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1.  Dissecting the Transcriptional Response to Elicitors in Vitis vinifera Cells 
PLoS ONE  2014;9(10):e109777.
The high effectiveness of cyclic oligosaccharides like cyclodextrins in the production of trans-resveratrol in Vitis vinifera cell cultures is enhanced in the presence of methyl jasmonate. In order to dissect the basis of the interactions among the elicitation responses triggered by these two compounds, a transcriptional analysis of grapevine cell cultures treated with cyclodextrins and methyl jasmonate separately or in combination was carried out. The results showed that the activation of genes encoding enzymes from phenylpropanoid and stilbene biosynthesis induced by cyclodextrins alone was partially enhanced in the presence of methyl jasmonate, which correlated with their effects on trans-resveratrol production. In addition, protein translation and cell cycle regulation were more highly repressed in cells treated with cyclodextrins than in those treated with methyl jasmonate, and this response was enhanced in the combined treatment. Ethylene signalling was activated by all treatments, while jasmonate signalling and salicylic acid conjugation were activated only in the presence of methyl jasmonate and cyclodextrins, respectively. Moreover, the combined treatment resulted in a crosstalk between the signalling cascades activated by cyclodextrins and methyl jasmonate, which, in turn, provoked the activation of additional regulatory pathways involving the up-regulation of MYB15, NAC and WRKY transcription factors, protein kinases and calcium signal transducers. All these results suggest that both elicitors cause an activation of the secondary metabolism in detriment of basic cell processes like the primary metabolism or cell division. Crosstalk between cyclodextrins and methyl jasmonate-induced signalling provokes an intensification of these responses resulting in a greater trans-resveratrol production.
PMCID: PMC4196943  PMID: 25314001
2.  Correction: Berry Flesh and Skin Ripening Features in Vitis vinifera as Assessed by Transcriptional Profiling 
PLoS ONE  2012;7(10):10.1371/annotation/fd93800a-3b3c-484d-97a9-190043309e4b.
PMCID: PMC3935808
3.  Berry Flesh and Skin Ripening Features in Vitis vinifera as Assessed by Transcriptional Profiling 
PLoS ONE  2012;7(6):e39547.
Ripening of fleshy fruit is a complex developmental process involving the differentiation of tissues with separate functions. During grapevine berry ripening important processes contributing to table and wine grape quality take place, some of them flesh- or skin-specific. In this study, transcriptional profiles throughout flesh and skin ripening were followed during two different seasons in a table grape cultivar ‘Muscat Hamburg’ to determine tissue-specific as well as common developmental programs.
Methodology/Principal Findings
Using an updated GrapeGen Affymetrix GeneChip® annotation based on grapevine 12×v1 gene predictions, 2188 differentially accumulated transcripts between flesh and skin and 2839 transcripts differentially accumulated throughout ripening in the same manner in both tissues were identified. Transcriptional profiles were dominated by changes at the beginning of veraison which affect both pericarp tissues, although frequently delayed or with lower intensity in the skin than in the flesh. Functional enrichment analysis identified the decay on biosynthetic processes, photosynthesis and transport as a major part of the program delayed in the skin. In addition, a higher number of functional categories, including several related to macromolecule transport and phenylpropanoid and lipid biosynthesis, were over-represented in transcripts accumulated to higher levels in the skin. Functional enrichment also indicated auxin, gibberellins and bHLH transcription factors to take part in the regulation of pre-veraison processes in the pericarp, whereas WRKY and C2H2 family transcription factors seems to more specifically participate in the regulation of skin and flesh ripening, respectively.
A transcriptomic analysis indicates that a large part of the ripening program is shared by both pericarp tissues despite some components are delayed in the skin. In addition, important tissue differences are present from early stages prior to the ripening onset including tissue-specific regulators. Altogether, these findings provide key elements to understand berry ripening and its differential regulation in flesh and skin.
PMCID: PMC3386993  PMID: 22768087
4.  A 48 SNP set for grapevine cultivar identification 
BMC Plant Biology  2011;11:153.
Rapid and consistent genotyping is an important requirement for cultivar identification in many crop species. Among them grapevine cultivars have been the subject of multiple studies given the large number of synonyms and homonyms generated during many centuries of vegetative multiplication and exchange. Simple sequence repeat (SSR) markers have been preferred until now because of their high level of polymorphism, their codominant nature and their high profile repeatability. However, the rapid application of partial or complete genome sequencing approaches is identifying thousands of single nucleotide polymorphisms (SNP) that can be very useful for such purposes. Although SNP markers are bi-allelic, and therefore not as polymorphic as microsatellites, the high number of loci that can be multiplexed and the possibilities of automation as well as their highly repeatable results under any analytical procedure make them the future markers of choice for any type of genetic identification.
