Our data showed that the expression of flavonoids/PAs related-genes was highly increased in V9-berries at the third harvest time compared to V7- berries, resulting in the accumulation of more PAs in V9-berries.Developing and producing table grapes with high quality is of utmost importance for the success of grapevine breeding programs. Scarlet Royal table grape , variety , is one such success story, producing premium fruit quality and becoming one of the major red varieties in California. However, under certain unknown circumstances, the berry quality of Scarlet Royal grapes can be affected by undesirable astringent taste, which can negatively impact marketability and consumer acceptance. Research on the relationship between astringency and phenolic composition in table grapes is still scarce, especially on the transcriptomic level. In this study, we aimed to understand the molecular events involved in the development of berry astringency, which is a complex set of sensations resulting from the shrinking, drying, drawing, or puckering of the mouth epithelium . We focused on Scarlet Royal berries from two different vineyards with contrasting astringency and analyzed the changes in phenolic-related compounds at six different time points from veraison until the last harvesting time.
Our panel test revealed that the V9-berries were perceived as more astringent, blueberry packaging a characteristic that could be attributed to their elevated levels of tannins . Differences observed between the two vineyards under study indicate that V7 vines yield more compared to V9. This difference may be attributed to the lower cluster count in V9, a factor known to potentially contribute to astringency, as suggested by Cañon et al., 2014 in wine grapes. Additionally, petiole analysis revealed higher levels of nitrogen and potassium in V9 vines compared to V7. Vine nutrient levels contribute to the final berry quality at harvest . These factors may also contribute to the higher levels of tannins detected in V9 berries; however, further research is needed to confirm this theory. Nevertheless, the two vineyards are located in close proximity to each other, and weather data collected from the same station in the Delano area indicates similar conditions. Therefore, it is unlikely that astringency or higher phenolic compounds are induced by weather factors. In fact, several studies, mainly in wine, have pointed to PAs as a determining factor for astringency intensity . For example, Vidal et al. reported that the total amount of tannins is the most plausible factor for sensory astringency, with flavan-3-ols dimers, trimers, and non-galloylated tetramers contributing to the astringency sensation. The PAs are a group of oligomers and polymers of flavan-3-ols and are the naturally occurring and predominant type of tannins in grapes and wine . Another study on aronia berry juice confirmed PAs as the key astringent compounds using sensory evaluation and phenolic profile approaches along with in-vitro models .
The study found that PAs with higher degrees of polymerization were responsible for the strong astringent mouthfeel . The composition of phenolic substances, especially PAs, seems to play a crucial role in determining berry astringency, and further exploration of this relationship is warranted in fresh fruits of different species and cultivars. Understanding the molecular basis of astringency development in Scarlet Royal berries can provide valuable insights for improving grape breeding programs and enhancing the overall quality of table grapes.The relationship between astringency and the berry polyphenols content has not been explored yet in table grape at the molecular level. To the best of our knowledge, the present study provides the first transcriptome profiling along with the changes of polyphenols in grape berries of the same variety but having astringency diversity . The transcriptome profile of both V7-berries and V9-berries underlined the remarkable transcriptional shift during berry ripening at different vineyards . Commonly, berry transcriptome profiles may widely vary based on many factors, including genotypic variations among varieties/species and developmental stages , as well as environmental circumstances. In our case of study, stemmed differences from the developmental stages and genotypic variations were eliminated, and hence the difference of the vineyard locations was the main source of variance with 97% of variance . The identified DEGs output of V9-berries compared to V7-berries is highly explanatory, including polyphenolic-related genes that are robustly expressed and co-regulated with astringency development, particularly in the V9-berries. The enrichment of the up-regulated genes with BP GO terms related to the biosynthesis of secondary metabolites, phenylpropanoid, and nitrogen compound metabolic process , commensurate with the higher amounts of N found in V9- berries . In fact, not only the levels of N fertilization but also its different forms highly affected the composition of phenolic compounds in leaves and wine . However, the synergistic/antagonistic effects of other macro- and micro-nutrients should also be considered. Our results highlighted the negative impact of above-normal amounts of macro-nutrients, mainly N, and to a lesser extent P and K, on the desirable attributes of grape berries.
