Characterization of major ripening events during softening in grape: turgor, sugar accumulation, abscisic acid metabolism, colour development, and their relationship with growth

Abstract

Along with sugar accumulation and colour development, softening is an important physiological change during the onset of ripening in fruits. In this work, we investigated the relationships among major events during softening in grape (Vitis vinifera L.) by quantifying elasticity in individual berries. In addition, we delayed softening and inhibited sugar accumulation using a mechanical growth-preventing treatment in order to identify processes that are sugar and/or growth dependent. Ripening processes commenced on various days after anthesis, but always at similarly low elasticity and turgor. Much of the softening occurred in the absence of other changes in berry physiology investigated here. Several genes encoding key cell wall-modifying enzymes were not up-regulated until softening was largely completed, suggesting softening may result primarily from decreases in turgor. Similarly, there was no decrease in solute potential, increase in sugar concentration, or colour development until elasticity and turgor were near minimum values, and these processes were inhibited when berry growth was prevented. Increases in abscisic acid occurred early during softening and in the absence of significant expression of the V. vinifera 9-cis-epoxycarotenoid dioxygenases. However, these increases were coincident with decreases in the abscisic acid catabolite diphasic acid, indicating that initial increases in abscisic acid may result from decreases in catabolism and/or exogenous import. These data suggest that softening, decreases in turgor, and increases in abscisic acid represent some of the earliest events during the onset of ripening. Later, physical growth, further increases in abscisic acid, and the accumulation of sugar are integral for colour development.

Keywords: Anthocyanins; Vitis vinifera L.; cell wall; elasticity; firmness; fruit development.

Fig. 1.

Berry diameter, weight, and total soluble solids (TSS). (A, B) Berry diameter and weight in Control and Box berries (n = 20–100). (C) Degrees Brix in Control (n = 6–9), Box (n = 6), and Box Control (n = 3 at 85 DAA and n = 1 at 90 DAA). (D) Anthocyanin accumulation in berry skins as a function of DAA in Control and Box (n = 3). Boxes were removed at 79 DAA (arrows) and Box Control remained boxed on the cluster. Bars represent ± SE and different letters signify significant differences (Tukey’s HSD, P < 0.05).

Fig. 2.

Cell turgor (P) and berry elasticity. (A) Cell turgor through berry development in Control, Box, and Box Control (n = 12–42). (B) Elasticity in Control and Box berries (n = 16). Boxes were removed at 79 DAA (arrow) and Box Control remained boxed on the cluster. Bars represent ± SE and different letters signify significant differences (Tukey’s HSD, P < 0.05). (C) Relationship between elasticity and cell turgor. Points are means of 2–6 berries and bars are ±SE.

Fig. 3.

Expression of key cell wall metabolism enzymes in skin and flesh as a function of elasticity. Expression of VviExp1 (A), VviExp2 (B), VviPL (C), VviXTH (D), and VviPME (E) in skin and flesh as a function of the elasticity in Control and Box berries. Each point represents the level of expression in the skin or flesh of a single berry at the corresponding measured E. Points are coloured according to berry skin colour.

Fig. 4.

Relationships between Ψ_s ^Berry, elasticity, and sugar concentration. (A) Ψ_s ^Berry in Control and Box berries. Boxes were removed at 79 DAA (arrow). Bars represent ±SE and different letters signify significant differences (n = 3–9 for Control and n = 3 for Box; Tukey’s HSD, P < 0.05). (B) The relationship of berry Ψ_s ^Berry to the elasticity of Control and Box berries. (C) Developmental changes in berry sugar concentration (glucose + fructose) in Control and Box berries. Boxes were removed at 79 DAA (arrow). Bars represent ±SE and different letters signify significant differences (n = 3–9 for Control and n = 3 for Box; Tukey’s HSD, P < 0.05). (D) The relationship between the elasticity and sugar concentration in Control and Box berries.

Fig. 5.

Expression of key sugar metabolism enzymes in skin and flesh as a function of elasticity. Expression of VviINV (A), VviHT1 (B), VviHT2 (C), and VviHT3 (D) in skin and flesh as a function of the elasticity of Control and Box berries. Each point represents the level of expression in the skin or flesh of a single berry at the corresponding measured E. Points are coloured according to berry skin colour.

Fig. 6.

ABA metabolism. Concentration of ABA species, including active ABA (A), its inactive glucose ester conjugate ABA-GE (B), and the committed catabolites 7′-OH ABA (C), diphasic acid (D), and neophasic acid (E), as a function of elasticity. Box treatment data include both boxed and unboxed berries because treatment had no significant effect. Each point represents the level of metabolite in the skin or flesh a single berry. Points are coloured according to berry skin colour.

Fig. 7.

Relationships among VviNCED expression, ABA concentration, and sugars. (A) VviNCED expression as a function of elasticity. (B) The relationship between VviNCED expression and ABA concentration. (C) The relationship between sugar and ABA concentrations. Box treatment data include both boxed and unboxed berries because treatment had no significant effect. Each point represents the level of metabolite or expression in the skin or flesh of a single berry. The colour of the berries comprising each pool is shown.

Fig. 8.

Summary of the relationship between decreases in elasticity (i.e. softening), sugar accumulation, increases in ABA, and anthocyanin accumulation (i.e. colour development). Points represent data for E, sugar concentration, and anthocyanin concentration measured in individual berries. Average ABA concentrations for selected elasticity pools are represented in the columns. Three sequential steps can be identified: 1 (light green) – E (and P) decreases, small increase in ABA concentration, no change in sugar concentration; 2 (dark green) – sugar concentration increases, large increase in ABA concentration, E (and P) decreases further; 3 (purple) – colour development begins.