Synthesis of light-inducible and light-independent anthocyanins regulated by specific genes in grape 'Marselan' (V. vinifera L.)

Abstract

Anthocyanin is an important parameter for evaluating the quality of wine grapes. However, the effects of different light intensities on anthocyanin synthesis in grape berry skin and its regulation mechanisms are still unclear. In this experiment, clusters of wine grape cv. 'Marselan' were bagged using fruit bags with different light transmittance of 50%, 15%, 5%, and 0, designated as treatment A, B, C and D, respectively. Fruits that were not bagged were used as the control, designated as CK. The anthocyanin composition and concentration, as well as gene expression profiles in the berry skin were determined. The results showed that the degree of coloration of the berry skin reduced with the decrease of the light transmittance, and the veraison was postponed for 10 days in D when compared with the CK. Total anthocyanin concentration in the berry skin treated with D decreased by 51.50% compared with CK at the harvest stage. A total of 24 and 21 anthocyanins were detected in CK and D, respectively. Among them, Malvidin-3-O-coumaroylglucoside (trans), which showed a significant positive correlation with the total concentration of anthocyanins at the harvest stage (r = 0.775) and was not detected in D, was presumed to be light-induced anthocyanin. Other anthocyanins which were both synthesized in CK and D were considered to be light-independent anthocyanins. Among them, Malvidin-3-O-coumaroylglucoside (cis) and Malvidin-3-O-acetylglucoside were typical representatives. Remarkably, the synthesis of light-inducible anthocyanins and light-independent anthocyanins were regulated by different candidate structural genes involved in flavonoid biosynthesis pathway and members of MYB and bHLH transcription factors.

Keywords: Anthocyanin composition; Grapevine; LC-MS/MS; Light intensity; Transcriptome analysis.

Figure 1. Effects of different fruit bags on berry skin color.

Bags with light transmittance of 50%, 15%, 5% and 0 were performed at 45 days after flowering and designated as CK, A, B, C and D, respectively. Unbagged berries as a control . (A) Close-up views of wine grape cv. ‘Marselan’ berry fruit from different bagging treatments at S1, S2 and S3. (B) Changes of a nthocyanin concentration at the three development stages. Error bars represent the ±SE of three biological replicates, and the asterisks represent significant differences from the control, with P < 0.05 (*) or P < 0.01(∗∗).

Figure 2. Heatmap of the concentration of anthocyanins in different bagging treatments at S1, S2 and S3.

Figure 3. The proportion of individual anthocyanins in total anthocyanin concentration from CK (A) and D (B) treatments at S3.

Figure 4. Heatmap of expressed genes assigned to anthocyanins synthesis in different bagging treatments.

Colors indicate expression values of the genes. Expression values of ten libraries are presented as FPKM normalized log2 transformed counts.

Figure 5. Heatmap of MYB and basic helix-loop-helix (bHLH) transcription factors in different bagging treatments.

Colors indicate expression values of the genes. Expression values of ten libraries are presented as FPKM normalized log2 transformed counts.

Figure 6. qRT-PCR validation of four candidate genes related to anthocyanins synthesis from the different bagging treatments.

qRT-PCR validation of four candidate genes related to anthocyanins synthesis from the different bagging treatments (A, C, E and G) indicate relative expression of PAL(VIT_08s0040g01710), F3H(VIT_18s0001g14310), CHS(VIT_05s0136g00260) and F3′5′H (VIT_06s0009g02970) at S2, respectively. (B, D, F and H) indicate relative expression of PAL(VIT_08s0040g01710), F3H(VIT_18s0001g14310), CHS(VIT_05s0136g00260) and F3′5′H (VIT_06s0009g02970) at S3, respectively. The left y-axis indicates relative gene expression levels were determined by qRT-PCR and analyzed using 2^−ΔΔCT Method. The x-axis indicates different treatments. All qRT-PCR for each gene used three biological replicates, with three technical replicates per experiments. Error bars indicate ±SE, and the asterisks represent significant differences from the control, with P < 0.05(∗) or P < 0.01(∗∗).