Comparative Proteomics Reveals Differential Mechanism of Root Cold-resistance between Vitis. Riparia × V. Labrusca and Cabernet Sauvignon

Grapevines, containing large amounts of bioactive metabolites that offer health benefits, are widely cultivated around the world. The cold damage of growing outside with extreme low temperature during overwintering stage limits the expansion of production. Although the levels of morphological, biochemical and molecular in different Vitis species exposure to different temperatures have been investigated, differential expression of proteins in roots is still limited. Here, the roots of cold-resistant (Vitis. riparia × V. labrusca, T1) and cold-sensitive varieties (Cabernet Sauvignon, T3) at −4°C as well as of the former at −15°C (T2) were measured by iTRAQ-based proteomic analysis, expression levels of genes encoding candidate proteins were validated by qRT-PCR. The results showed that the root activity of cold-resistant variety was stronger than that of cold-sensitive variety, and it declined with the decrease of temperature. A total of 25 proteins were differentially co-expressed at T2 versus (vs) T1 and T1 vs T3, and these proteins were involved in stress response (e.g. DHN1, SHSPCP and USPCP), bio-signaling (e.g. PKCP, S/TPP and nsS/TP), metabolism (e.g. GluP, GluBE and PE), energy (e.g. AAC, AAACP and NADCP), and translation (e.g. rpL14, rpS21 and PPI). The relative expression levels of the candidate 13 genes were consistent with their fold-change values of proteins. The signature translation pattern for the roots at spatio-temporal treatments of varieties and temperatures provides insight into the differential mechanism of cold resistance of grapevines.

Features of identification grape varieties and clones of Western European origin

С помощью молекулярно-генетических и ампелографических методов проведена идентификация сортов винограда, относящихся к наиболее распространенным в мире техническим сортам западно-европейского происхождения. Генотипирование образцов проводилось с использованием 9-ядерных и 3-хлоропластных микросателлитных маркеров. На основании полученных профилей, по данным базы VIVC было установлено, что образец № 2 является сортом Каберне-Совиньон, образец № 4 - сортом Рисланер. Профиль образца № 1 совпадает с профилем сорта Мерло, за исключением разницы в двух парах нуклеотидов (п.н.) в одном аллеле локуса VVMD27, что можно объяснить редким случаем мутации в микросателлитной последовательности и не является достаточным основанием утверждать, что образец № 1 и Мерло являются разными сортами. Генетические профили образцов № 3 и № 6 соответствовали сортам сортогруппы Темпранильо, № 5 - сортам сортогруппы Рислинг рейнский, №7 - сортам сортогруппы Пино черный. Сорта в сортогруппах, полученные в результате соматических мутаций (связанных в основном с окраской ягод), имели одинаковый профиль. Принадлежность образцов к указанным сортам в сортогруппах была подтверждена ампелографическим методом. Использование для идентификации сортов в сортогруппах 6-9-ти SSR-маркеров в сочетании с ампелографическими методами позволяет получить достоверные результаты без удорожания работ. Однако дифференциация клонов и сортов, полученных в результате соматических мутаций, только SSR-маркерами потребует значительного увеличения их количества на 1-2 порядка либо использования высоковариабельных SSR-маркеров, таких как VRG ( Vitis riparia Götzhof). Таким образом, целесообразен более целенаправленный поиск полиморфизмов непосредственно в генах, отвечающих за определенные хозяйственно ценные признаки. В случае возникновения отличия в окраске ягод для дифференциации возможно использовать полиморфизм локуса гена VvMybA1, при изменении во вкусе и аромате ягод - в локусе гена VviDXS, при изменении лигнификации семян - в локусе гена VviAGL11, при повышении устойчивости к заболеваниям - в локусах соответствующих генов резистентности. The identification of grapes related to the most widespread wine varieties of West-European origin was carried out using molecular-genetic and ampelographic methods. Genotyping of samples was provided using 9- nuclear and 3-chloroplast microsatellite markers. Basing on the profiles obtained according to the VIVC database, it was established that Sample No. 2 is a ‘Cabernet-Sauvignon’ variety, and Sample No. 4 is a ‘Rieslaner’ variety. The profile of Sample No. 1 coincides with the ‘Merlot’ profile, except for the difference in 2 base pairs (bp) in one allele of the VVMD27 locus, which can be explained by a rare case of mutation in microsatellite sequence, and is not a sufficient reason to insist that Sample No. 1 and the ‘Merlot’ are different varieties. The genetic profiles of Samples No. 3 and No. 6 corresponded to the varieties of ‘Tempranillo’ group, No. 5 - to the varieties of ‘Rhein Riesling’ group, and No. 7 - to the varieties of ‘Pinot Noir’ group. The varieties of the groups, obtained as a result of somatic mutations (mainly associated with color of berries), had the same profile. The ampelographic method confirmed the origin of samples in the mentioned groups of varieties. Using of 6-9-SSR-markers in combination with ampelographic methods to identify the varieties of groups allows obtaining reliable results without increasing the cost of work. However, differentiation of clones and varieties in groups with only SSR-markers will require a significant increase in their number by 1-2 orders, or using of highly variable SSR-markers, such as VRG ( Vitis riparia Götzhof). Thus, a more targeted search for polymorphisms directly in genes responsible for certain economically valuable traits is advisable. In case of occurrence a difference in the color of berries, it is possible to use for differentiation the polymorphism of VvMybA1 gene locus, when flavor and aroma of berries change - to use the VviDXS gene locus, when seed lignification changes - the VviAGL11 gene locus, when disease resistance increases - the loci of the corresponding resistance genes.

