DREB1A介导褪黑素调控番茄耐盐性的信号途径和分子机制

Melatonin is a widely concerned plant regulator in recent years, the preliminary experiment showed that transcription factor DREB1A was involved in melatonin alleviating salt stress on tomato, but the physiological mechanism is not clear. In our previous study, it was found that the physiological function of melatonin was closely related with NO, ABA and H2O2 , Based on this, in the present study, both exogenous melatonin treatment and endogenous melatonin regulation were used to combine with ABA, NO and H2O2 synthesis inhibitors or scavengers, and the effect of different combined treatments on DREB1A expression in tomato plants under salt stress will be studied. RNAi technology is used to inhibit the expression of DREB1A, and then the effect of DREB1A on ABA, NO and H2O2 will be investigated, based on this, the signal pathway of DREB1A involved in melatonin regulating tomato tolerance to salt stress is analyzed. yeast two-hybrid technology, bimolecular fluorescence complementation and Pull-down are combine to screening proteins interacting with DREB1A, and the relationships between the interacted protein with DREB1A and melatonin, ABA, NO, H2O2 are further investigated, and the molecular mechanism of DREB1A involved in melatonin regulating tomato tolerance to salt stress will be made clear.The expected results will not only enrich the physiological pathway of melatonin function involved in stress tolerance, but also provide the theoretical basis for the application of DREB1A in tomato tolerance to salt stress.

褪黑素是近几年广受关注的植物调节物质，初步试验表明转录因子DREB1A参与了褪黑素缓解番茄盐胁迫过程，但其生理机制尚不清楚。我们前期发现褪黑素生理功能与NO、ABA和H2O2密切相关，在此基础上，本项目拟采用外源褪黑素处理和内源褪黑素调控相结合的方法，与ABA、NO和H2O2合成抑制剂或清除剂进行组合处理，研究盐胁迫下不同处理组合对番茄DREB1A表达的影响，采用RNAi技术抑制DREB1A表达，研究DREB1A对ABA、NO和H2O2的影响，解析DREB1A介导褪黑素调控番茄耐盐性的信号途径；联合应用酵母双杂交、双分子荧光互补和Pull-down技术筛选DREB1A互作蛋白，研究互作蛋白与DREB1A、褪黑素、ABA、NO和H2O2的关系，探明DREB1A参与褪黑素调控番茄耐盐信号传递的分子机制。预期结果既可丰富褪黑素抗逆功能的生理途径，又可为DREB1A在番茄耐盐中的应用奠定理论基础。

首先利用转录组系统解析了褪黑素缓解番茄盐碱胁迫的生理机制，在此基础上，挖掘出DREB1A是介导褪黑素调控番茄耐盐碱的重要转录因子，进一步系统解析了DREB1A调控的IAA3表达及其一系列生理代谢过程是褪黑素提高番茄耐盐碱的重要生理途径。进而克隆了褪黑素合成关键基因SlCOMT1，发现过表达SlCOMT1可以通过调控离子平衡、水分代谢、物质转化等过程促进番茄发芽期及植株对盐胁迫和干旱胁迫的耐性，并解析了外源褪黑素缓解番茄缺钙胁迫的生理效应。为褪黑素在提高番茄对盐碱、干旱等非生物胁迫中的应用提供了理论依据。采用酵母双杂交、BiFC筛选获得了SlDREB1A互作的SlMORF2和SlMBD15，并发现盐胁迫能显著诱导SlMORF2和SlMBD15的表达，与SlDREB1A在盐胁迫下的表达模式类似，为进一步深入解析SlDREB1A介导褪黑素调控番茄耐盐性的分子机制奠定了基础。另外，研究发现SlGSNOR是番茄耐非生物胁迫的正调控因子，在盐碱、高温等非生物胁迫下，SlGSNOR的下调导致根部的NO过量积累，触发了MAPK和乙烯信号通路中蛋白的表达，同时增加了乙烯的释放，继而解析了SlGSNOR介导的番茄非生物胁迫响应在很大程度上依赖于SlMAPK3-SlACO1级联信号。在研究了调控番茄耐盐碱能力之后，我们又研究了盐胁迫对番茄品质的影响，发现适度盐胁迫可以显著提高番茄果实中糖等营养成分的积累，通过多组学分析，鉴定出番茄果实成熟过程和盐处理下成熟果实中的差异表达蛋白以及和N-糖基化蛋白，发现这些蛋白主要参与淀粉和蔗糖代谢以及半乳糖代谢等重要过程，进一步从蛋白表达和N-糖基化角度揭示盐处理调控番茄品质形成的生理机制奠定了基础。