We analyzed over 300 SNP in the genome of grapevine using a re-sequencing strategy in a selection of 11 genotypes. Among the identified polymorphisms, we selected 48 SNP spread across all grapevine chromosomes with allele frequencies balanced enough as to provide sufficient information content for genetic identification in grapevine allowing for good genotyping success rate. Marker stability was tested in repeated analyses of a selected group of cultivars obtained worldwide to demonstrate their usefulness in genetic identification.
We have selected a set of 48 stable SNP markers with a high discrimination power and a uniform genome distribution (2-3 markers/chromosome), which is proposed as a standard set for grapevine (Vitis vinifera L.) genotyping. Any previous problems derived from microsatellite allele confusion between labs or the need to run reference cultivars to identify allele sizes disappear using this type of marker. Furthermore, because SNP markers are bi-allelic, allele identification and genotype naming are extremely simple and genotypes obtained with different equipments and by different laboratories are always fully comparable.
PMCID: PMC3221639  PMID: 22060012
5.  Transcriptome changes in grapevine (Vitis vinifera L.) cv. Malbec leaves induced by ultraviolet-B radiation 
BMC Plant Biology  2010;10:224.
Ultraviolet-B radiation (UV-B, 280-315 nm) is a natural component of sunlight, which has numerous regulatory effects on plant physiology. The nature of the response to UV-B is dependent on fluence rate, dose, duration and wavelength of the UV-B treatment. Some reports have analyzed the changes in gene expression caused by UV-B light on several plant species using microarray technology. However, there is no information on the transcriptome response triggered by UV-B in grapevine. In this paper we investigate the gene expression responses of leaves from in vitro cultured Vitis vinifera cv. Malbec plants subjected to the same dose of biologically effective UV-B radiation (4.75 kJ m-2 d-1) administered at two different fluence rates (16 h at ≅ 8.25 μW cm-2, 4 h at ≅ 33 μW cm-2) using a new custom made GrapeGen Affymetrix GeneChip®.
The number of genes modulated by high fluence rate UV-B doubled the number of genes modulated by low fluence UV-B. Their functional analyses revealed several functional categories commonly regulated by both UV-B treatments as well as categories more specifically modulated depending on UV-B fluence rate. General protective responses, namely the induction of pathways regulating synthesis of UV-B absorbing compounds such as the Phenylpropanoid pathway, the induction of different antioxidant defense systems and the activation of pathways commonly associated with pathogen defense and abiotic stress responses seem to play critical roles in grapevine responses against UV-B radiation. Furthermore, high fluence rate UV-B seemed to specifically modulate additional pathways and processes in order to protect grapevine plantlets against UV-B-induced oxidative stress, stop the cell cycle progression, and control protein degradation. On the other hand, low fluence rate UV-B regulated the expression of specific responses in the metabolism of auxin and abscisic acid as well as in the modification of cell walls that could be involved in UV-B acclimation-like processes.
Our results show the UV-B radiation effects on the leaf transcriptome of grapevine (Vitis vinifera cv. Malbec) plantlets. Functional categories commonly modulated under both UV-B treatments as well as transcripts specifically regulated in an UV-B-intensity dependent way were identified. While high fluence rate UV-B had regulatory effects mainly on defense or general multiple-stress responses pathways, low fluence rate UV-B promoted the expression of genes that could be involved in UV-B protection or the amelioration of the UV-B-induced damage. This study also provides an extensive list of genes regulating multiple metabolic pathways involved in the response of grapevine to UV-B that can be used for future researches.
PMCID: PMC3017828  PMID: 20959019
6.  Synergistic effect of methyljasmonate and cyclodextrin on stilbene biosynthesis pathway gene expression and resveratrol production in Monastrell grapevine cell cultures 
BMC Research Notes  2008;1:132.
Plant cell cultures have been shown as feasible systems for the production of secondary metabolites, being the elicitation with biotic or abiotic stimuli the most efficient strategy to increase the production of those metabolites. Vitaceae phytoalexins constitute a group of molecules belonging to the stilbene family which are derivatives of the trans-resveratrol structure and are produced by plants and cell cultures as a response to biotic and abiotic stresses. The potential benefits of resveratrol on human health have made it one of the most thoroughly studied phytochemical molecules. The aim of this study was to evaluate the elicitor effect of both cyclodextrin (CD) and methyljasmonate (MeJA) on grapevine cell cultures by carrying out a quantitative analysis of their role on resveratrol production and on the expression of stilbene biosynthetic genes in Vitis vinifera cv Monastrell albino cell suspension cultures.
