葡萄低温锻炼期蔗糖积累与ABA信号转导的分子调控机制

The northern of China is one of the world's best wine grapes producing areas, but the winter is cold tolerance, frequency of extreme cold weather, so it is often occured that the wine grapes suffered the invasion of cold winter and early spring of cold current, this has seriously affected the development of the wine industry characteristics.Therefore, improving the ability of the wine grape cold-resistence has very important significance for the production and development of wine grapes in northern China. After grape branches go through the cold acclimation period, there is a big difference in the degree of tolerance to low temperature. The studies suggest that sucrose accumulation and ABA signaling of molecules are the main material changes in the plant of cold-resistance, there was little about how they regulate grape hardiness has not been reported. Therefore, in this research, cold-resistence of Vitis amurensis is regarded as study material, with "sucrose accumulation - ABA signal- protein kinase - gene expression" as the main line, this research focus on cold-related genes of expression and changes of the key signal components in the signal transduction process which will be caused by the cold acclimation period， the growth, low-tempreture ( begin, mid and later) and dormancy, and sucrose accumulation and ABA signal transduction play critical role in this pathway. Clearing the critical period of cold-related genes expression in the grape and understanding the molecular mechanism of cold-resistence to lay a foundation for the application of grapes genomics function and transgenic technology making molecular improvement.

我国西北地区是世界酿酒葡萄的优势产区之一，但是该区冬季极端气温低，酿酒葡萄主栽品种难以露地越冬，埋土防寒是该区葡萄安全越冬的主要方式，严重制约该区葡萄酒现代产业的发展。因此，研究葡萄抗寒相关分子调控机理，加快葡萄抗寒育种进程具有重要意义。葡萄枝条经过低温锻炼可以显著提高其抗寒能力，而蔗糖积累与ABA信号传导是诱导植物产生抗寒力的重要途径，但他们是如何调控葡萄抗寒性表达的分子机理还不明确。本研究以抗寒性强的山葡萄为材料，以"蔗糖积累-ABA-蛋白激酶-基因表达"为主线，重点研究在生长期、低温锻炼期（低温锻炼初期、低温锻炼中期、低温锻炼后期）和深休眠期蔗糖的积累与ABA的信号转导会引起哪些抗寒相关基因的表达及其信号转导过程中的关键信号组分的变化。明确葡萄体内抗寒相关基因的表达关键时期，认识葡萄抗寒的分子调控机制。

冬季极端低温限制了我国西北地区酿酒葡萄主栽品种的露地越冬，利用埋土防寒是该区葡萄安全越冬的主要方式，但是由于其高成本及枝蔓损伤严重严重制约该区葡萄酒现代产业的发展。因此，研究葡萄抗寒相关分子调控机理，加快葡萄抗寒育种进程具有重要意义。本项目从葡萄低温锻炼期蔗糖积累、ABA合成途径以及转录水平基因表达等层面研究了在低温锻炼期蔗糖积累与ABA信号转导的分子调控机制主要取得以下成果。ABA信号在植物抗逆中扮演重要的角色，本研究利用葡萄基因组数据对其体内ABA信号途径相关的基因（SnRK2、PYL、PP2C、CIPK和LBD等）进行了克隆与表达分析，结果表明SnRK2、PYL、PP2C、CIPK和LBD基因家族能够明显的响应植物逆境信号。利用转录组学结合物质合成与积累在不同低温锻炼阶段对其ABA与蔗糖信号进行分析发现，在葡萄低温锻炼期共检测到差异表达基因8935个，其中上调的有4064（45.48%）个，下调的有4871（54.52%）个，这些基因在不同的低温表达阶段波动出现。通过转录组数据能够明确蔗糖相关基因的表达在葡萄低温锻炼阶段中共获得了关键的57个显著差异表达基因。这些基因包括了β-呋喃果糖苷酸酶基因，海藻糖磷酸合成酶基因，蔗糖合成酶，蔗糖磷酸合成酶，果糖激酶，葡聚糖分支酶，果胶酯酶/果胶酯酶抑制剂，葡聚糖磷酸化酶，多聚半乳糖醛酸酶，果胶酯酶，己糖激酶，内切葡糖糖苷酶，galacturonosyl转移酶，鸟苷二磷酸葡萄糖 4-差向异构酶，葡聚糖转移酶，纤维素合成酶的类似蛋白，木糖苷酶，β-葡萄糖苷酶，β-淀粉酶和葡萄糖磷酸腺苷转移酶大亚基1等基因；ABA信号途径中相关的PYL、SnRK2、PP2C、CIPK和LBD等基因的表达显著变化。并对转录组数据筛选到蔗糖信号途径相关基因VvBAM1和Vv AGPS进行了克隆与转化进行相应的功能验证，进一步明确了其在葡萄低温锻炼期蔗糖参与的分子调控机制。