Table_3_Constitutive Contribution by the Rice OsHKT1;4 Na+ Transporter to Xylem Sap Desalinization and Low Na+ Accumulation in Young Leaves Under Low as High External Na+ Conditions.pdf

HKT Na+ transporters correspond to major salt tolerance QTLs in different plant species and are targets of great interest for breeders. In rice, the HKT family is composed of seven or eight functional genes depending on cultivars. Three rice HKT genes, OsHKT1;1, OsHKT1;4 and OsHKT1;5, are known to contribute to salt tolerance by reducing Na+ accumulation in shoots upon salt stress. Here, we further investigate the mechanisms by which OsHKT1;4 contributes to this process and extend this analysis to the role of this transporter in plants in presence of low Na+ concentrations. By analyzing transgenic rice plants expressing a GUS reporter gene construct, we observed that OsHKT1;4 is mainly expressed in xylem parenchyma in both roots and leaves. Using mutant lines expressing artificial microRNA that selectively reduced OsHKT1;4 expression, the involvement of OsHKT1;4 in retrieving Na+ from the xylem sap in the roots upon salt stress was evidenced. Since OsHKT1;4 was found to be also well expressed in the roots in absence of salt stress, we extended the analysis of its role when plants were subjected to non-toxic Na+ conditions (0.5 and 5 mM). Our finding that the transporter, expressed in Xenopus oocytes, displayed a relatively high affinity for Na+, just above 1 mM, provided first support to the hypothesis that OsHKT1;4 could have a physiological role at low Na+ concentrations. We observed that progressive desalinization of the xylem sap along its ascent to the leaf blades still occurred in plants grown at submillimolar Na+ concentration, and that OsHKT1;4 was involved in reducing xylem sap Na+ concentration in roots in these conditions too. Its contribution to tissue desalinization from roots to young mature leaf blades appeared to be rather similar in the whole range of explored external Na+ concentrations, from submillimolar to salt stress conditions. Our data therefore indicate that HKT transporters can be involved in controlling Na+ translocation from roots to shoots in a much wider range of Na+ concentrations than previously thought. This asks questions about the roles of such a transporter-mediated maintaining of tissue Na+ content gradients in non-toxic conditions.

HKT型钠离子转运蛋白（HKT Na+ transporters）在不同植物物种中对应主要的耐盐数量性状位点（QTLs），是育种学家高度关注的研究靶标。在水稻中，HKT家族根据栽培品种的不同，由7至8个功能基因组成。已知3个水稻HKT基因——OsHKT1;1、OsHKT1;4和OsHKT1;5——可通过在盐胁迫下减少地上部的钠离子积累来提升植株耐盐性。本研究进一步解析了OsHKT1;4参与该过程的分子机制，并将分析拓展至该转运蛋白在低钠离子浓度环境下植物中的作用。通过分析表达β-葡萄糖苷酸酶（β-glucuronidase, GUS）报告基因构建体的转基因水稻植株，我们发现OsHKT1;4主要在根系和叶片的木质部薄壁组织中表达。利用人工微RNA（artificial microRNA）特异性降低OsHKT1;4表达的突变株系，我们证实了盐胁迫下OsHKT1;4参与从根系木质部汁液中回收钠离子的过程。由于我们发现OsHKT1;4在无盐胁迫的根系中同样高水平表达，因此我们进一步分析了其在植物处于非毒性钠离子浓度（0.5 mM和5 mM）条件下的功能。我们在爪蟾卵母细胞（Xenopus oocytes）中表达该转运蛋白后发现，其对钠离子具有较高的亲和力，解离常数略高于1 mM，这为“OsHKT1;4在低钠离子浓度下具有生理功能”这一假说提供了首个实验证据。我们观察到，即使在亚毫摩尔浓度钠离子环境中生长的植株，其木质部汁液在向叶片输送的过程中仍会逐步脱盐；且在该条件下，OsHKT1;4同样参与降低根系木质部汁液的钠离子浓度。从根系到幼嫩成熟叶片的组织脱盐过程中，OsHKT1;4的贡献在整个检测的外源钠离子浓度范围（从亚毫摩尔至盐胁迫条件）内均较为相似。因此，我们的研究数据表明，HKT型转运蛋白在调控钠离子从根系向地上部的转运方面，可作用的钠离子浓度范围远较此前认知更广。这一发现引发了关于在非毒性环境下，此类转运蛋白介导的组织钠离子含量梯度维持究竟发挥何种生理作用的疑问。