MaGPX介导膜脂氧化损伤修复的香蕉果实冷害的作用机制

Banana is one of the dominating fruits in China, and chilling injury occurs once banana fruit are preserved in low temperature accompanied by cell membrane damage and redox imbalance. Glutathione peroxidase (GPX) is an important redox-regulating protein to repair oxidative damage in response to abiotic and biotic stress. Our previous studies showed that the expressions of MaGPX in banana could be significantly induced by low temperature. However, the role of MaGPX in chilling injury of postharvest banana fruit is still unclear. We put forward a hypothesis that MaGPX could alleviate banana pericarp browning and inhibit starch degradation of fruit pulp via repairing lipid oxidation, thus delay chilling injury process. Therefore, we will take harvested banana as the experimental materails and room temperature, temperature close to chilling injury and that could induce chilling injury will be set as three treatment. The characteristics of transcriptional expression of MaGPX gene, the expression characteristics of MaGPX protein and the enzyme activity of MaGPX are studied to determine the MaGPX expression and regulation. Meanwhile, the target proteins of MaGPX are screened and the manner of protein-protein interaction will be verified. In addition, transgenic banana plants of MaGPX gene knock-out and overexpression were cultivated to further verify the function of MaGPX on chilling injury. In particular, the structure and content of oxidized lipids will be explored to illuminate the lipid oxidative degradation model in chilling injury of postharvest banana fruit. The results are expected to illustrate the role and regulation mechanism of MaGPX in fruit chilling injury, which may provide further evidence to elucidate the mechanism of chilling injury occurance and new approach to control chilling injury.

香蕉是我国主要热带水果之一，但果实低温贮藏易发生冷害，冷害伴随着膜系统损伤和氧化还原水平失衡。谷胱甘肽过氧化物酶（GPX）是生物体内重要的过氧化物分解酶，响应胁迫反应，参与修复膜脂过氧化损伤。申请人前期研究发现，低温显著诱导香蕉MaGPX的表达。目前，MaGPX在香蕉冷害过程的作用机制尚不明确。假设MaGPX通过修复膜脂过氧化损伤，抑制香蕉果皮褐变，抑制果肉淀粉降解。因此，本项目以采后香蕉果实为材料，设置常温、近冷害温度和冷害温度三个处理，通过分析MaGPX基因转录表达和蛋白表达特性，MaGPX酶活性，筛选MaGPX下游目标蛋白并验证其互作关系，构建基因敲除和超表达转基因香蕉株系，验证MaGPX在香蕉冷害过程的功能；同时鉴定MaGPX下游底物结构，明确膜脂质氧化降解模式。研究结果有助于明确MaGPX在果实冷害过程的功能及其调控机制，为果实采后冷害机理和控制新途径提供理论依据。

谷胱甘肽过氧化物酶（GPX）是生物体内重要的过氧化物分解酶，响应胁迫反应，参与修复膜脂过氧化损伤过程进而调控采后香蕉果实冷害进程。本研究克隆了香蕉GPX基因全长、生物信息学分析、并构建原核表达载体、诱导并纯化重组蛋白以及酶活测定等技术，从而揭示其生化特性，进一步研究发现，香蕉冷害过程中膜脂质降解与香蕉果皮褐变有明显的关系，低温储存会导致膜脂降解和过氧化，从而导致香蕉果实的冷害症状。本研究筛选出PC、PE、PA、DGDG和MGDG作为冷害过程的生物标志物。我们还证实了脂质相关酶、PLD 和 LOX 参与了与 ROS 产生相关的膜降解过程。此外，冷害的香蕉果实中PA显著积累，DGDG与MGDG比值升高，MDA含量增加。除此之外，我们还研究了膜脂代谢引起木质素累积情况，包括发现了柚子果实木质素积累过程中的糖类和能量代谢情况，发现柚果实在采后贮藏过程中，木质素累积与膜脂降解，糖类消耗，能量亏损密切相关。综上所述，我们的研究成果有望为降低水果品质劣变、确保水果安全的防治措施提供技术支撑。