盐胁迫下菌根共生维持酸枣钾、钠离子稳态平衡机制的研究

Chinese jujube (Ziziphus jujuba) is an important fruit tree, and mostly growing in the saline-alkaline soil conditions. Previous studies have revealed arbuscular mycorrhizal symbiosis has been shown to confer abiotic stress tolerance to sour jujube (Z. jujuba var. spinosa). However, the physiological and molecular mechanism of improving jujube Na+、K+ homeostasis by mycorrhizal symbiosis is still unknown. In this study, we perform a pot experiment to study the salt tolerance of sour jujube seedlings inoculated with mycorrhizal fungi (Rhizophagus irregularis) by addition of NaCl into the medium, and then investigate the physiological response of photosynthetic parameters, and Na+、K+ distribution. To determine the effect of mycorrhizal colonization on K+/Na+ homeostasis of sour jujube seedlings, we analyze salt-induced fluxes of Na+, K+, etc. of root tips using a scanning ion-selective electrode technique. In addition, the total RNA from fine roots under different treatments were extracted and then submitted to RNA-seq on Hiseq platform. We performed the comparative transcriptomic analysis among different treatments to revealed the differential gene expression profiles of sour jujube roots induced by mycorrhizal colonization under salt stress. We annotate and identify Na+/K+ transporters/channel proteins in Z. jujuba and R. irregularis, respectively. We then determine those members in mycorrhizal roots of sour jujube regulated by salt stress. Thereafter, the cell tissues harboring arbuscules were captured in the mycorrhizal tips using laser capture microdissection technology and then subjected to RNA extraction. We further analyze the gene expression pattern of Na+/K+ transporters/channel proteins belonging to sour jujube and arbuscular mycorrhizal fungi by qRT-PCR, respectively, and characterize their relations with the Na+, K+ homeostasis. Finally, we perform complementation assays on the Na+/ K+ channels and transporters using yeast mutant strainW6 defective in K+ uptake. In general, we try to characterize the molecular mechanism of mycorrhizal symbiosis improving jujube Na+、K+ homeostasis through analyzing the gene expression of Na+、K+ channels and transporters of both sour jujube and R. irregularis. This research would provide new insights into the mechanism of plant salt tolerance, and finally contributes to the jujube industry in the saline-alkaline area.

枣树主要栽培区土壤次生盐碱化严重，丛枝菌根（AM）可显著提高枣树的耐盐性，但盐胁迫下AM维持枣树Na+、K+稳态的机制尚不明确。本研究拟以枣树砧木-酸枣作为试材，采用盆栽试验，接种菌根菌（Rhizophagus irregularis）和添加NaCl，监测AM的Na+、K+等离子流，从动态的角度研究AM对盐离子选择吸收的调控；基于转录组水平，分析菌根共生对酸枣Na+、K+通道/转运蛋白基因表达的调控方式；采用激光显微切割技术结合qRT-PCR，研究AM含丛枝细胞中酸枣-菌根菌的离子通道/转运蛋白等基因对盐胁迫的响应方式；进一步采用酵母功能互补试验验证菌根菌 Na+/K+离子转运/通道蛋白的功能。本研究旨在从酸枣-菌根菌双方Na+、K+转运蛋白的协同表达方式来阐释菌根维持酸枣Na+、K+稳态平衡的机制，可丰富植物抗盐胁迫的理论，对保障盐碱地枣树可持续发展具有重要科学意义。

枣树是原产我国的重要的经济林树种，目前干旱和土壤盐渍化是枣主产区枣树生产的主要逆境限制因素。丛枝菌根（AM）真菌能与多数陆生植物形成互惠共生关系，促进宿主植物的生长，提高宿主植物在逆境条件下的适应能力。目前，菌根共生对枣树适应盐胁迫的生理及分子机制尚不明确。本研究以酸枣幼苗和异型根孢囊为材料，通过不同浓度 NaCl处理，研究菌根共生和盐胁迫对酸枣幼苗的生长、光合作用、激素含量和对离子稳态的影响，并通过比较转录组分析，揭示盐胁迫下菌根共生提高酸枣耐盐性的调控机制。.研究表明 酸枣幼苗具有一定的耐盐性，但超过100 mM NaCl时会抑制其生长，并且盐胁迫浓度越高、持续时间越长对生长的抑制更显。菌根共生显著提高了酸枣的生长和耐盐性，光合系统、抗氧化酶系统、内源激素代谢和离子稳态均受到菌根共生调。.研究发现菌根共生对Na+的吸收和转运具有促进作用，但对K+在叶中的积累效应更强，同时，菌根共生显著增强酸枣根系的H+外排和K+内流。结果表明质膜型H+-ATPase （ZjAHA7）和K+转运蛋白基因（ZjHAK2）的表达受到菌根共生特异诱后，通过酵母功能互补证明了ZjHAK2具有K+ 转运活性。表明菌根共生通过诱导ZjHAH7的表达启动H+的外排，提高酸枣对K+的吸收和转运是维持叶中K+/Na+离子稳态的主要途径。 .此外，通过菌根化的酸枣杨树和苜蓿植株的脂肪酸测定，证实了菌根共生提高叶片中脂肪酸含量的是一种保守的共生生理效应。对脂肪酸合成通路相关基因的表达模式进行分析，发现根中ZjACP2、ZjGRAS4、ZjFAE1.3、ZjFAE1等基因被菌根共生特异诱导表达或上调表达，而叶中没有菌根共生特异诱导和上调表达的基因。研究还发酸枣根中甘油三酯的增加主要来自菌根真菌中储存利用的部分，而叶中虽然脂肪酸含量显著提高，但并不以甘油三酯的形式进行储存。.研究为菌根共生提高木本植物对盐胁迫的适应性提供了一个全面的认识，阐明了菌根共生提高宿主植物离子稳态的机制，发现了菌根共生调控宿主植物脂肪酸代谢的新效应。