A high-affinity potassium transporter (MeHKT1) from cassava (Manihot esculenta) negatively regulates the response of transgenic Arabidopsis to salt stress.

BACKGROUND: High-affinity potassium transporters (HKTs) are crucial in facilitating potassium uptake by plants. Many types of HKTs confer salt tolerance to plants through regulating K+ and Na+ homeostasis under salinity stress. However, their specific functions in cassava (Manihot esculenta) remain unclear. RESULTS: Herein, an HKT gene (MeHKT1) was cloned from cassava, and its expression is triggered by exposure to salt stress. The expression of a plasma membrane-bound protein functions as transporter to rescue a low potassium (K+) sensitivity of yeast mutant strain, but the complementation of MeHKT1 is inhibited by NaCl treatment. Under low K+ stress, transgenic Arabidopsis with MeHKT1 exhibits improved growth due to increasing shoot K+ content. In contrast, transgenic Arabidopsis accumulates more Na+ under salt stress than wild-type (WT) plants. Nevertheless, the differences in K+ content between transgenic and WT plants are not significant. Additionally, Arabidopsis expressing MeHKT1 displayed a stronger salt-sensitive phenotype. CONCLUSION: These results suggest that under low K+ condition, MeHKT1 functions as a potassium transporter. In contrast, MeHKT1 mainly transports Na+ into cells under salt stress condition and negatively regulates the response of transgenic Arabidopsis to salt stress. Our results provide a reference for further research on the function of MeHKT1, and provide a basis for further application of MeHKT1 in cassava by molecular biological means.

Potassium transporter OsHAK9 regulates seed germination under salt stress by preventing gibberellin degradation through mediating OsGA2ox7 in rice.

Soil salinity has a major impact on rice seed germination, severely limiting rice production. Herein, a rice germination defective mutant under salt stress (gdss) was identified by using chemical mutagenesis. The GDSS gene was detected via MutMap and shown to encode potassium transporter OsHAK9. Phenotypic analysis of complementation and mutant lines demonstrated that OsHAK9 was an essential regulator responsible for seed germination under salt stress. OsHAK9 is highly expressed in germinating seed embryos. Ion contents and non-invasive micro-test technology results showed that OsHAK9 restricted K+ efflux in salt-exposed germinating seeds for the balance of K+/Na+. Disruption of OsHAK9 significantly reduced gibberellin 4 (GA4) levels, and the germination defective phenotype of oshak9a was partly rescued by exogenous GA3 treatment under salt stress. RNA sequencing (RNA-seq) and real-time quantitative polymerase chain reaction analysis demonstrated that the disruption of OsHAK9 improved the GA-deactivated gene OsGA2ox7 expression in germinating seeds under salt stress, and the expression of OsGA2ox7 was significantly inhibited by salt stress. Null mutants of OsGA2ox7 created using clustered, regularly interspaced, short palindromic repeat (CRISPR)/CRISPR-associated nuclease 9 approach displayed a dramatically increased seed germination ability under salt stress. Overall, our results highlight that OsHAK9 regulates seed germination performance under salt stress involving preventing GA degradation by mediating OsGA2ox7, which provides a novel clue about the relationship between GA and OsHAKs in rice.

Ethylene-induced potassium transporter AcKUP2 gene is involved in kiwifruit postharvest ripening.

BACKGROUND: Potassium (K) is important in the regulation of plant growth and development. It is the most abundant mineral element in kiwifruit, and its content increases during fruit ripening. However, how K+ transporter works in kiwifruit postharvest maturation is not yet clear. RESULTS: Here, 12 K+ transporter KT/HAK/KUP genes, AcKUP1 ~ AcKUP12, were isolated from kiwifruit, and their phylogeny, genomic structure, chromosomal location, protein properties, conserved motifs and cis-acting elements were analysed. Transcription analysis revealed that AcKUP2 expression increased rapidly and was maintained at a high level during postharvest maturation, consistent with the trend of K content; AcKUP2 expression was induced by ethylene, suggesting that AcKUP2 might play a role in ripening. Fluorescence microscopy showed that AcKUP2 is localised in the plasma membrane. Cis-elements, including DER or ethylene response element (ERE) responsive to ethylene, were found in the AcKUP2 promoter sequence, and ethylene significantly enhanced the AcKUP2 promoter activity. Furthermore, we verified that AcERF15, an ethylene response factor, directly binds to the AcKUP2 promoter to promote its expression. Thus, AcKUP2 may be an important potassium transporter gene which involved in ethylene-regulated kiwifruit postharvest ripening. CONCLUSIONS: Therefore, our study establishes the first genome-wide analysis of the kiwifruit KT/HAK/KUP gene family and provides valuable information for understanding the function of the KT/HAK/KUP genes in kiwifruit postharvest ripening.

