MaDof转录因子调控香蕉果实成熟的分子机制分析

The Dof is plant-specific transcription factor (TF), involving in a variety of plant biological processes, such as growth and development. However, little information is available regarding its role in regulating fruit ripening. Banana, as a climacteric fruit, is very susceptible to postharvest losses due to rapid softening. Thus, the ripening mechanism of banana fruit and its regulation is merited to study and is very important for extending postharvest life of banana fruit. In the present project, in order to reveal the role and mechanism of Dof TF in regulating banana fruit ripening, ripening-related MaDof TFs are firstly selecetd according to their expression patterns during banana fruit ripening, then their direct targets and interaction proteins were further identified., Moreover, the function of MaDofs involving in fruit ripening were confirmed by over-expressing ripening-related MaDofs in tomato. The results obtained in the project will not only expand new areas of function of Dof TF, but also give essential theoretical support about the regulation network of fruit ripening, as well as providing novel insight to improve the technology of maintaining fruit quality

Dof是一类植物特有的转录因子，参与调控植物生长发育等生物学过程，但目前有关Dof 转录因子在果实成熟中的作用及调控机制，尚未见报道。香蕉是典型的呼吸跃变型果实，对乙烯极其敏感，果实采后极易成熟、软化并腐烂，因此探讨香蕉果实采后成熟衰老及调控机制对于延长其货架期及供应期具有重要意义。 本项目在获得香蕉果实成熟相关的MaDof基因基础上，筛选、鉴定MaDof的下游靶基因及互作蛋白，并进一步在番茄中超表达与香蕉果实成熟相关的MaDof基因，鉴定MaDof的功能，以期阐明MaDof转录因子在采后香蕉果实成熟衰老过程中的作用及其分子机制。研究成果可以拓展Dof转录因子功能的新领域，同时为构建果实成熟的调控网络提供理论依据，对于改进果实贮藏的综合技术及维持果实品质，也提供了新思路。

Dof是一类植物特有的转录因子，参与调控植物生长发育等生物学过程，但目前有关Dof 转录因子在果实成熟中的作用及调控机制，尚未见报道。香蕉是典型的呼吸跃变型果实，对乙烯极其敏感，果实采后极易成熟、软化并腐烂，因此探讨香蕉果实采后成熟衰老及调控机制对于延长其货架期及供应期具有重要意义。本项目在获得香蕉果实成熟相关的MaDof基因基础上，筛选、鉴定MaDof的下游靶基因及互作蛋白，并进一步并分析其下游调控的靶基因。研究成果可以拓展Dof转录因子功能的新领域，对于改进果实贮藏的综合技术及维持果实品质，也提供了新思路。主要研究结果如下：.1. 从香蕉基因组中获得25个Dof基因。荧光定量分析表明，外源乙烯促进果肉中MaDof10/23/24/25的表达，其表达趋势随果实成熟及内源乙烯的释放明显增强，表明它们可能与香蕉果实成熟相关。.2. MaDof23与MaERF9互作参与调控香蕉果实成熟：酵母双杂交和双分子荧光互补（BiFC）实验证实，MaDof23与乙烯信号通路关键转录因子ERF类蛋白MaERF9相互作用。双荧光素酶报告基因检测（DLR）表明MaDof23能够显著抑制MaEXP1/2/3/5、MaXET7、MaPG1、MaPME3、MaPL2、MaGAL、MaCAT和MaPDC的启动子活性，而MaERF9则明显促进了MaEXP1/2/3/5、MaXET7、MaPG1、MaPME3、MaPL2、MaGAL、MaCAT 和MaPDC基因启动子的活性。共转后MaERF9则削弱了MaDof23对下游基因的抑制效果。这些结果说明，MaDof23可能通过与乙烯信号转导途径中的MaERF9互作形成蛋白复合体调控下游的成熟相关基因表达从而参与调控香蕉果实成熟。.3. MaDof24与MaCBF1和MaICE互作参与香蕉果实耐冷性调控。低温（7°C）逆境下，MaDof24在果皮中的表达则随着冷害进程而减弱，外源MeJA在提高香蕉果实的耐冷性同时也上调MaDof24的表达量。酵母双杂交和双分子荧光互补实验表明，MaDof24与植物抗冷途径关键因子MaCBF1和MaICE互作；此外，MaDof24 能够抑制 JA 合成途径相关基因 MaAOC2, MaLOX2, MaOPR3 的启动子活性，推测 MaDof24 可能通过介导 JA 合成而参与香蕉果实抗冷调控。