高粱耐盐相关基因HKT的功能及其耐盐作用机制研究

Sweet sorghum (Sorghum bicolor) is a C4 annual crop with a high biomass, and high tolerance to unfavorable conditions such as drought, salinity, poor nutrient supplies. Currently, sweet sorghum is considered the crop of best choice for the biofuels. However, sweet sorghum has a relative short history of breeding and many bottlenecks still need to be overcome before it is transformed into a dedicated biofuel crop. The high-affinity potassium transporter carrier protein family (HKT), is an important role in plant salt K + transport carrier protein superfamily. Little is known about the function of the HKT transporters in C4 plants which include economical and societal important food and bio-energy crops such as sorghum. Our current study shows that the glycophytic sorghum HKT gene family members have selective absorbing in the high salt stress, which is shown to play a critical role in regulating Na+/K+ balance under salt stress in the whole plant level. The SbHKTs had different express patterns in various tissues and upon treatments with external Na+ and K+ concentrations. Ectopic expression of SbHKT1;4 and SbHKT2;1 in K+ uptake deficient yeast strain altered the Na+ sensitivity and enhanced the K+ transport.In this project, we will take advantage of the GUS histochemical positioning or the immunogold labeling technique to determine the sorghum the four SbHKTs in gene expression patterns in sorghum different tissues,which may be involved in salt tolerance. At the same time,we also identify sbHKTs in representative sorghum inbred lines to explore the HKT allele mutants. The research is expected to show a possible existence of the halophytic mechanism of maintaining high K+-permeable transportation under Na+ stress in glycophytic C4 crops and provide novel insights in the use of sbHKTs variations for genetic improvement of sorghum and related crops.

甜高粱是 C4 植物，是发展生物质能源的最佳作物之一。为满足国家能源和粮食安全需求，急需培育适宜沿海滩涂、盐碱等边际土壤种植的甜高粱新品种。高亲和性钾离子转运载体蛋白家族(HKT)，是一类在植物耐盐方面有重要调控作用的K+转运载体蛋白超级家族。目前的研究表明，淡土植物高粱中的HKT基因家族中的成员已在拟南芥athkt1-1突变体、酵母缺钾互补菌株w△6的互补实验中证实，具有盐土植物中HKT基因在高Na+下选择吸收K+的离子转运特性，可能参与在高盐胁迫下，维持细胞及整株水平的K+含量的生理过程中起重要调控作用。我们希望通过GUS组织化学定位或免疫金标记等技术进一步确定高粱中4个SbHKTs基因在高粱不同组织、细胞中的表达模式，明晰其在抗盐中可能参与机制。同时利用遗传多样性丰富的高粱自交系，发掘HKT等位基因相关突变材料，为高粱及相关作物耐盐分子育种提供新思路。

由于不合理的耕作灌溉措施而引起土壤的次生盐渍化，使全球约占1/3 的农作物产量受到严重影响，盐害已成为亟待解决的问题。而如何提高作物耐盐能力，培育高耐盐作物应用于生产越来越受到广泛重视。高亲和性钾离子转运蛋白家族，是位于膜上的离子转运载体，在拟南芥、水稻、小麦等多种作物中的抗盐机制已经有很多研究报道。而高粱（Sorghum bicolour）属于淡土植物，相比水稻、小麦及玉米耐盐能力强，但高粱HKT蛋白在钠钾调控中的作用研究较少。本论文主要研究了高粱HKT基因家族在其钠钾离子平衡调控中的功能作用。高粱HKT蛋白家族有4个成员，基因启动子区存在11-15个参与应答干旱、高盐等非生物胁迫及逆境信号传导途径相关的顺式作用元件。SbHKTs蛋白均具有保守的功能结构域TrkH。烟草表皮细胞瞬时表达分析表明SbHKTs蛋白定位于细胞膜。SbHKTs基因具有组织表达特异性，SbHKT1;4在根、叶鞘及叶脉中表达量相对较高；SbHKT1;5基因主要在茎中表达；SbHKT1;3和 SbHKT2;1在地上部的表达明显高于根中的表达。在外界高Na+胁迫下，添加足量KCl的条件下，SbHKT1;4基因的表达水平在耐盐品种与盐敏感品种中存在显著差异。在耐盐品种2007中，SbHKT1;4基因仍然保持较高水平的上调表达，而在Ji2731中则无明显调控应答。高粱SbHKT1;4蛋白具有明显的Na+转运特性，并且具有高Na+离子条件下对K+的选择吸收来调控其体内钠钾平衡，从而提高耐盐能力。SbHKT1;4蛋白的高亲和性钾离子选择吸收特性可能是高粱较耐盐的一个重要原因，这也为我们下一步进行耐盐作物的改良及培育提供一种新思路。