枳GRAS家族PtrPAT1基因调控甜菜碱积累响应低温的分子机制

低温胁迫是制约柑橘分布及产业发展的主要环境因子之一，适当的冷驯化可以增强植株的抗寒性，但极度寒冷会造成树体的死亡。甜菜碱（GB）作为一种渗透调节物质，参与响应植物的多种逆境胁迫，其具体的转录调控机制尚不清晰。前期研究发现外源施加甜菜碱明显提高了柑橘抗逆种质资源枳的耐寒性，通过构建枳低温表达谱鉴定出了枳甜菜碱合成关键酶基因PtrBADH-like，经酵母单杂交筛库分离出PtrMYC2和PtrPAT1两个上游转录因子，GRAS家族转录因子PtrPAT1强烈响应低温，且Y1H实验点对点验证了PtrPAT1与PtrBADH-l启动子的互作。本项目拟通过一系列生理生化实验进一步挖掘鉴定PtrPAT1-PtrBADH-l模块上游调控因子，旨在解析GA信号介导PtrPAT1调控枳冷诱发甜菜碱积累的分子机理，丰富并完善植物甜菜碱调控网络，同时为生产上应用甜菜碱增强柑橘抗寒性提供理论依据。

Low temperature is one of the main environmental factors that restrict the distribution and industrial development of citrus. Appropriate cold acclimation can enhance cold resistance of plants, but extreme freeze will cause the death of the trees. As an important osmotic protector in plants, glycine betaine (GB) is involved in response to various stress of plants, however, its specific transcriptional regulation mechanism and network are still unclear. Our previous study found that exogenous glycine betaine could significantly improve the cold tolerance of Poncirus trifoliata, a citrus cold-resistant germplasm. Based on low temperature transcriptome data of trifoliate orange, a key enzyme gene of GB biosynthesis was identified and named PtrBADH-like, and two transcription factors PtrMYC2 and PtrPAT1, which directly regulated PtrBADH-l, were isolated respectively through yeast one-hybrid screening library. The GRAS family transcription factor, PtrPAT1, was responded strongly to low temperature, and could binding to the promoter of PtrBADH-l via Y1H assay. This project intends to further identify the upstream regulatory factors of PTRPAT1-PtrBADH-L model by a series of physiological and biochemical experiments, aiming to uncover the molecular mechanism of GA signal mediated PtrPAT1 in regulating the accumulation of GB induced by low temperature, enrich the regulatory network of GB in plant, and provide theoretical basis for the application of GB to enhance the cold resistance of citrus in orange industry.