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.
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).
Project description:MicroRNA (miRNA) is a class of functional non-coding small RNA with 19-25 nucleotides in length. Amur grape (Vitis amurensis Rupr.) is an important wild fruit crop with the strongest cold resistance in the Vitis genus and is used as an excellent breeding parent for grapevine, and with growing interest in terms of wine production. To date, there is a relatively large number of grapevine miRNAs (vv-miRNAs) from cultivated grapevine varieties such as Vitis vinifera L. and hybrids of V. vinifera and V. labrusca, but there is no report on miRNAs from Vitis amurensis Rupr, a wild grapevine species. In this study, a small RNA library from Amur grapes was constructed and Solexa technology used to perform deep sequencing of the library followed by subsequent bioinformatics analysis to identify new miRNAs. In total, 126 conserved miRNA belonging to 27 miRNA families were identified, and 34 known but non-conserved miRNAs were also found. Significantly, 72 new potential Amur grapevine-specific miRNAs were discovered. The sequences of these new potential va-miRNAs were further validated through miR-RACE, accumulation of 18 new va-miRNAs in seven tissues of grapevines were also confirmed by real time RT-PCR (qRT-PCR) analysis, and expression levels of va-miRNAs in flowers and berries were basically consistent in identity to those from deep sequenced sRNAs libraries of independent corresponding tissues. We also describe the conservation and variation of va-miRNAs using miR-SNPs and miR-LDs during plant evolution based on comparison of orthologous sequences, and revealed the number and sites of miR-SNP of diverse miRNA families exhibited distinct divergence. Finally, 346 target genes for the new miRNAs were predicted and they include a number of Amur grapevine stress tolerance genes and many genes regulating anthocyanin systhesis and sugar metabolism. Deep sequencing of short RNAs from Amur grapes flowers and fruits identified 72 new potential miRNAs and 34 known but non-conserved miRNAs, indicating that specific miRNAs exist in Amur grapes. These results show that a number of regulatory miRNAs exist in Amur grapes and play an important role in Amur grape growth, development, and response to abiotic or biotic stress. High throughput sequencing was employed to identify miRNAs in Amur grapevine and try to describe their functions in Amur grapevine growth and development.
Project description:Cultivated grapevine (Vitis vinifera) is susceptible to many pathogens which cause significant losses to viticulture worldwide. Chemical control is available, but agro-ecological concerns have raised interest in alternative methods, especially in elicitation of plant immunity by bio-molecules such as Pathogen Associated Molecular Patterns (PAMPs). We have demonstrated that the beta-glucan laminarin (Lam) and its sulfated derivative (PS3) induce a PAMP-triggered immunity in grapevine against downy mildew (Plasmopara viticola). However, if Lam elicits classical grapevine defenses, PS3 triggered grapevine resistance via a poorly understood priming phenomenon. The aim of this study was to discover the mechanism of the PS3-induced resistance. On uninfected grapevine, we first investigated defense signaling and performed microarray experiments to identify early events and genes directly triggered by PS3. Our results showed that PS3 i) was unable to elicit ROS and NO production, cytosolic Ca2+ variations, MAPK activation but triggered a long lasting plasma membrane depolarization in grapevine cells ii) up-regulated a stress-responsive transcriptome close to the one induced by Lam but only partly overlapping the ones triggered by salicylate (SA) or jasmonate (JA). Finally, in response to P. viticola infection, PS3 specifically primed the SA- and ROS-dependent defense pathways leading to grapevine triggered immunity against this biotroph. Keywords: cell death, induced resistance, oomycete, priming, reactive oxygen species, salicylate, sulfated laminarin, transcriptomics, Vitis vinifera.
Project description:Cultivated grapevine (Vitis vinifera) is susceptible to many pathogens which cause significant losses to viticulture worldwide. Chemical control is available, but agro-ecological concerns have raised interest in alternative methods, especially in elicitation of plant immunity by bio-molecules such as Pathogen Associated Molecular Patterns (PAMPs). We have demonstrated that the beta-glucan laminarin (Lam) and its sulfated derivative (PS3) induce a PAMP-triggered immunity in grapevine against downy mildew (Plasmopara viticola). However, if Lam elicits classical grapevine defenses, PS3 triggered grapevine resistance via a poorly understood priming phenomenon. The aim of this study was to discover the mechanism of the PS3-induced resistance. On uninfected grapevine, we first investigated defense signaling and performed microarray experiments to identify early events and genes directly triggered by PS3. Our results showed that PS3 i) was unable to elicit ROS and NO production, cytosolic Ca2+ variations, MAPK activation but triggered a long lasting plasma membrane depolarization in grapevine cells ii) up-regulated a stress-responsive transcriptome close to the one induced by Lam but only partly overlapping the ones triggered by salicylate (SA) or jasmonate (JA). Finally, in response to P. viticola infection, PS3 specifically primed the SA- and ROS-dependent defense pathways leading to grapevine triggered immunity against this biotroph. Keywords: cell death, induced resistance, oomycete, priming, reactive oxygen species, salicylate, sulfated laminarin, transcriptomics, Vitis vinifera. 6 samples (Adj, PS3, Lam, ctrl, SA, JA) were analized with 3 biological replicates each, Adj and ctrl samples are reference samples
Project description:The growth and fruit quality of grapevine are widely affected by abnormal climatic conditions such as extreme temperature. But how grapevine responds to cold stress is still largely unknown. Here we found that VaMyb14, a member of R2R3 Myb transcription factor family, was up-regulated dramatically during cold, drought and salinity treatments in Vitis amurensis, a cold and drought-hardiness wild Vitis species. Overexpression VaMyb14 in Arabidopsis increased antioxidant enzyme activity, especially POD activity, than that of the wild type and decreased the MDA content. A series of ABA metabolism and signal transduction genes in transgenic Arabidopsiswere were up-regulated in microarry results, including several nsLTPs, PP2Cs, RD29B, COR78 and other structural genes, suggesting that VaMyb14 not only affect the ABA signaling pathways, but also activates the CBF-COR independent nsLTP genes. Collectively, these results illustrate that Vitis Myb14 could represent a node of convergence regulating grapevine stress responses, including improve defence induced phytoalexin resveratrol against necrotrophic as well as drought and/or cold stress tolerance, highlighting Myb14 as a potential gene resource in future grapevine breeding.