Rice OsHAK5 is a major potassium transporter that functions in potassium uptake with high specificity but contributes less to cesium uptake.

Cesium (Cs) in the environment is primarily absorbed by a potassium (K) transporter. OsHAK5 is a KT/HAK/KUP family K-transporter showing a high affinity for K. We created cultured rice cells whose OsHAK5 was knocked down by RNAi (named KD). In the medium containing 1.0 m m and less K, the growth of KD was significantly suppressed, suggesting that OsHAK5 greatly contributed to K absorption under limited K conditions. Although Cs suppressed the growth of KD and WT, stronger inhibition was observed on KD. Both KD and WT accumulated similar amounts of Cs when they were cultured in a medium containing Cs, whereas lower amounts of K were detected in KD. These results suggest that OsHAK5 was less involved in the absorption of Cs, although it was essential to K absorption under limited K conditions. In contrast, this means that another transporter may contribute to cesium uptake in rice.

Plant HAK/KUP/KT K+ transporters: Function and regulation.

The HAK/KUP/KT family of potassium (K+) transporters belongs to the amino acid-polyamine-organocation (APC) superfamily of carriers for secondary active transport and has been widely associated with K+ transport across membranes in bacteria, fungi, and plants. The plant genome contains large number of HAK/KUP/KT transporters, and they show the diverse roles in K+ uptake and translocation, salt tolerance and osmotic potential regulation, as well as in controlling root morphology and shoot phenotyping. Recently, significant progress has been achieved towards uncovering the regulatory mechanisms of HAK/KUP/KT transporters at both transcriptional and post-translational levels. Most of the HAK/KUP/KT genes were regulated at transcriptional level, and such regulation may contribute to the alteration of root cell membrane potential by different growth conditions. At least six transcription factors have been identified as positive or negative regulators of HAK/KUP/KT gene expression in responding to external K+ supply. The HAK/KUP/KT transporter proteins can be phosphorylated by CIPK-CBL complexes for activating their function in K+ uptake and probably signaling. Nevertheless, it is still not known if HAK/KUP/KT transporters are involved in K+-sensing and K+-compartmentation in plant cells. Some orthologues of the HAK/KUP/KT transporters from different species show varied physiological functions and some plant species lack an entire sub-clade of HAK/KUT/KT transporters. We are still a long way from unraveling the molecular mechanism of HAK/KUP/KT involved in K+-sensing and signaling pathways in plants.

Genome-Wide Survey and Expression Analysis of the KT/HAK/KUP Family in Brassica napus and Its Potential Roles in the Response to K+ Deficiency.

The KT/HAK/KUP (HAK) family is the largest potassium (K+) transporter family in plants, which plays key roles in K+ uptake and homeostasis, stress resistance, and root and embryo development. However, the HAK family has not yet been characterized in Brassica napus. In this study, 40 putative B. napus HAK genes (BnaHAKs) are identified and divided into four groups (Groups I-III and V) on the basis of phylogenetic analysis. Gene structure analysis revealed 10 conserved intron insertion sites across different groups. Collinearity analysis demonstrated that both allopolyploidization and small-scale duplication events contributed to the large expansion of BnaHAKs. Transcription factor (TF)-binding network construction, cis-element analysis, and microRNA prediction revealed that the expression of BnaHAKs is regulated by multiple factors. Analysis of RNA-sequencing data further revealed extensive expression profiles of the BnaHAKs in groups II, III, and V, with limited expression in group I. Compared with group I, most of the BnaHAKs in groups II, III, and V were more upregulated by hormone induction based on RNA-sequencing data. Reverse transcription-quantitative polymerase reaction analysis revealed that the expression of eight BnaHAKs of groups I and V was markedly upregulated under K+-deficiency treatment. Collectively, our results provide valuable information and key candidate genes for further functional studies of BnaHAKs.

