Project description:BACKGROUND:Roots play a central role in plant response to water stress (WS). They are involved in its perception and signalling to the leaf as well as in allowing the plant to adapt to maintaining an adequate water balance. Only a few studies have investigated the molecular/biochemical responses to WS in roots of perennial plants, such as grapevine. This study compares two grapevine rootstock genotypes (i.e. 101.14 and M4) with different tolerance to WS, evaluating the responses at proteomic and metabolite levels. RESULTS:WS induced changes in the abundance of several proteins in both genotypes (17 and 22% of the detected proteins in 101.14 and M4, respectively). The proteomic analysis revealed changes in many metabolic pathways that fitted well with the metabolite data. M4 showed metabolic responses which were potentially able to counteract the WS effects, such as the drop in cell turgor, increased oxidative stress and loss of cell structure integrity/functionality. However, in 101.14 it was evident that the roots were suffering more severely from these effects. We found that many proteins classified as active in energy metabolism, hormone metabolism, protein, secondary metabolism and stress functional classes showed particular differences between the two rootstocks. CONCLUSION:The proteomic/metabolite comparative analysis carried out provides new information on the possible biochemical and molecular strategies adopted by grapevine roots to counteract WS. Although further work is needed to define in detail the role(s) of the proteins and metabolites that characterize WS response, this study, involving the M4 rootstock genotype, highlights that osmotic responses, modulations of C metabolism, mitochondrial functionality and some specific responses to stress occurring in the roots play a primary role in Vitis spp. tolerance to this type of abiotic stress.
Project description:Most of the vineyards around the world are in areas characterized by seasonal drought, where water deficits and high temperatures represent severe constraints on the regular grapevine growth cycle. Although grapevines are well adapted to arid and semi-arid environments, water stress can cause physiological changes, from mild to irreversible. Screening of available Vitis spp. genetic diversity for new rootstock breeding programs has been proposed as a way for which new viticulture challenges may be faced. In 2014, novel genotypes (M-rootstocks) were released from the University of Milan. In this work, the behavior of M1, M3 and M4 in response to decreasing water availabilities (80%, 50% and 20% soil water content, SWC) was investigated at the physiological and gene expression levels, evaluating gas exchange, stem water potential and transcript abundances of key genes related to ABA (abscisic acid) biosynthesis (VvZEP, VvNCED1 and VvNCED2) and signaling (VvPP2C4, VvSnRK2.6 and VvABF2), and comparing them to those of cuttings of nine commercial rootstocks widely used in viticulture. M-rootstocks showed a change at physiological levels in severe water-stressed conditions (20% soil water content, SWC), reducing the stomatal conductance and stem water potential, but maintaining high photosynthetic activity. Water use efficiency was high in water-limiting conditions. The transcriptional changes were observed at 50% SWC, with an increment of transcripts of VvNCED1 and VvNCED2 genes. M-rootstocks showed similar behavior to 1103P and 110R rootstocks, two highly tolerant commercial genotypes. These rootstocks adopted a tolerant strategy to face water-stressed conditions.
Project description:Salinity is ubiquitous abiotic stress factor limiting viticulture productivity worldwide. However, the grapevine is vulnerable to salt stress, which severely affects growth and development of the vine. Hence, it is crucial to delve into the salt resistance mechanism and screen out salt-resistance prediction marker genes; we implicated RNA-sequence (RNA-seq) technology to compare the grapevine transcriptome profile to salt stress. Results showed 2472 differentially-expressed genes (DEGs) in total in salt-responsive grapevine leaves, including 1067 up-regulated and 1405 down-regulated DEGs. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) annotations suggested that many DEGs were involved in various defense-related biological pathways, including ROS scavenging, ion transportation, heat shock proteins (HSPs), pathogenesis-related proteins (PRs) and hormone signaling. Furthermore, many DEGs were encoded transcription factors (TFs) and essential regulatory proteins involved in signal transduction by regulating the salt resistance-related genes in grapevine. The antioxidant enzyme analysis showed that salt stress significantly affected the superoxide dismutase (SOD), peroxidase (POD), catalase (CAT) and glutathione S-transferase (GST) activities in grapevine leaves. Moreover, the uptake and distribution of sodium (Na?), potassium (K?) and chlorine (Cl-) in source and sink tissues of grapevine was significantly affected by salt stress. Finally, the qRT-PCR analysis of DE validated the data and findings were significantly consistent with RNA-seq data, which further assisted in the selection of salt stress-responsive candidate genes in grapevine. This study contributes in new perspicacity into the underlying molecular mechanism of grapevine salt stress-tolerance at the transcriptome level and explore new approaches to applying the gene information in genetic engineering and breeding purposes.
