Comparative Analysis of Cuticular Wax in Various Grape Cultivars During Berry Development and After Storage

Cuticular wax covering the surface of fleshy fruit is closely related to fruit glossiness, development, and post-harvest storage quality. However, the information about formation characteristics and molecular mechanisms of cuticular wax in grape berry is limited. In this study, crystal morphology, chemical composition, and gene expression of cuticular wax in grape berry were comprehensively investigated. Morphological analysis revealed high density of irregular lamellar crystal structures, which were correlated with the glaucous appearances of grape berry. Compositional analysis showed that the dominant wax compounds were triterpenoids, while the most diverse were alkanes. The amounts of triterpenoids declined sharply after véraison, while those of other compounds maintained nearly constant throughout the berry development. The amounts of each wax compounds varied among different cultivars and showed no correlation with berry skin colors. Moreover, the expression profiles of related genes were in accordance with the accumulation of wax compounds. Further investigation revealed the contribution of cuticular wax to the water preservation capacity during storage. These findings not only facilitate a better understanding of the characteristics of cuticular wax, but also shed light on the molecular basis of wax biosynthesis in grape.

Alternative splicing regulation appears to play a crucial role in grape berry development and is also potentially involved in adaptation responses to the environment

Background Alternative splicing (AS) produces transcript variants playing potential roles in proteome diversification and gene expression regulation. AS modulation is thus essential to respond to developmental and environmental stimuli. In grapevine, a better understanding of berry development is crucial for implementing breeding and viticultural strategies allowing adaptation to climate changes. Although profound changes in gene transcription have been shown to occur in the course of berry ripening, no detailed study on splicing modifications during this period has been published so far. We report here on the regulation of gene AS in developing berries of two grapevine (Vitis vinifera L.) varieties, Gewurztraminer (Gw) and Riesling (Ri), showing distinctive phenotypic characteristics. Using the software rMATS, the transcriptomes of berries at four developmental steps, from the green stage to mid-ripening, were analysed in pairwise comparisons between stages and varieties. Results A total of 305 differential AS (DAS) events, affecting 258 genes, were identified. Interestingly, 22% of these AS events had not been reported before. Among the 80 genes that underwent the most significant variations during ripening, 22 showed a similar splicing profile in Gw and Ri, which suggests their involvement in berry development. Conversely, 23 genes were subjected to splicing regulation in only one variety. In addition, the ratios of alternative isoforms were different in Gw and Ri for 35 other genes, without any change during ripening. This last result indicates substantial AS differences between the two varieties. Remarkably, 8 AS events were specific to one variety, due to the lack of a splice site in the other variety. Furthermore, the transcription rates of the genes affected by stage-dependent splicing regulation were mostly unchanged, identifying AS modulation as an independent way of shaping the transcriptome. Conclusions The analysis of AS profiles in grapevine varieties with contrasting phenotypes revealed some similarity in the regulation of several genes with developmental functions, suggesting their involvement in berry ripening. Additionally, many splicing differences were discovered between the two varieties, that could be linked to phenotypic specificities and distinct adaptive capacities. Together, these findings open perspectives for a better understanding of berry development and for the selection of grapevine genotypes adapted to climate change.

Identification of miRNAs-mediated seed and stone-hardening regulatory networks and their signal pathway of GA-induced seedless berries in grapevine (V. vinifera L.)

Background Stone-hardening stage is crucial to the development of grape seed and berry quality. A significant body of evidence supports the important roles of MicroRNAs in grape-berry development, but their specific molecular functions during grape stone-hardening stage remain unclear. Results Here, a total of 161 conserved and 85 species-specific miRNAs/miRNAs* (precursor) were identified in grape berries at stone-hardening stage using Solexa sequencing. Amongst them, 30 VvmiRNAs were stone-hardening stage-specific, whereas 52 exhibited differential expression profiles during berry development, potentially participating in the modulation of berry development as verified by their expression patterns. GO and KEGG pathway analysis showed that 13 VvmiRNAs might be involved in the regulation of embryo development, another 11 in lignin and cellulose biosynthesis, and also 28 in the modulation of hormone signaling, sugar, and proline metabolism. Furthermore, the target genes for 4 novel VvmiRNAs related to berry development were validated using RNA Ligase-Mediated (RLM)-RACE and Poly(A) Polymerase-Mediated (PPM)-RACE methods, and their cleavage mainly occurred at the 9th–11th sites from the 5′ ends of miRNAs at their binding regions. In view of the regulatory roles of GA in seed embryo development and stone-hardening in grape, we investigated the expression modes of VvmiRNAs and their target genes during GA-induced grape seedless-berry development, and we validated that GA induced the expression of VvmiR31-3p and VvmiR8-5p to negatively regulate the expression levels of CAFFEOYL COENZYME A-3-O-METHYLTRANSFERASE (VvCCoAOMT), and DDB1-CUL4 ASSOCIATED FACTOR1 (VvDCAF1). The series of changes might repress grape stone hardening and embryo development, which might be a potential key molecular mechanism in GA-induced grape seedless-berry development. Finally, a schematic model of miRNA-mediated grape seed and stone-hardening development was proposed. Conclusion This work identified 30 stone-hardening stage-specific VvmiRNAs and 52 significant differential expression ones, and preliminary interpreted the potential molecular mechanism of GA-induced grape parthenocarpy. GA negatively manipulate the expression of VvCCoAOMT and VvDCAF1 by up-regulation the expression of VvmiR31-3p and VvmiR8-5p, thereby repressing seed stone and embryo development to produce grape seedless berries.