Otherwise, the positive effect of Ca, Mg, and Mn were achieved as their levels were maintained within the normal range . These data should be also seen in the light of the highly enriched BP GO terms in the down regulated genes . Particularly, those for hormonal signaling pathways such as auxin, abscisic acid, strigolactones, as well as the KEGG pathway for the carotenoid biosynthetic pathway .The transcriptome profiling identified the common and unique molecular events featuring the development of tannins/astringency in grape berries. It is well-documented that the synthesis of PAs in grapevines is achieved via three sequential pathways: the shikimate pathway, the phenylpropanoid pathway, and ultimately the flavonoid pathway . Our results revealed that the expression levels of flavonoids/PAs-related genes were highly induced in V9- berries at the third harvesting time compared to V7-berries. The 19 selected genes were involved in the three pathways: the shikimate pathway, phenylpropanoids pathway, and flavonoids pathway. The shikimate pathway is an alternative route to produce aromatic compounds, including phenylalanine, tyrosine, and tryptophan, which serve as precursors for various metabolites, such as phenolic compounds . The up-regulation of genes like chorismate synthase and chorismate mutase in V9-berries may lead to the accumulation of phenylalanine, which is a critical precursor for the phenylpropanoid pathway. The latter pathway is responsible for synthesizing several end products, including PAs, anthocyanins, lignin, lignans, hydroxycinnamic acid esters, and hydroxycinnamic acid amides . Under the conditions of the V9 vineyard, several PAs/flavonoids structural genes such as PAL, C4H, 4CL, CHS, CHI, F3H, LDOX, LAR, and ANR were induced in V9-berries, leading to the accumulation of PAs in the berries . This process is facilitated by GSTs and transported by multidrug and toxic compound extrusion transporters. The activation of the PAs biosynthetic pathway in V9- berries may lead to a reduction in the necessary substrates for anthocyanin synthesis, resulting in low red color intensity in V9- berries compared to V7-berries. Additionally, the accumulation of PAs is associated with the development of astringency taste in V9-berries. Our study provides valuable insights into the molecular events underlying astringency development in Scarlet Royal berries. By integrating transcriptome profiling with polyphenolic composition analysis, the research shed light on the co-regulation of genes involved in the shikimate, phenylpropanoid, and flavonoid pathways, leading to the synthesis of PAs and ultimately influencing astringency. The findings from this research have implications for grapevine breeding programs and the production of high-quality table grapes. Understanding the molecular mechanisms underlying astringency development can help breeders in selecting and developing grape varieties with desirable attributes. Additionally, the knowledge gained from this study can inform vineyard management practices, such as nutrient fertilization, to optimize polyphenolic composition and berry quality. In conclusion, the data presented in this study indicates that berry astringency is strongly correlated with a high tannin content, blueberry packaging box likely resulting from the activation of nineteen genes within the phenylpropanoid pathway. The activation of these genes shifts the flavonoid biosynthesis pathway towards proanthocyanins, leading to increased tannin accumulation in the berries. It appears that triggering these events is associated with nutritional imbalances and a lower number of clusters per vine, as confirmed by petiole nutrient levels and the observed lower berry soluble solids and higher titratable acidity levels. The identification of these genes holds significant value for table grape genetic improvement programs. The nutrient imbalance theory derived from this research could be applied worldwide to optimize grapevine fertilization programs. Furthermore, it paves the way for further research in this area, with a particular focus on vine nutrients, crop load management, and berry astringency, thereby contributing to advancements in the field of table grape cultivation.The cultivated garden strawberry , an allo-octoploid , has a unique natural and domestication history, originating as an interspecific hybrid between wild octoploid progenitor species approximately 300 years before present1 . The genomes of the progenitor species, Fragaria virginiana and Fragaria chiloensis, are the products of polyploid evolution: they were formed by the fusion of and interactions among genomes from four diploid progenitor species approximately 1 million years before present.