MeJA and CD significantly but transiently induced the expression of stilbene biosynthetic genes when independently used to treat grapevine cells. This expression correlated with resveratrol production in CD-treated cells but not in MeJA-treated cells, which growth was drastically affected. In the combined treatment of CD and MeJA cell growth was similarly affected, however resveratrol production was almost one order of magnitude higher, in correlation with maximum expression values for stilbene biosynthetic genes.
The effect of MeJA on cell division combined with a true and strong elicitor like CD could be responsible for the observed synergistic effect of both compounds on resveratrol production and on the expression of genes in the stilbene pathway.
PMCID: PMC2628674  PMID: 19102745
7.  High throughput SNP discovery and genotyping in grapevine (Vitis vinifera L.) by combining a re-sequencing approach and SNPlex technology 
BMC Genomics  2007;8:424.
Single-nucleotide polymorphisms (SNPs) are the most abundant type of DNA sequence polymorphisms. Their higher availability and stability when compared to simple sequence repeats (SSRs) provide enhanced possibilities for genetic and breeding applications such as cultivar identification, construction of genetic maps, the assessment of genetic diversity, the detection of genotype/phenotype associations, or marker-assisted breeding. In addition, the efficiency of these activities can be improved thanks to the ease with which SNP genotyping can be automated. Expressed sequence tags (EST) sequencing projects in grapevine are allowing for the in silico detection of multiple putative sequence polymorphisms within and among a reduced number of cultivars. In parallel, the sequence of the grapevine cultivar Pinot Noir is also providing thousands of polymorphisms present in this highly heterozygous genome. Still the general application of those SNPs requires further validation since their use could be restricted to those specific genotypes.
In order to develop a large SNP set of wide application in grapevine we followed a systematic re-sequencing approach in a group of 11 grape genotypes corresponding to ancient unrelated cultivars as well as wild plants. Using this approach, we have sequenced 230 gene fragments, what represents the analysis of over 1 Mb of grape DNA sequence. This analysis has allowed the discovery of 1573 SNPs with an average of one SNP every 64 bp (one SNP every 47 bp in non-coding regions and every 69 bp in coding regions). Nucleotide diversity in grape (π = 0.0051) was found to be similar to values observed in highly polymorphic plant species such as maize. The average number of haplotypes per gene sequence was estimated as six, with three haplotypes representing over 83% of the analyzed sequences. Short-range linkage disequilibrium (LD) studies within the analyzed sequences indicate the existence of a rapid decay of LD within the selected grapevine genotypes. To validate the use of the detected polymorphisms in genetic mapping, cultivar identification and genetic diversity studies we have used the SNPlex™ genotyping technology in a sample of grapevine genotypes and segregating progenies.
These results provide accurate values for nucleotide diversity in coding sequences and a first estimate of short-range LD in grapevine. Using SNPlex™ genotyping we have shown the application of a set of discovered SNPs as molecular markers for cultivar identification, linkage mapping and genetic diversity studies. Thus, the combination a highly efficient re-sequencing approach and the SNPlex™ high throughput genotyping technology provide a powerful tool for grapevine genetic analysis.
PMCID: PMC2212664  PMID: 18021442
8.  Genome-wide comparative phylogenetic analysis of the rice and Arabidopsis Dof gene families 
Dof proteins are a family of plant-specific transcription factors that contain a particular class of zinc-finger DNA-binding domain. Members of this family have been found to play diverse roles in gene regulation of processes restricted to the plants. The completed genome sequences of rice and Arabidopsis constitute a valuable resource for comparative genomic analyses, since they are representatives of the two major evolutionary lineages within the angiosperms. In this framework, the identification of phylogenetic relationships among Dof proteins in these species is a fundamental step to unravel functionality of new and yet uncharacterised genes belonging to this group.
We identified 30 different Dof genes in the rice Oryza sativa genome and performed a phylogenetic analysis of a complete collection of the 36-reported Arabidopsis thaliana and the rice Dof transcription factors identified herein. This analysis led to a classification into four major clusters of orthologous genes and showed gene loss and duplication events in Arabidopsis and rice, that occurred before and after the last common ancestor of the two species.
According to our analysis, the Dof gene family in angiosperms is organized in four major clusters of orthologous genes or subfamilies. The proposed clusters of orthology and their further analysis suggest the existence of monocot specific genes and invite to explore their functionality in relation to the distinct physiological characteristics of these evolutionary groups.
PMCID: PMC184357  PMID: 12877745

Results 1-8 (8)