Project description:In agricultural soils, nitrate (NO<sub>3</sub><sup>-</sup>) is the major nitrogen (N) nutrient for plants, but few studies have analyzed molecular and biochemical responses involved in its acquisition by grapevine roots. In viticulture, considering grafting, NO<sub>3</sub><sup>-</sup> acquisition is strictly dependent on rootstock. To improve the knowledge about N nutrition in grapevine, this study analyzed biochemical and proteomic changes induced by, NO<sub>3</sub><sup>-</sup> availability, in a hydroponic system, in the roots of M4, a recently selected grapevine rootstock. The evaluation of biochemical parameters, such as NO<sub>3</sub><sup>-</sup>, sugar and amino acid contents in roots, and the abundance of nitrate reductase, allowed us to define the time course of the metabolic adaptations to NO<sub>3</sub><sup>-</sup> supply. On the basis of these results, the proteomic analysis was conducted by comparing the root profiles in N-starved plants and after 30 h of NO<sub>3</sub><sup>-</sup> resupply. The analysis quantified 461 proteins, 26% of which differed in abundance between conditions. Overall, this approach highlighted, together with an increased N assimilatory metabolism, a concomitant rise in the oxidative pentose phosphate pathway and glycolysis, needed to fulfill the redox power and carbon skeleton demands, respectively. Moreover, a wide modulation of protein and amino acid metabolisms and changes of proteins involved in root development were observed. Finally, some results open new questions about the importance of redox-related post-translational modifications and of NO<sub>3</sub><sup>-</sup> availability in modulating the dialog between root and rhizosphere.
Project description:In modern viticulture, grafting commercial grapevine varieties on interspecific rootstocks is a common practice required for conferring resistance to many biotic and abiotic stresses. Nevertheless, the use of rootstocks to gain these essential traits is also known to impact grape berry development and quality, although the underlying mechanisms are still poorly understood. In grape berries, the onset of ripening (véraison) is regulated by a complex network of mobile signals including hormones such as auxins, ethylene, abscisic acid, and brassinosteroids. Recently, a new rootstock, designated M4, was selected based on its enhanced tolerance to water stress and medium vigor. This study investigates the effect of M4 on Cabernet Sauvignon (CS) berry development in comparison to the commercial 1103P rootstock. Physical and biochemical parameters showed that the ripening rate of CS berries is faster when grafted onto M4. A multifactorial analysis performed on mRNA-Seq data obtained from skin and pulp of berries grown in both graft combinations revealed that genes controlling auxin action (ARF and Aux/IAA) represent one of main categories affected by the rootstock genotype. Considering that the level of auxin tightly regulates the transcription of these genes, we investigated the behavior of the main gene families involved in auxin biosynthesis and conjugation. Molecular and biochemical analyses confirmed a link between the rate of berry development and the modulation of auxin metabolism. Moreover, the data indicate that this phenomenon appears to be particularly pronounced in skin tissue in comparison to the flesh.