Characterization of CRN-Like Genes From Plasmopara viticola: Searching for the Most Virulent Ones

Grapevine downy mildew is an insurmountable disease that endangers grapevine production and the wine industry worldwide. The causal agent of the disease is the obligate biotrophic oomycete Plasmopara viticola, for which the pathogenic mechanism remains largely unknown. Crinkling and necrosis proteins (CRN) are an ancient class of effectors utilized by pathogens, including oomycetes, that interfere with host plant defense reactions. In this study, 27 CRN-like genes were cloned from the P. viticola isolate YL genome, hereafter referred to as PvCRN genes, and characterized in silico and in planta. PvCRN genes in ‘YL’ share high sequence identities with their ortholog genes in the other three previously sequenced P. viticola isolates. Sequence divergence among the genes in the PvCRN family indicates that different PvCRN genes have different roles. Phylogenetic analysis of the PvCRN and the CRN proteins encoded by genes in the P. halstedii genome suggests that various functions might have been acquired by the CRN superfamily through independent evolution of Plasmopara species. When transiently expressed in plant cells, the PvCRN protein family shows multiple subcellular localizations. None of the cloned PvCRN proteins induced hypersensitive response (HR)-like cell death on the downy mildew-resistant grapevine Vitis riparia. This was in accordance with the result that most PvCRN proteins, except PvCRN11, failed to induce necrosis in Nicotiana benthamiana. Pattern-triggered immunity (PTI) induced by INF1 was hampered by several PvCRN proteins. In addition, 15 PvCRN proteins prevented Bax-induced plant programmed cell death. Among the cell death-suppressing members, PvCRN17, PvCRN20, and PvCRN23 were found to promote the susceptibility of N. benthamiana to Phytophthora capsici, which is a semi-biotrophic oomycete. Moreover, the nucleus-targeting member, PvCRN19, promoted the susceptibility of N. benthamiana to P. capsici. Therefore, these PvCRN proteins were estimated to be virulent effectors involved in the pathogenicity of P. viticola YL. Collectively, this study provides comprehensive insight into the CRN effector repertoire of P. viticola YL, which will help further elucidate the molecular mechanisms of the pathogenesis of grapevine downy mildew.

