热水处理诱导转录因子WRKY调控桃果实GSH合成代谢的抗冷性机制研究

Peach fruit are sensitive to low temperature and exhibit chilling injury when stored at an inapposite temperature or for a long period, significantly affecting fruit quality and storage ability. During previous study, the applicant found that hot water treatment (48 ℃, 10 min) could effectively enhance chilling resistance and affect GSH metabolism in peach fruit. WRKY transcript factors are recognized as response factors to chilling stress, which may be involved in the regulation of GSH synthesis and metabolism pathway. Based on above research results and background, the regulation mechanisms of hot water treatment on GSH synthesis and metabolism are to be analyzed at molecular level in this project. Transcriptomic analysis is to be performed to identify key WRKY transcript factors and key genes in GSH synthesis and metabolism pathway. Moreover, the correlation between key WRKY transcript factors and promotors of key genes in GSH synthesis and metabolism is also to be analyzed. In this study, the relationship between the regulation mechanisms of GSH synthesis and metabolism in response to hot water treatment and the enhancement of chilling resistance in peach fruit is to be clarified. The expected results from this study would provide a novel theoretic base for understanding the effect of hot water treatment on chilling injury in peach fruit and on WRKY transcript factors in regulation of GSH synthesis and metabolism. Meanwhile, this study would also provide a guide at theoretical and technical level for improving chilling resistance in peach fruit by using hot water treatment during production practice.

桃属于冷敏型果实，不适宜的低温或长期低温贮藏易引起桃果实发生冷害，严重影响桃果实的品质和贮藏期。申请人前期研究发现热水处理(48 ℃，10 min) 能够有效提高桃果实的抗冷性，影响桃果实中的谷胱甘肽(GSH)代谢。相关报道指出WRKY转录因子作为果实中的低温胁迫响应因子，可能参与调控GSH合成代谢途径。基于以上研究基础和背景，本项目拟重点从分子水平进一步研究热水处理对桃果实GSH合成代谢的作用机制，利用转录组学技术筛选出受热水处理调控的关键WRKY转录因子和GSH合成代谢途径的关键基因，分析WRKY转录因子与GSH合成代谢关键基因启动子的结合关系，阐明热水处理诱导WRKY转录因子调控桃果实GSH合成代谢的作用机制与其提高果实抗冷性的关系。通过本项目的实施，将为探究热水处理调控桃果实冷害的机理提供新的理论依据，并为今后在生产实践中采用热水处理提高桃果实抗冷性提供了理论及技术指导。

冷害（CI）是限制桃果实低温贮藏期和商品价值的重要因素。本项目基于转录组学分析出了桃果实低温冷害形成过程中的6条关键代谢通路，并筛选出了谷胱甘肽（GSH）代谢途径中的关键基因及参与响应低温胁迫信号的关键WRKY转录因子成员。通过不同热水处理（HWT）条件对比分析发现，52 ℃ 3 min的热水处理能够在贮藏后期维持桃果实较高抗氧化物质含量，抑制活性氧(ROS)的积累，减轻桃果实的CI症状并延缓桃果实的品质下降。通过进一步探究热水处理对GSH代谢通路和WRKY转录因子的调控机制，发现HWT处理可以诱导贮藏末期GSH含量，而在GSH代谢途径中，PpGS1表达水平的显著上调可能是影响GSH含量变化的关键因素；PpGR1、PpGPX1、PpGST1表达水平的变化可能是影响GSH-GSSG间转化水平的关键因素；而PpGST2受HWT处理的显著上调，可能是贮藏后期减轻桃果实ROS损伤的重要因素。通过酵母单杂交和瞬时过表达分析发现，PpWRKY33可以分别与PpGS1和PpGST2的启动子结合，并对PpGST2表达水平起到显著正调控作用，从而参与调控桃果实抗冷性的提升。此外，研究还发现内源茉莉酸合成和信号转导途径是响应茉莉酸甲酯处理减轻桃果实冷害中的关键通路。基于以上研究，PpWRKY33-PpGST2分子模块在抗冷性方面的功能验证及上游调控机制有待进一步探索，为挖掘抗冷新通路提供理论指导。