木薯MeCBF1介导水孔蛋白MePIP2;7响应冷胁迫的机制研究

It is important to implement cold resistance breeding to promote northward expansion of cassava cultivation. Aquaporin plays a vital role in plant abiotic stresses through regulating the water transportation. Analyzing the regulation network will benefit to explore the internal mechanism under cold stress in cassava. Previous study showed that the expression of MePIP2;7 was repressed under cold condition, which was opposite to the expression of MeCBF1. The promoter of MePIP2;7 has the binding site of MeCBF1 and MeCBF1 could attenuate the promoter activity of MePIP2;7, which implied that MeCBF1 negatively regulated MePIP2;7 responding to cold stress. Overexpressing MeCBF1 transgenic plants had higher cold resistance, overexpressing MePIP2;7 transgenic plants were sensitive to cold stress but RNAi transgenic plants were resistant. The related traits of transgenic plants under cold treatment will be analyzed including the phenotype, physiologic and biochemical index, RNA-seq. Methods including the yeast one-hybrid will be used to verify the regulation function of MeCBF1 on MePIP2;7. Our objective is to elucidate the regulation mechanism of MeCBF1 on MePIP2;7 under cold stress in cassava. This work can provide the fundamental knowledge to understand the cold-sensitive phenotype and novel approaches for improving cold resistance in cassava.

抗寒分子育种是实现木薯北移种植的重要手段。水孔蛋白通过调控水分转运在植物的抗冷等逆境胁迫中发挥重要作用，解析其调控网络有助于揭示木薯不耐低温的调控机制。前期研究表明水孔蛋白MePIP2;7的表达受低温抑制，与MeCBF1的诱导表达成负相关。MePIP2;7启动子上具有MeCBF1的结合位点且活性受MeCBF1抑制，暗示MeCBF1负调控MePIP2;7响应冷胁迫。过表达MeCBF1木薯抗冷能力提高；而过表达MePIP2;7植株对冷敏感，RNAi植株抗冷能力增强。下一步将分析冷胁迫条件下转基因植株与野生型的表型、生理生化、RNA-seq等指标，开展酵母单杂等验证其调控模式，建立MeCBF1-MePIP2;7参与调控木薯冷胁迫响应的调控模式，为解析木薯冷敏感机制及抗寒改良提供理论依据。

木薯对低温敏感、耐贫瘠，其内在调控网络很大程度上是未知的。水孔蛋白MePIP2;7在冷处理基因芯片中被鉴定，其表达受冷、干旱、盐等非生物胁迫诱导，在根茎中表达量高，尤其在维管组织中高表达。低温下，PIP2;7-OE转基因植株离子渗透率和MDA的含量高于野生型，表现出对冷敏感。而PIP2;7-RNAi中含量较低，显示出抗冷表型。并且冷敏感导致H2O2在过表达植株叶片中富集。PIP2;7-RNAi植株基部成熟叶片表现出明显缺镁表型，ICP-MS证实了叶片中镁离子含量明显降低，PIP2;7-OE植株中镁离子富集。野生型和转基因植株缺镁处理进一步证实了MePIP2;7对镁离子平衡的调控作用。酵母双杂、BiFC和共定位表明MePIP2;7通过与MeMGT9互作实现对镁离子平衡的调控。这些结果表明MePIP2;7通过协调冷和镁离子的吸收调控木薯的生长发育。