Na+ Transporter SvHKT1;1 from a Halophytic Turf Grass Is Specifically Upregulated by High Na+ Concentration and Regulates Shoot Na+ Concentration.

We characterized an Na+ transporter SvHKT1;1 from a halophytic turf grass, Sporobolus virginicus. SvHKT1;1 mediated inward and outward Na+ transport in Xenopus laevis oocytes and did not complement K+ transporter-defective mutant yeast. SvHKT1;1 did not complement athkt1;1 mutant Arabidopsis, suggesting its distinguishable function from other typical HKT1 transporters. The transcript was abundant in the shoots compared with the roots in S. virginicus and was upregulated by severe salt stress (500 mM NaCl), but not by lower stress. SvHKT1;1-expressing Arabidopsis lines showed higher shoot Na+ concentrations and lower salt tolerance than wild type (WT) plants under nonstress and salt stress conditions and showed higher Na+ uptake rate in roots at the early stage of salt treatment. These results suggested that constitutive expression of SvHKT1;1 enhanced Na+ uptake in root epidermal cells, followed by increased Na+ transport to shoots, which led to reduced salt tolerance. However, Na+ concentrations in phloem sap of the SvHKT1;1 lines were higher than those in WT plants under salt stress. Based on this result, together with the induction of the SvHKT1;1 transcription under high salinity stress, it was suggested that SvHKT1;1 plays a role in preventing excess shoot Na+ accumulation in S. virginicus.

Rice OsHAK16 functions in potassium uptake and translocation in shoot, maintaining potassium homeostasis and salt tolerance.

MAIN CONCLUSION: OsHAK16 mediates K uptake and root-to-shoot translocation in a broad range of external K concentrations, thereby contributing to the maintenance of K homeostasis and salt tolerance in the rice shoot. The HAK/KUP/KT transporters have been widely associated with potassium (K) transport across membranes in both microbes and plants. Here, we report the physiological function of OsHAK16, a member belonging to the HAK/KUP/KT family in rice (Oryza sativa L.). Transcriptional expression of OsHAK16 was up-regulated by K deficiency or salt stress. OsHAK16 is localized at the plasma membrane. OsHAK16 knockout (KO) dramatically reduced root K net uptake rate and growth at both 0.1 mM and 1 mM K supplies, while OsHAK16 overexpression (OX) increased total K uptake and growth only at 0.1 mM K level. OsHAK16-KO decreased the rate of rubidium (Rb) uptake and translocation compared to WT at both 0.2 mM and 1 mM Rb levels. OsHAK16 disruption decreased while its overexpression increased K concentration in root slightly but in shoot remarkably. The relative distribution of total K between shoot and root decreased by 30% in OsHAK16-KO lines and increased by 30% in its OX lines compared to WT. OsHAK16-KO diminished K uptake and K/Na ratio, while OsHAK16-OX improved K uptake and translocation from root to shoot, resulting in increased sensitivity and tolerance to salt stress, respectively. Expression of OsHAK16 enhanced the growth of high salt-sensitive yeast mutant by increasing its K but no Na content. Taking all these together, we conclude that OsHAK16 plays crucial roles in maintaining K homeostasis and salt tolerance in rice shoot.

Combined genetic disruption of K-Cl cotransporters and Gardos channel KCNN4 rescues erythrocyte dehydration in the SAD mouse model of sickle cell disease.

