or air (control, CT) before vinification. The composition of the grapes, must and wine was different in the CT and ET samples. In the ET wine, higher concentrations of specific phenol compounds, belonging to the classes of flavonols, anthocyanins, flavan-3-ols, and stilbenes, were detected. ET induced a significant change in the wine aroma profile by increasing free volatile categories such as phenols and fatty acids, and reducing the content of carbonyl compounds and, in particular, of esters. Less pronounced differences between CT and ET wines were observed in terms of glycosidically-bound volatile compounds. The activity of pectin methyl esterase and ß-glucosidase was enhanced in ET-treated berry skins, suggesting that cell wall properties and changes in the hydrolytic activity are effective in modulating the composition of CT and ET wines.

The high-quality draft genome of peach (Prunus persica) identifies unique patterns of genetic diversity, domestication and genome evolution.

Rosaceae is the most important fruit-producing clade, and its key commercially relevant genera (Fragaria, Rosa, Rubus and Prunus) show broadly diverse growth habits, fruit types and compact diploid genomes. Peach, a diploid Prunus species, is one of the best genetically characterized deciduous trees. Here we describe the high-quality genome sequence of peach obtained from a completely homozygous genotype. We obtained a complete chromosome-scale assembly using Sanger whole-genome shotgun methods. We predicted 27,852 protein-coding genes, as well as noncoding RNAs. We investigated the path of peach domestication through whole-genome resequencing of 14 Prunus accessions. The analyses suggest major genetic bottlenecks that have substantially shaped peach genome diversity. Furthermore, comparative analyses showed that peach has not undergone recent whole-genome duplication, and even though the ancestral triplicated blocks in peach are fragmentary compared to those in grape, all seven paleosets of paralogs from the putative paleoancestor are detectable.

Short-term postharvest carbon dioxide treatments induce selective molecular and metabolic changes in grape berries.

Detached wine grapes ( Vitis vinifera cv. 'Trebbiano', white skinned) were treated for 3 days with 30 kPa of CO(2) and then transferred to air for an additional 9 days to partially dehydrate (about 20% weight loss). At the end of the CO(2) treatment on withering berries, total polyphenols and flavonoids were maintained in the skin, but to a more limited extent in the pulp. An induction of the proanthocyanidin synthesis appeared to be one of the responses to the treatment because both (+)-catechin and (-)-epicatechin concentrations increased in the skin. The skin and pulp of the grape berries showed different molecular responses to a high CO(2) treatment. As revealed by microarray hybridizations, 217 and 75 genes appeared differentially expressed in the skin and pulp of treated samples, respectively. Functional categorization and gene enrichment analyses pointed out that epicarp cells undergo more pronounced changes in transcript profiling at the end of the incubation period. Highly represented categories in both tissues were related to protein, stress, transcript, RNA, and hormone (ethylene, ABA) metabolism. Fermentation, CHO metabolism, and redox regulation functional categories were represented only in the skin.

Computational annotation of genes differentially expressed along olive fruit development.

BACKGROUND: Olea europaea L. is a traditional tree crop of the Mediterranean basin with a worldwide economical high impact. Differently from other fruit tree species, little is known about the physiological and molecular basis of the olive fruit development and a few sequences of genes and gene products are available for olive in public databases. This study deals with the identification of large sets of differentially expressed genes in developing olive fruits and the subsequent computational annotation by means of different software. RESULTS: mRNA from fruits of the cv. Leccino sampled at three different stages [i.e., initial fruit set (stage 1), completed pit hardening (stage 2) and veraison (stage 3)] was used for the identification of differentially expressed genes putatively involved in main processes along fruit development. Four subtractive hybridization libraries were constructed: forward and reverse between stage 1 and 2 (libraries A and B), and 2 and 3 (libraries C and D). All sequenced clones (1,132 in total) were analyzed through BlastX against non-redundant NCBI databases and about 60% of them showed similarity to known proteins. A total of 89 out of 642 differentially expressed unique sequences was further investigated by Real-Time PCR, showing a validation of the SSH results as high as 69%. Library-specific cDNA repertories were annotated according to the three main vocabularies of the gene ontology (GO): cellular component, biological process and molecular function. BlastX analysis, GO terms mapping and annotation analysis were performed using the Blast2GO software, a research tool designed with the main purpose of enabling GO based data mining on sequence sets for which no GO annotation is yet available. Bioinformatic analysis pointed out a significantly different distribution of the annotated sequences for each GO category, when comparing the three fruit developmental stages. The olive fruit-specific transcriptome dataset was used to query all known KEGG (Kyoto Encyclopaedia of Genes and Genomes) metabolic pathways for characterizing and positioning retrieved EST records. The integration of the olive sequence datasets within the MapMan platform for microarray analysis allowed the identification of specific biosynthetic pathways useful for the definition of key functional categories in time course analyses for gene groups. CONCLUSION: The bioinformatic annotation of all gene sequences was useful to shed light on metabolic pathways and transcriptional aspects related to carbohydrates, fatty acids, secondary metabolites, transcription factors and hormones as well as response to biotic and abiotic stresses throughout olive drupe development. These results represent a first step toward both functional genomics and systems biology research for understanding the gene functions and regulatory networks in olive fruit growth and ripening.