Table_7_Fruit From Two Kiwifruit Genotypes With Contrasting Softening Rates Show Differences in the Xyloglucan and Pectin Domains of the Cell Wall.docx

Fruit softening is controlled by hormonal and developmental cues, causing an upregulation of cell wall-associated enzymes that break down the complex sugar matrices in the cell wall. The regulation of this process is complex, with different genotypes demonstrating quite different softening patterns, even when they are closely related. Currently, little is known about the relationship between cell wall structure and the rate of fruit softening. To address this question, the softening of two Actinidia chinensis var. chinensis (kiwifruit) genotypes (a fast ‘AC-F’ and a slow ‘AC-S’ softening genotype) was examined using a range of compositional, biochemical, structural, and molecular techniques. Throughout softening, the cell wall structure of the two genotypes was fundamentally different at identical firmness stages. In the hemicellulose domain, xyloglucanase enzyme activity was higher in ‘AC-F’ at the firm unripe stage, a finding supported by differential expression of xyloglucan transglycosylase/hydrolase genes during softening. In the pectin domain, differences in pectin solubilization and location of methyl-esterified homogalacturonan in the cell wall between ‘AC-S’ and ‘AC-F’ were shown. Side chain analyses and molecular weight elution profiles of polyuronides and xyloglucans of cell wall extracts revealed fundamental differences between the genotypes, pointing towards a weakening of the structural integrity of cell walls in the fast softening ‘AC-F’ genotype even at the firm, unripe stage. As a consequence, the polysaccharides in the cell walls of ‘AC-F’ may be easier to access and hence more susceptible to enzymatic degradation than in ‘AC-S’, resulting in faster softening. Together these results suggest that the different rates of softening between ‘AC-F’ and ‘AC-S’ are not due to changes in enzyme activities alone, but that fundamental differences in the cell wall structure are likely to influence the rates of softening through differential modification and accessibility of specific cell wall polysaccharides during ripening.

果实软化受激素与发育信号调控，可诱导细胞壁相关酶（cell wall-associated enzymes）的表达上调，进而分解细胞壁中的复杂糖基质。该调控过程极为复杂：即使亲缘关系相近的基因型，其果实软化模式也存在显著差异。目前学界对细胞壁结构与果实软化速率之间的关联仍缺乏深入认知。为解答这一科学问题，本研究采用一系列组分分析、生化检测、结构解析与分子生物学技术，对两个中华猕猴桃（Actinidia chinensis var. chinensis，猕猴桃）基因型——快速软化型‘AC-F’与慢速软化型‘AC-S’的果实软化过程开展了分析。在整个软化进程中，两个基因型的细胞壁结构在相同果实硬度阶段即已呈现根本性差异。在半纤维素结构域（hemicellulose domain）中，未成熟硬果阶段的‘AC-F’材料的木葡聚糖酶（xyloglucanase）活性更高，这一结果得到了果实软化过程中木葡聚糖转糖基酶/水解酶（xyloglucan transglycosylase/hydrolase）基因差异表达数据的佐证。在果胶结构域中，‘AC-S’与‘AC-F’在果胶溶解释放以及细胞壁内甲基酯化同聚半乳糖醛酸（methyl-esterified homogalacturonan）的定位方面均存在差异。对细胞壁提取物中的聚醣醛酸（polyuronides）与木葡聚糖开展侧链分析与分子量洗脱谱分析后发现，两个基因型之间存在根本性差异，这表明即使处于未成熟硬果阶段，快速软化型‘AC-F’的细胞壁结构完整性已出现弱化。因此，与‘AC-S’相比，‘AC-F’细胞壁中的多糖更易被酶接触，进而更易发生酶解降解，最终导致果实软化速率更快。综合上述结果可知，‘AC-F’与‘AC-S’之间的软化速率差异并非仅由酶活性变化所致，细胞壁结构的根本性差异很可能通过调控果实成熟过程中特定细胞壁多糖的修饰程度与可及性，进而影响果实软化速率。