Excessive red cell dehydration contributes to the pathophysiology of sickle cell disease (SCD). The densest fraction of sickle red cells (with the highest corpuscular hemoglobin concentration) undergoes the most rapid polymerization of deoxy-hemoglobin S, leading to accelerated cell sickling and increased susceptibility to endothelial activation, red cell adhesion, and vaso-occlusion. Increasing red cell volume in order to decrease red cell density can thus serve as an adjunct therapeutic goal in SCD. Regulation of circulating mouse red cell volume and density is mediated largely by the Gardos channel, KCNN4, and the K-Cl cotransporters, KCC3 and KCC1. Whereas inhibition of the Gardos channel in subjects with sickle cell disease increased red cell volume, decreased red cell density, and improved other hematological indices in subjects with SCD, specific KCC inhibitors have not been available for testing. We therefore investigated the effect of genetic inactivation of KCC3 and KCC1 in the SAD mouse model of sickle red cell dehydration, finding decreased red cell density and improved hematological indices. We describe here generation of mice genetically deficient in the three major red cell volume regulatory gene products, KCNN4, KCC3, and KCC1 in C57BL6 non-sickle and SAD sickle backgrounds. We show that combined loss-of-function of all three gene products in SAD mice leads to incrementally increased MCV, decreased CHCM and % hyperchromic cells, decreased red cell density (phthalate method), increased resistance to hypo-osmotic lysis, and increased cell K content. The data show that combined genetic deletion of the Gardos channel and K-Cl cotransporters in a mouse SCD model decreases red cell density and improves several hematological parameters, supporting the strategy of combined pharmacological inhibition of these ion transport pathways in the adjunct treatment of human SCD.

Pumpkin CmHKT1;1 Controls Shoot Na⁺ Accumulation via Limiting Na⁺ Transport from Rootstock to Scion in Grafted Cucumber.

Soil salinity adversely affects the growth and yield of crops, including cucumber, one of the most important vegetables in the world. Grafting with salt-tolerant pumpkin as the rootstock effectively improves the growth of cucumber under different salt conditions by limiting Na⁺ transport from the pumpkin rootstock to the cucumber scion. High-affinity potassium transporters (HKTs) are crucial for the long distance transport of Na⁺ in plants, but the function of pumpkin HKTs in this process of grafted cucumber plants remains unclear. In this work, we have characterized CmHKT1;1 as a member of the HKT gene family in Cucurbita moschata and observed an obvious upregulation of CmHKT1;1 in roots under NaCl stress conditions. Heterologous expression analyses in yeast mutants indicated that CmHKT1;1 is a Na⁺-selective transporter. The transient expression in tobacco epidermal cells and in situ hybridization showed CmHKT1;1 localization at plasma membrane, and preferential expression in root stele. Moreover, ectopic expression of CmHKT1;1 in cucumber decreased the Na⁺ accumulation in the plants shoots. Finally, the CmHKT1;1 transgenic line as the rootstock decreased the Na⁺ content in the wild type shoots. These findings suggest that CmHKT1;1 plays a key role in the salt tolerance of grafted cucumber by limiting Na⁺ transport from the rootstock to the scion and can further be useful for engineering salt tolerance in cucurbit crops.

Cesium Uptake by Rice Roots Largely Depends Upon a Single Gene, HAK1, Which Encodes a Potassium Transporter.

Incidents at the Fukushima and Chernobyl nuclear power stations have resulted in widespread environmental contamination by radioactive nuclides. Among them, 137cesium has a 30 year half-life, and its persistence in soil raises serious food security issues. It is therefore important to prevent plants, especially crop plants, from absorbing radiocesium. In Arabidopsis thaliana, cesium ions are transported into root cells by several different potassium transporters such as high-affinity K+ transporter 5 (AtHAK5). Therefore, the cesium uptake pathway is thought to be highly redundant, making it difficult to develop plants with low cesium uptake. Here, we isolated rice mutants with low cesium uptake and reveal that the Oryza sativa potassium transporter OsHAK1, which is expressed on the surfaces of roots, is the main route of cesium influx into rice plants, especially in low potassium conditions. During hydroponic cultivation with low to normal potassium concentrations (0-206 µM: the normal potassium level in soil), cesium influx in OsHAK1-knockout lines was no greater than one-eighth that in the wild type. In field experiments, knockout lines of O. sativa HAK1 (OsHAK1) showed dramatically reduced cesium concentrations in grains and shoots, but their potassium uptake was not greatly affected and their grain yields were similar to that of the wild type. Our results demonstrate that, in rice roots, potassium transport systems other than OsHAK1 make little or no contribution to cesium uptake. These results show that low cesium uptake rice lines can be developed for cultivation in radiocesium-contaminated areas.

SbHKT1;4, a member of the high-affinity potassium transporter gene family from Sorghum bicolor, functions to maintain optimal Na⁺ /K⁺ balance under Na⁺ stress.