Project description:Grapevine trunk diseases (GTDs) are caused by cryptic complexes of fungal pathogens and have become a growing problem for new grapevine (<i>Vitis vinifera</i>) plantations. We studied the role of the nursery, variety, and rootstock in the composition of the fungal communities in root collars and graft unions of planting material in Catalonia (NE Spain). We compared necrosis and fungal communities in graft unions and root collars at harvest, and then after three months of cold storage. We evaluated combinations of eleven red and five white varieties with four common rootstocks coming from six nurseries. Fungal communities were characterized by isolation and metabarcoding of the ITS2 region. Our data suggests that nursery followed by rootstock and variety had significant effects on necrosis and fungal community structure in graft and root tissues. Within the plant, we found large differences in terms fungal community distribution between graft and root tissues. Graft unions housed a significantly higher relative abundance of GTD-related Operational Taxonomic Units (OTUs) than root collars. More severe necrosis was correlated with a lower relative abundance of GTD-related OTUs based on isolation and metabarcoding analyses. Our results suggest that nurseries and therefore their plant production practices play a major role in determining the fungal and GTD-related fungal community in grapevine plants sold for planting. GTD variation across rootstocks and varieties could be explored as a venue for minimizing pathogen load in young plantations.
Project description:Plants belonging to <i>Vitis vinifera</i> varieties are usually grafted on different rootstocks to enhance the plant defenses against pathogens and increase productivity under harsh environmental conditions. Particularly, in Emilia-Romagna region (Italy), <i>Vitis vinifera</i> cultivar Lambrusco can be grafted on a hybrid of <i>V. berlandieri</i> × <i>V. riparia</i> (5BB) or <i>V. berlandieri</i> × <i>V. rupestris</i> (1103P). However, the latter shows potassium absorption problems, with a consequent reduction in grapevine production. Since it has recently been demonstrated that the rootstock has the potential to select for different microorganisms at the root-soil interface, here we hypothesized that the potassium deficiency of 1103P could be partially accounted for by the peculiarities of the rootstock microbiome. We thus employed 16S rRNA sequencing to compare root and rhizosphere microbiomes in plants of <i>V. vinifera</i> cultivar Lambrusco grafted on the two aforementioned rootstocks. According to our findings, 1103P shows a reduced diversity in root and rhizosphere microbiomes, including members of potassium-solubilizing microorganisms, possibly explaining the inadequate potassium absorption of this hybrid. Besides confirming the importance of the rootstock as a determinant of the composition of plant microbiomes, our data indicate the relevance of rootstock-selected microbiomes as possible regulators of potassium absorption by <i>V. vinifera</i>.
Project description:In this project a proteomic comparative analysis was used to analyse the changes induced by different nitrate availability in roots of M4 grapevine rootstock. Plants, previously adapted to hydroponic condition, were grown for 5 weeks, afterwards nitrogen was removed from the growing medium for eight days to obtain N-starved plants. After this period, the analysis compared plants maintained in absence of nitrogen with plants exposed to 10 mM nitrate for 30 hours.
Project description:Oxidative stress can arise when in vitro propagated plants developed under low light conditions are exposed to high light during transfer to ex vitro conditions. In such a situation, among the many potential stresses to which the transferred plant can be exposed, oxidative stress is commonly experienced, most likely brought about by absorption of light energy in excess of that required for very low levels of photosynthetic metabolism. In vitro propagated grapevine when transferred to ex vitro conditions with a 4 fold increase in PPFD shows an initial inhibition of PET accompanied by an accumulation of H2O2, suggesting a signal for the upregulation in gene expression and antioxidant enzyme activity, which peaked at 48h after transfer of in vitro grapevine to ex vitro growing conditions. When in vitro propagated plants are exposed to high light upon transfer to ex vitro oxidative stress symptoms occur. To determine whether the underlying pathways activated at the transfer of in vitro grapevine to ex vitro conditions reflect the processes occurring upon light stress we used microarrays. Leaves were harvested from in vitro grown plants immediately prior to transfer to ex vitro condition and 48h after transfer to compare gene expression before and after exposure of these plants to the high light conditions typical of ex vitro growth.