Whereas two of the diploid progenitor species have been identified, the other two diploid progenitor species have remained unknown. Moreover, the history of events leading to the formation of the octoploid lineage and the evolutionary dynamics among the four sub-genomes that restabilized cellular processes after ‘genomic shock’ in allopolyploids remain poorly understood. Here, we present what is, to our knowledge, the first chromosome-scale assembly of an octoploid strawberry genome, the identities of the extant diploid progenitor species of each sub-genome, and novel insights into the collective evolutionary processes involved in establishing a dominant sub-genome in this highly polyploid species. The Rosaceae are a large eudicot family including a rich diversity of crops with major economic importance worldwide, such as nuts , ornamentals , pome fruits , stone fruits , and berries. Strawberries are prized by consumers, largely because of their complex array of flavors and aromas. The genus Fragaria was named by the botanist Carl Linnaeus, on the basis of the Latin word ‘fragrans’, meaning ‘sweet scented’, describing its striking, highly aromatic fruit . A total of 22 wild species of Fragaria have been described, ranging from diploid to decaploid. The genus Fragaria is highly interfertile between and within ploidy levels, thus leading to the natural formation of higher-polyploid species. Polyploid events, also known as whole-genome duplications, have been an important recurrent process throughout the evolutionary history of eukaryotes and have probably contributed to novel and varied phenotypes. Polyploids are grouped into two main categories: autopolyploids and allopolyploids, involving either a single or multiple diploid progenitor species, respectively. Many crop species are allopolyploids, thus contributing to the emergence of important agronomic traits such as spinnable fibers in cotton, diversified morphotypes in Brassica, and varied aroma and flavor profiles in strawberry. Allopolyploids face the challenge of organizing distinct parental sub-genomes—each with a unique genetic and epigenetic makeup shaped by independent evolutionary histories—residing within a single nucleus. Previous studies have proposed, as part of the ‘sub-genome dominance’ hypothesis, that the establishment of a single dominant sub-genome may resolve various genetic conflicts in allopolyploids. However, understanding of the underlying mechanisms and ultimate consequences of sub-genome dominance remains largely incomplete. sub-genome-level analyses in most allopolyploid systems are greatly hindered by the inability to confidently assign parental gene copies to each sub-genome, owing to both large-scale chromosomal changes and homoeologous exchanges that shuffle and replace homoeologs among parental chromosomes. Octoploid strawberry still has a complete set of homoeologous chromosomes from all four parental sub-genomes, thus greatly simplifying homoeolog assignment. Furthermore, gene sequences from extant relatives of the diploid progenitor species, which probably still exist for octoploid strawberry, can be used to accurately assign homoeologs to each parental sub-genome. However, a highquality reference genome for the octoploid is needed to fully exploit strawberry as a model system for studying allopolyploidy as well as to provide a platform for identifying biologically and agriculturally important genes and applying genomic-enabled breeding approaches. The assembly of the octoploid strawberry genome, with an estimated genome size of 813.4 Mb, has been particularly challenging because of its high heterozygosity and ploidy level. For example, the most recently published version of the octoploid strawberry genome is highly fragmented, with more than 625,000 scaffolds, and largely incomplete, with less than 660 Mb assembled after removal of the numerous gaps. Thus, that version of the genome, owing to its overall highly fragmented nature, has not been a useful resource for genome-wide analyses including the discovery of molecular markers for breeding.Our goal was to obtain a high-quality reference genome for the Fragaria×ananassa cultivar ‘Camarosa’, one of the most historically important and widely grown strawberry cultivars worldwide.