In halophytic plants, the high-affinity potassium transporter HKT gene family can selectively uptake K⁺ in the presence of toxic concentrations of Na⁺. This has so far not been well examined in glycophytic crops. Here, we report the characterization of SbHKT1;4, a member of the HKT gene family from Sorghum bicolor. Upon Na⁺ stress, SbHKT1;4 expression was more strongly upregulated in salt-tolerant sorghum accession, correlating with a better balanced Na⁺ /K⁺ ratio and enhanced plant growth. Heterogeneous expression analyses in mutants of Saccharomyces cerevisiae and Arabidopsis thaliana indicated that overexpressing SbHKT1;4 resulted in hypersensitivity to Na⁺ stress, and such hypersensitivity could be alleviated with the supply of elevated levels of K⁺, implicating that SbHKT1;4 may mediate K⁺ uptake in the presence of excessive Na⁺. Further electrophysiological evidence demonstrated that SbHKT1;4 could transport Na⁺ and K⁺ when expressed in Xenopus laevis oocytes. The relevance of the finding that SbHKT1;4 functions to maintain optimal Na⁺ /K⁺ balance under Na⁺ stress to the breeding of salt-tolerant glycophytic crops is discussed.

Positive Regulatory Roles of Manihot esculenta HAK5 under K+ Deficiency or High Salt Stress.

HAK/KUP/KT family members have been identified as playing key roles in K+ uptake and salt tolerance in numerous higher plants. However, their functions in cassava (Manihot esculenta Cantz) remain unknown. In this study, a gene encoding for a high-affinity potassium transporter (MeHAK5) was isolated from cassava and its function was investigated. Subcellular localization analysis showed that MeHAK5 is a plasma membrane-localized transporter. RT-PCR and RT-qPCR indicated that MeHAK5 is predominantly expressed in cassava roots, where it is upregulated by low potassium or high salt; in particular, its highest expression levels separately increased by 2.2 and 2.9 times after 50 µM KCl and 150 mM NaCl treatments. When heterologously expressed in yeast, MeHAK5 mediated K+ uptake within the cells of the yeast strain CY162 and rescued the salt-sensitive phenotype of AXT3K yeast. MeHAK5 overexpression in transgenic Arabidopsis plants exhibited improved growth and increased shoot K+ content under low potassium conditions. Under salt stress, MeHAK5 transgenic Arabidopsis plants accumulated more K+ in the shoots and roots and had reduced Na+ content in the shoots. As a result, MeHAK5 transgenic Arabidopsis demonstrated a more salt-tolerant phenotype. These results suggest that MeHAK5 functions as a high-affinity K+ transporter under K+ starvation conditions, improving K+/Na+ homeostasis and thereby functioning as a positive regulator of salt stress tolerance in transgenic Arabidopsis. Therefore, MeHAK5 may be a suitable candidate gene for improving K+ utilization efficiency and salt tolerance.

Mechanism of high affinity potassium transporter (HKT) towards improved crop productivity in saline agricultural lands.

Glycophytic plants are susceptible to salinity and their growth is hampered in more than 40 mM of salt. Salinity not only affects crop yield but also limits available land for farming by decreasing its fertility. Presence of distinct traits in response to environmental conditions might result in evolutionary adaptations. A better understanding of salinity tolerance through a comprehensive study of how Na+ is transported will help in the development of plants with improved salinity tolerance and might lead to increased yield of crops growing in strenuous environment. Ion transporters play pivotal role in salt homeostasis and maintain low cytotoxic effect in the cell. High-affinity potassium transporters are the critical class of integral membrane proteins found in plants. It mainly functions to remove excess Na+ from the transpiration stream to prevent sodium toxicity in the salt-sensitive shoot and leaf tissues. However, there are large number of HKT proteins expressed in plants, and it is possible that these members perform in a wide range of functions. Understanding their mechanism and functions will aid in further manipulation and genetic transformation of different crops. This review focuses on current knowledge of ion selectivity and molecular mechanisms controlling HKT gene expression. The current review highlights the mechanism of different HKT transporters from different plant sources and how this knowledge could prove as a valuable tool to improve crop productivity.

Genome-wide identification and expression analysis of HAK/KUP/KT potassium transporter provides insights into genes involved in responding to potassium deficiency and salt stress in pepper (Capsicum annuum L.).

