Comparative Omics Analysis of Four Grape Varieties and Exploration of Their Anthocyanin Synthesis Mechanisms

Abstract

Background: Vitis vinifera L. exhibits diverse varietal traits influencing fruit quality and stress tolerance. The summer black grape (Xiahei), known for its superior tolerance to abiotic stress and intense pigmentation, was hypothesized to possess distinct metabolic and genetic profiles, particularly in flavonoid and anthocyanin biosynthesis. This study aimed to elucidate the metabolic and molecular basis underlying these phenotypic traits by comparing carbohydrate composition and metabolomic and transcriptomic profiles of four grape varieties (summer black, flame seedless, black grape, and red milk). Methods: Grapes were consistently sampled five days after full maturity, and metabolites were analyzed using UPLC-MS/MS and GC-MS, while transcriptome analysis employed RNA sequencing followed by qRT-PCR validation. Results: The results demonstrated that carbohydrate content was similar among all grape varieties, whereas the summer black grape showed significantly higher levels of flavonoids, particularly anthocyanins such as delphinidin-3-O-glucoside, cyanidin-3-O-glucoside, and pelargonidin-3-O-glucoside. Metabolomic analyses revealed substantial enrichment of metabolites involved in flavonoid biosynthesis pathways, in agreement with transcriptomic data showing significant upregulation of key regulatory genes (CHS, DFR, and ANS) specific to anthocyanin biosynthesis. These findings suggest that the pronounced anthocyanin accumulation in summer black grape contributes to its distinctive dark pigmentation and enhanced resistance to abiotic stresses compared to other varieties. Conclusion: This study provides novel insights into the molecular and metabolic mechanisms driving anthocyanin accumulation in summer black grapes, which could inform future breeding programs aimed at improving grape resilience.

Keywords: Vitis vinifera L.; anthocyanin biosynthesis; gene expression; metabolomics; transcriptomics.

Figure 1

Characteristics of V. vinifera samples with different varieties: (A) summer black grape, (B) flame seedless grape, (C) black grape, (D) red milk grape.

Figure 2

Metabolomic and transcriptomic analysis of V. vinifera samples with different varieties. XH represents Xiahei grape; HYWH represents Huoyanwuhe grape; HT represents Heiti grape; HR represents Hongru grape. Principal component analysis of metabolome (A) and transcriptome (B). Cluster heatmap of V. vinifera metabolome (C) and transcriptome (D) with different varieties. Multiple volcano plots of differential metabolites (E) and genes (F). The part on the horizontal axis of the coordinate axis represents the upregulated metabolites or genes, while the part below the horizontal axis represents the downregulated metabolites or genes.

Figure 3

The quantitative histogram plot of carbohydrate contents. Summer black grape (XH), flame seedless grape (HYWH), black grape (HT), and red milk grape (HR). A one-way analysis of variance (ANOVA) was performed on the statistical data, revealing significant differences in the contents of various sugars among different groups. Notably, D-fructose (Fru) exhibited a high F-value of 49.3224 with a p-value of 0.0000, indicating a statistically significant difference. Similarly, sucrose (Suc) showed an F-value of 6.3374 and a p-value of 0.0000. D-galactose (Glu) also demonstrated a significant variation, with an F-value of 52.7433 and a p-value of 0.0000.

Figure 4

(A) The Kmeans analysis of metabolites. (B) The KEGG enrichment results of differential metabolites. (C) The Kmeans analysis of genes. (D) The KEGG enrichment results of differential genes.

Figure 5

The expression levels of structural genes and anthocyanin content involved in the anthocyanin biosynthesis pathway were examined in different colored flowers of V. vinifera. The heatmap was utilized to illustrate the expression patterns of these structural genes in four V. vinifera cultivars, and the pie chart represents the anthocyanin content, with colors ranging from blue-green to red, indicating low to high expression levels of the respective genes. The enzymes represented in this pathway are as follows: chalcone synthase (CHS), chalcone isomerase (CHI), flavanone 3-hydroxylase (F3H), flavonoid 3′-hydroxylase (F3′H), flavonoid 3′,5′-hydroxylase (F3′5′H), dihydroflavonol 4-reductase (DFR), anthocyanidin synthase (ANS), UDP-glucose:flavonoid 3-O-glucosyltransferase (UFGT), and glutathione S-transferase (GST).

Figure 6

The relative gene expression in the anthocyanin pathway. A one-way analysis of variance (ANOVA) was performed on the statistical data, revealing significant differences in the expression of regulated genes among different groups.