The Effect of Water Deficit on Two Greek Vitis vinifera L. Cultivars: Physiology, Grape Composition and Gene Expression during Berry Development

Plants are exposed to numerous abiotic stresses. Drought is probably the most important of them and determines crop distribution around the world. Grapevine is considered to be a drought-resilient species, traditionally covering semiarid areas. Moreover, in the case of grapevine, moderate water deficit is known to improve the quality traits of grape berries and subsequently wine composition. However, against the backdrop of climate change, vines are expected to experience sustained water deficits which could be detrimental to both grape quality and yield. The influence of water deficit on two Greek Vitis vinifera L. cultivars, ‘Agiorgitiko’ and ‘Assyrtiko’, was investigated during the 2019 and 2020 vintages. Vine physiology measurements in irrigated and non-irrigated plants were performed at three time-points throughout berry development (green berry, veraison and harvest). Berry growth and composition were examined during ripening. According to the results, water deficit resulted in reduced berry size and increased levels of soluble sugars, total phenols and anthocyanins. The expression profile of specific genes, known to control grape color, aroma and flavor was altered by water availability during maturation in a cultivar-specific manner. In agreement with the increased concentration of phenolic compounds due to water deficit, genes of the phenylpropanoid pathway in the red-skinned Agiorgitiko exhibited higher expression levels and earlier up-regulation than in the white Assyrtiko. The expression profile of the other genes during maturation or in response to water deficit was depended on the vintage.

Transcriptomic analysis of temporal shifts in berry development between two grapevine cultivars of the Pinot family reveals potential genes controlling ripening time

Background Grapevine cultivars of the Pinot family represent clonally propagated mutants with major phenotypic and physiological differences, such as different colour or shifted ripening time, as well as changes in important viticultural traits. Specifically, the cultivars ‘Pinot Noir’ (PN) and ‘Pinot Noir Precoce’ (PNP, early ripening) flower at the same time, but vary in the beginning of berry ripening (veraison) and, consequently, harvest time. In addition to genotype, seasonal climatic conditions (i.e. high temperatures) also affect ripening times. To reveal possible regulatory genes that affect the timing of veraison onset, we investigated differences in gene expression profiles between PN and PNP throughout berry development with a closely meshed time series and over two separate years. Results The difference in the duration of berry formation between PN and PNP was quantified to be approximately two weeks under the growth conditions applied, using plant material with a proven PN and PNP clonal relationship. Clusters of co-expressed genes and differentially expressed genes (DEGs) were detected which reflect the shift in the timing of veraison onset. Functional annotation of these DEGs fit to observed phenotypic and physiological changes during berry development. In total, we observed 3,342 DEGs in 2014 and 2,745 DEGs in 2017 between PN and PNP, with 1,923 DEGs across both years. Among these, 388 DEGs were identified as veraison-specific and 12 were considered as berry ripening time regulatory candidates. The expression profiles revealed two candidate genes for ripening time control which we designated VviRTIC1 and VviRTIC2 (VIT_210s0071g01145 and VIT_200s0366g00020, respectively). These genes likely contribute the phenotypic differences observed between PN and PNP. Conclusions Many of the 1,923 DEGs show highly similar expression profiles in both cultivars if the patterns are aligned according to developmental stage. In our work, putative genes differentially expressed between PNP and PN which could control ripening time as well as veraison-specific genes were identified. We point out connections of these genes to molecular events during berry development and discuss potential candidate genes which may control ripening time. Two of these candidates were observed to be differentially expressed in the early berry development phase. Several down-regulated genes during berry ripening are annotated as auxin response factors / ARFs. Conceivably, general changes in auxin signaling may cause the earlier ripening phenotype of PNP.