Large-scale proteomics of the cassava storage root and identification of a target gene to reduce post-harvest deterioration

Cassava (Manihot esculenta) is the most important root crop in the tropics, but rapid postharvest physiological deterioration (PPD) of the root is a major constraint to commercial cassava production. We established a reliable method for image-based PPD symptom quantification and used label-free quantitative proteomics to generate an extensive cassava root and PPD proteome. Over 2600 unique proteins were identified in the cassava root, and nearly 300 proteins showed significant abundance regulation during PPD. We identified protein abundance modulation in pathways associated with oxidative stress, phenylpropanoid biosynthesis (including scopoletin), the glutathione cycle, fatty acid α-oxidation, folate transformation, and the sulfate reduction II pathway. Increasing protein abundances and enzymatic activities of glutathione-associated enzymes, including glutathione reductases, glutaredoxins, and glutathione S-transferases, indicated a key role for ascorbate/glutathione cycles. Based on combined proteomics data, enzymatic activities, and lipid peroxidation assays, we identified glutathione peroxidase as a candidate for reducing PPD. Transgenic cassava overexpressing a cytosolic glutathione peroxidase in storage roots showed delayed PPD and reduced lipid peroxidation as well as decreased H2O2 accumulation. Quantitative proteomics data from ethene and phenylpropanoid pathways indicate additional gene candidates to further delay PPD. Cassava root proteomics data are available at www.pep2pro.ethz.ch for easy access and comparison with other proteomics data.

木薯（Manihot esculenta）是热带地区最重要的块根作物，但其块根快速发生的采后生理劣变（postharvest physiological deterioration，PPD）是制约木薯商业化生产的主要限制因素。我们建立了一套可靠的基于图像的PPD症状量化方法，并采用无标记定量蛋白质组学（label-free quantitative proteomics）构建了覆盖全面的木薯块根及PPD相关蛋白质组。在木薯块根中共鉴定到2600余种独特蛋白质，其中近300种蛋白质在PPD进程中呈现显著的丰度调控变化。我们鉴定到与氧化应激、苯丙烷类生物合成（包括东莨菪素（scopoletin））、谷胱甘肽循环、脂肪酸α-氧化、叶酸转化以及硫酸盐还原II通路相关的蛋白质丰度调控事件。谷胱甘肽相关酶类，包括谷胱甘肽还原酶、谷氧还蛋白与谷胱甘肽S-转移酶，其丰度与酶活性均显著升高，表明抗坏血酸-谷胱甘肽循环在PPD进程中发挥关键调控作用。基于整合的蛋白质组学数据、酶活性测定结果及脂质过氧化实验，我们将谷胱甘肽过氧化物酶鉴定为缓解PPD的候选靶标。在贮藏块根中过表达胞质型谷胱甘肽过氧化物酶的转基因木薯，表现出PPD延迟、脂质过氧化程度降低以及过氧化氢（H₂O₂）积累量减少的表型。来自乙烯（ethene）与苯丙烷类通路的定量蛋白质组学数据，进一步揭示了可延缓PPD的其他候选基因。木薯块根蛋白质组数据可通过网址www.pep2pro.ethz.ch公开获取，方便研究人员便捷访问并与其他蛋白质组数据开展比对分析。