Project description:Background: Grape cultivars and wines are distinguishable by their color, flavor and aroma profiles. Omic analyses (transcripts, proteins and metabolites) are powerful tools for assessing biochemical differences in biological systems. Results: Berry skins of red- (Cabernet Sauvignon, Merlot, Pinot Noir) and white-skinned (Chardonnay, Semillon) wine grapes were harvested near optimum maturity from the same experimental vineyard and Ë?Brix-to-titratable acidity ratio. Identical sample aliquots were analyzed for transcripts by grapevine whole-genome oligonucleotide microarray and RNA-seq technologies, proteins by nano-liquid chromatography-mass spectroscopy, and metabolites by gas chromatography-mass spectroscopy and liquid chromatography-mass spectroscopy. Principal components analysis of each of five Omic technologies showed similar results across cultivars in all Omic datasets. Comparison of the processed data of genes mapped in RNA-seq and microarray data revealed a strong Pearson's correlation (0.80). The exclusion of probesets associated with genes with potential for cross-hybridization on the microarray improved the correlation to 0.93. The overall concordance of protein with transcript data was low with a Pearson's correlation of 0.27 and 0.24 for the RNA-seq and microarray data, respectively. Integration of metabolite with protein and transcript data produced an expected model of phenylpropanoid biosynthesis, which distinguished red from white grapes, yet provided detail of individual cultivar differences. Conclusions: The five Omic technologies were consistent in distinguishing cultivar variation. There was high concordance between transcriptomic technologies, but generally protein abundance did not correlate well with transcript abundance. The integration of multiple high-throughput Omic datasets revealed complex biochemical variation amongst five cultivars of an ancient and economically important crop species. Vitis vinifera L. cv. Cabernet Sauvignon, Chardonnay, Merlot, Pinot Noir, Semillon berries were harvested from Nevada Agricultural Experiment Station Valley Road Vineyard, Reno, NV, USA. Whole-genome microarray analysis was used to assess the transcriptomic response of berry skins at harvest, approximately 24 Â°Brix (2011 vintage). Vines were grown under water deficit and well-watered conditions. At least two clusters harvested from non-adjacent vines were used for each of five experimental replicates.

Project description:Grapevine red blotch is a recently identified viral disease that was first recognized in the Napa Valley of California. Infected plants showed foliar symptoms similar to leafroll, another grapevine viral disease, on vines testing negative for known grapevine leafroll-associated virus. Later, the Grapevine red blotch virus (GRBV) was independently discovered in the US states of California and New York and was demonstrated to be the causal agent of red blotch disease. Due to its wide occurrence in the US, vector transmission and impacts on grape industry, this virus has the potential to cause serious economic losses. Despite numerous attempts, it was not possible to isolate or visualize viral particles from GRBV infected plants. Consequently, this has hampered the development of a serological assay that would facilitate GRBV detection in grapevine. We therefore decided to explore mass spectrometry approaches in order to quantify GRBV in infected plants and to identify potential biomarkers for viral infection. We present for the first time the physical detection on the protein level of the two GRBV genes V1 (coat protein) and V2 in grapevine tissue lysates. The GRBV coat protein load in leaf petioles was determined to be in the range of 100 to 900 million copies per milligram wet weight by using three heavy isotope labeled reference peptides as internal standards. The V1 copy number per unit wet tissue weight in leaves appeared to be about six times lower, and about 200-times lower in terms of protein concentration in the extractable protein mass than in petioles. We found a consistent upregulation of several enzymes involved in flavonoid biosynthesis in leaf and petiole extracts of GRBV-infected plants by label-free shotgun proteomics, indicating the activation of a defense mechanism against GRBV, a plant response already described for grapevine leafroll associated virus infection on the transcriptome level. Last but not least, we identified some other microorganisms belonging to the grapevine leaf microbiota, two bacterial species (Novosphingobium sp. Rr 2-17 and Methylobacterium) and one virus, Grapevine rupestris stem pitting associated virus.

Project description:Grapevine rupestris stem pitting-associated virus (GRSPaV) is a widespread virus affecting Vitis spp. Although it has established a compatible viral interaction in Vitis vinifera L. without the development of phenotypic alterations, it can occur as distinct variants that show different symptoms in diverse Vitis species. We investigated the changes induced by GRSPaV in V. vinifera cv Bosco, an Italian white grape variety, by combining agronomic, physiological, and molecular approaches, in order to provide comprehensive information about the global effects of GRSPaV. In two consecutive years, this virus caused a moderate decrease in physiological efficiency, yield performance, and sugar content in berries associated with several transcriptomic alterations. Transcript profiles were analysed by microarray techniques in petiole, leaf, and berry samples collected at véraison and by quantitative real-time RT-PCR (qRT-PCR) in a time course carried out at five relevant grapevine developmental stages. Global gene expression analyses showed that transcriptomic changes were highly variable among the different organs and the different phenological phases. GRSPaV triggers some unique responses in the grapevine at véraison, never reported before for other plant-virus interactions, such as an increase in transcripts involved in photosynthesis and CO2 fixation, associated with a moderate reduction in the photosynthesis rate and some defence mechanisms, and to an overlap with responses to water and salinity stresses. We hypothesise that the long co-existence between grapevine and GRSPaV has resulted in the evolution of a form of mutual adaptation between the virus and its host. This study contributes to elucidating alternative mechanisms used by infected plants to contend with viruses. The study was carried out in a vineyard planted in 2002 in Albenga (Liguria), North-West Italy, where a row was established with the white grape cultivar Bosco (V. vinifera L.). Microarray analysis was carried out on leaves, petioles, and berries collected at véraison (E-L35) in 2010. For each of the six GRSPaV-free and six GRSPaV-infected vines selected for the physiological and agronomical parameters evaluation, we collected 6 leaves (3 basal and 3 apical) with the related petioles and 12 berries from 3 different bunches. Samples from each organ were arbitrary pooled in 3 independent biological replicates and total RNA was extracted according to the method described by Gambino et al. (2008).

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