Habitat use by Veery (Catharus fuscescens) in southern Ontario

Veery (Catharus fuscescens) is a breeding migrant thrush that nests throughout much of the temperate forests within Canada. Habitat loss and degradation is thought to be responsible for a steady decline in Veery populations since 1970. We studied habitat characteristics of occupied Veery territories versus unoccupied adjacent areas in southern Ontario during the 2016 breeding season. Occupied territories were characterized as riparian deciduous forests dominated by ash (Fraxinus spp.), Black Cherry (Prunus serotina), and Red Maple (Acer rubrum) trees with an understorey of Balsam Fir (Abies balsamea) and ferns (order Polypodiales); the presence of fruit-producing plants such as Riverbank Grape (Vitis riparia) and Bunchberry (Cornus canadensis) also was important.

The Grapevine E3 Ubiquitin Ligase VriATL156 Confers Resistance against the Downy Mildew Pathogen Plasmopara viticola

Downy mildew, caused by Plasmopara viticola, is one of the most severe diseases of grapevine (Vitis vinifera L.). Genetic resistance is an effective and sustainable control strategy, but major resistance genes (encoding receptors for specific pathogen effectors) introgressed from wild Vitis species, although effective, may be non-durable because the pathogen can evolve to avoid specific recognition. Previous transcriptomic studies in the resistant species Vitis riparia highlighted the activation of signal transduction components during infection. The transfer of such components to V. vinifera might confer less specific and therefore more durable resistance. Here, we describe the generation of transgenic V. vinifera lines constitutively expressing the V. riparia E3 ubiquitin ligase gene VriATL156. Phenotypic and molecular analysis revealed that the transgenic plants were less susceptible to P. viticola than vector-only controls, confirming the role of this E3 ubiquitin ligase in the innate immune response. Two independent transgenic lines were selected for detailed analysis of the resistance phenotype by RNA-Seq and microscopy, revealing the profound reprogramming of transcription to achieve resistance that operates from the earliest stages of pathogen infection. The introduction of VriATL156 into elite grapevine cultivars could therefore provide an effective and sustainable control measure against downy mildew.

Physico-chemical characterization of wines produced by different rootstock and Vitis vinifera cv. Tannat clones in vineyards of subtropical climate region

Tannat wine trees are well characterized in Uruguay and the French region of Madiran for their high colour and phenolic concentrations. In addition to the cultivar, the rootstock, clone and region of production can influence the phenolic concentrations of wines. In this context, this study evaluated the rootstocks 'SO4' (Vitis berlandieri x Vitis riparia), 'Gravesac' ('161-49C' x '3309C') and '3309C' (Vitis riparia x Vitis rupestris) grafted with Tannat cultivar clones ('Californian', '944', '717', '398' and '794') to assess the physicochemical, phenolic and sensorial composition of the wine produced in the Campanha Gaúcha (RS) region, Southern Brazil, in a subtropical climate region. A vineyard planted in 2007 was used in this study (for 3 years during 2015, 2016 and 2017). The wine composition and the sensorial profile were evaluated as dependent variables. This study showed that the rootstocks and the Tannat clones did not influence the dependent variables evaluated and that the genetic materials and their combinations presented high oenological potential, providing wines with high alcohol content, colour and phenolic compound concentrations. This study suggests the diversification of rootstocks and clones as a way of increasing genetic variability, avoiding the cultivation of a single rootstock and clone.

Genome Sequences of Both Organelles of the Grapevine Rootstock Cultivar ‘Börner’

Genomic long reads of the interspecific grapevine rootstock cultivar ‘Börner’ (Vitis riparia GM183 × Vitis cinerea Arnold) were used to assemble its chloroplast and mitochondrion genome sequences. We annotated 133 chloroplast and 172 mitochondrial genes, including the RNA editing sites. The organelle genomes in ‘Börner’ were maternally inherited from Vitis riparia.

Genome sequences of both organelles of the grapevine rootstock cultivar 'Börner'

Genomic long reads of the interspecific grapevine rootstock cultivar 'Börner' (Vitis riparia GM183 x Vitis cinerea Arnold) were used to assemble its chloroplast and mitochondrion genome sequences. We annotated 133 chloroplast and 172 mitochondrial genes including the RNA-editing sites. The organellar genomes were maternally inherited to 'Börner' from Vitis riparia.