VvSWEET10 Mediates Sugar Accumulation in Grapes

Abstract

Sugar accumulation is a critical event during grape berry ripening that determines the grape market values. Berry cells are highly dependent on sugar transporters to mediate cross-membrane transport. However, the role of sugar transporters in improving sugar accumulation in berries is not well established in grapes. Herein we report that a Sugars Will Eventually be Exported Transporter (SWEET), that is, VvSWEET10, was strongly expressed at the onset of ripening (véraison) and can improve grape sugar content. VvSWEET10 encodes a plasma membrane-localized transporter, and the heterologous expression of VvSWEET10 indicates that VvSWEET10 is a hexose-affinity transporter and has a broad spectrum of sugar transport functions. VvSWEET10 overexpression in grapevine calli and tomatoes increased the glucose, fructose, and total sugar levels significantly. The RNA sequencing results of grapevine transgenic calli showed that many sugar transporter genes and invertase genes were upregulated and suggest that VvSWEET10 may mediate sugar accumulation. These findings elucidated the role of VvSWEET10 in sugar accumulation and will be beneficial for the improvement of grape berry quality in the future.

Keywords: SWEET; grape; invertase; overexpression; sugar accumulation; véraison.

Figure 1

Phylogenetic and expression analyses of VvSWEET genes. (A) Amino acid sequences from grapes (Vitis vinifera), tomatoes (Solanum lycopersicum), and Arabidopsis. Evolutionary analyses were conducted in MEGA5 using the neighbor-joining method. VvSWEET10 (red) is a member of group III. (B) Heat map showing the expression level of VvSWEET genes in grapes. qRT-PCR analyzed the relative RNA transcription of VvSWEET in grapes, and the gene expression level in different organs showing the highest CT values was set to 1. The expression levels presented in the heat map were log2-based. The color scale represents transcript abundance in which blue to red represents a change in the expression level from low to high.

Figure 2

Developmental expression patterns of VvSWEET10 in grapes. The five developmental stages (i.e., E-L stages 27, 31, 35, 37, and 38) of the berry were used for qRT-PCR. The results are presented as means ± SD (n = 3).

Figure 3

Subcellular localization of VvSWEET10 proteins. The transient expression of VvSWEET10-GFP and free GFP (as a control) under the control of the 35S promoter in tobacco protoplasts. The fluorescent signal was imaged by confocal microscopy, GFP green fluorescence, and chlorophyll autofluorescence; bright-field images and merged images are shown. Scale bars = 10 μm.

Figure 4

Functional characterization of VvSWEET10. (A) Low sugar supply stimulates the expression of VvSWEET10 in grape berry suspension cells. Berry suspension cells were cultured in a sugar-free medium for a starvation condition, and the transcript levels of VvSWEET10 were analyzed within 24 h. (B) Complementation of yeast EBY.VW4000 with VvSWEET10; drop tests were used to observe the cell growth on SD (-Ura) media supplemented with 2% maltose, 2% sucrose, 2% fructose or 2% glucose. Cells were grown at 30 °C for 3 days.

Figure 5

VvSWEET10 overexpression increases sugar accumulation in grapevine calli. The sugar contents of VvSWEET10-overexpressed calli (OE) and the control (CK) were analyzed by HPLC. Results are shown as means ± SD (n = 2).

Figure 6

qRT-PCR validated the selection of sugar metabolism/transport DEGs. Results are means ± SD (n = 3).

Figure 7

Hexose levels of transgenic lines (i.e., OE11 and OE13) and wild-type control in developing tomato fruit. (A) Photos of tomato fruit from VvSWEET10-overexpressed and wild-type (WT) lines; fruit ripening stages were determined by the day after flowering (DAF). (B) The glucose content in developing VvSWEET10-overexpressed (OE) and WT tomato fruits. (C) The fructose content of developing VvSWEET10-overexpressed (OE) and WT tomato fruits. Results are expressed as means ± SD (n = 3).