MsNAC29转录因子正调控能源作物—荻耐盐性的分子机制

The need for salt tolerance biofuel plants increase due to both the development of biomass energy industry and the saline soil utilization. Miscanthus sacchariflora is a choiceness biomass energy plant originated in China. It is also a salt tolerance plant and has extensive future as a biofuel plant in saline soil. NAC（NAM、ATAF1、ATAF2 and CUC2）are a family of genes previously known to be involved in regulating various abiotic stress tolerances. It was found that a members of NAC, MsNAC29, may be uniquely involved in the regulating salt-tolerance, while the molecular mechanism of MsNAC29 remained unclear. Our recent study showed that MsNAC29 cloned from Miscanthus sacchariflora positive regulated salt tolerance. Based on these results, we will perform researches on two aspects, (1) determine the downstream pathways directly regulated by MsNAC29, which will be revealed through several strategies including transcriptomics, bioinformatics, promoter cloning, protein interactions with nucleic acid, function identification of target gene, in transgenic materials of MsNAC29 overexpression and RNAi suppression expression; (2)elucidation of the post-translational regulation mechanism of MsNAC29 through systematicly exploriation of the interacting proteins and in-depth expression analysis and salt-tolerance functional characterizaiton of the interacting proteins. Execution of this research can not only enrich the understanding of the molecular mechanism how MsNAC29 regulates salt-tolerance of plants, but also provide excellent genes for molecular breeding of salt-tolerance Miscanthus based on MsNAC29 genes.

生物质能源产业的发展和盐渍地开发利用均增加了对耐盐生物质能源植物的需求。荻是起源于我国的优良能源植物，具有较强的耐盐性，在盐渍化土壤上有较大的发展潜力。已有研究表明植物特有的转录因子NAC介导了不同的非生物胁迫抗性，其中MsNAC29成员参与了耐盐调控，但分子机制尚不清楚。本项目组已在耐盐较强的荻种质资源中发现MsNAC29可正向调控植物耐盐性。在此基础上，本项目拟从两个方面展开研究：（1）获得MsNAC29过量和RNAi抑制表达的荻转基因材料，结合转录组学、生物信息学、启动子克隆、蛋白核酸互作、靶基因功能鉴定等手段解析MsNAC29特异调控荻耐盐性的下游分子途径；（2）通过对MsNAC29互作蛋白的筛选、验证及其表达特性分析和遗传转化后的耐盐功能鉴定，阐明MsNAC29蛋白互作调控机制。本研究的实施将深化对MsNAC29调控植物耐盐分子机制的理解，也将为能源作物分子育种提供优异基因资源。

荻是起源于我国的优良能源植物，具有较强的耐盐性，在盐渍化土壤上有较大的发展潜力。已有研究表明植物特有的转录因子NAC介导了不同的非生物胁迫抗性，其中MsNAC29成员参与了耐盐调控，但分子机制尚不清楚。本项目已在耐盐较强的荻种质资源中发现MsNAC29可正向调控植物耐盐性。在此基础上，本项目展开以下研究并取得相应结果：.（1）通过对MsNAC29过量和敲除表达的水稻和荻转基因材料的耐盐鉴定，发现过表达MsNAC29基因能显著提高水稻和荻的耐盐性，而敲除同源基因后显著降低了水稻的耐盐性。结合转录组学、生物信息学、启动子克隆、蛋白核酸互作、靶基因功能鉴定等方法，初步筛选出受NAC29调控的直接下游功能基因DREB2A；并克隆荻MsDREB2A基因及其启动子，发现在MsDREB2A的启动子序列上有保守的NAC结合位点CGT（A/G），通过酵母单杂交、EMSA等实验进行了验证。构建植物超表达MsDREB2A载体并转化水稻，发现过表达转基因株系的耐盐性强于野生型材料。.（2）通过对MsNAC29互作蛋白的酵母双杂交筛选，获得MsNAC29的互作蛋白MsNAC48，经BiFC和pull-down验证，证明MsNAC48和MsNAC29蛋白发生真实互作，初步推断MsNAC29与MsNAC48形成异源二聚体来发挥功能。进一步构建pEarleyGate103-MsNAC48过表达载体并遗传转化，结果表明MsNAC48基因过表达能够维持过表达株系体内更高的K+含量并限制Na+在植株体内尤其是地上部分的积累，从而缓解NaCl对其光合作用的影响并维持过表达株系相对较高的生物量，提高了其耐盐性。.本项目的实施，将深化对MsNAC29调控植物耐盐分子机制的理解，也将为基于MsNAC29基因的能源作物分子育种提供优异的耐盐基因资源。