In plants, the HAK/KUP/KT family is the largest group of potassium transporters, and it plays an important role in mineral element absorption, plant growth, environmental stress adaptation, and symbiosis. Although these important genes have been investigated in many plant species, limited information is currently available on the HAK/KUP/KT genes for Pepper (Capsicum annuum L.). In the present study, a total of 20 CaHAK genes were identified from the pepper genome and the CaHAK genes were numbered 1 - 20 based on phylogenetic analysis. For the genes and their corresponding proteins, the physicochemical properties, phylogenetic relationship, chromosomal distribution, gene structure, conserved motifs, gene duplication events, and expression patterns were analyzed. Phylogenetic analysis divided CaHAK genes into four cluster (I-IV) based on their structural features and the topology of the phylogenetic tree. Purifying selection played a crucial role in the evolution of CaHAK genes, while whole-genome triplication contributed to the expansion of the CaHAK gene family. The expression patterns showed that CaHAK proteins exhibited functional divergence in terms of plant K+ uptake and salt stress response. In particular, transcript abundance of CaHAK3 and CaHAK7 was strongly and specifically up-regulated in pepper roots under low K+ or high salinity conditions, suggesting that these genes are candidates for high-affinity K+ uptake transporters and may play crucial roles in the maintenance of the Na+/K+ balance during salt stress in pepper. In summary, the results not only provided the important information on the characteristics and evolutionary relationships of CaHAKs, but also provided potential genes responding to potassium deficiency and salt stress. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s13205-022-03136-z.

The Potassium Transporter OsHAK5 Alters Rice Architecture via ATP-Dependent Transmembrane Auxin Fluxes.

Plant HAK/KUP/KT family members function as plasma membrane (PM) H+/K+ symporters and may modulate chemiosmotically-driven polar auxin transport (PAT). Here, we show that inactivation of OsHAK5, a rice K+ transporter gene, decreased rootward and shootward PAT, tiller number, and the length of both lateral roots and root hairs, while OsHAK5 overexpression increased PAT, tiller number, and root hair length, irrespective of the K+ supply. Inhibitors of ATP-binding-cassette type-B transporters, NPA and BUM, abolished the OsHAK5-overexpression effect on PAT. The mechanistic basis of these changes included the OsHAK5-mediated decrease of transmembrane potential (depolarization), increase of extracellular pH, and increase of PM-ATPase activity. These findings highlight the dual roles of OsHAK5 in altering cellular chemiosmotic gradients (generated continuously by PM H+-ATPase) and regulating ATP-dependent auxin transport. Both functions may underlie the prominent effect of OsHAK5 on rice architecture, which may be exploited in the future to increase crop yield via genetic manipulations.

Comparative Functional Analysis of Class II Potassium Transporters, SvHKT2;1, SvHKT2;2, and HvHKT2;1, on Ionic Transport and Salt Tolerance in Transgenic Arabidopsis.

Class II high-affinity potassium transporters (HKT2s) mediate Na+-K+ cotransport and Na+/K+ homeostasis under K+-starved or saline conditions. Their functions have been studied in yeast and X. laevis oocytes; however, little is known about their respective properties in plant cells. In this study, we characterized the Na+ and K+ transport properties of SvHKT2;1, SvHKT2;2 and HvHKT2;1 in Arabidopsis under different ionic conditions. The differences were detected in shoot K+ accumulation and root K+ uptake under salt stress conditions, K+ accumulation in roots and phloem sap under K+-starved conditions, and shoot and root Na+ accumulation under K+-starved conditions among the HKT2s transgenic lines and WT plants. These results indicate the diverse ionic transport properties of these HKT2s in plant cells, which could not be detected using yeast or X. laevis oocytes. Furthermore, Arabidopsis expressing HKT2s showed reduced salt tolerance, while over-expression of HvHKT2;1 in barley, which has the ability to sequestrate Na+, showed enhanced salt tolerance by accumulating Na+ in the shoots. These results suggest that the coordinated enhancement of Na+ accumulation and sequestration mechanisms in shoots could be a promising strategy to confer salt tolerance to glycophytes.

K+ Release Assay and K+ Measurement in Oocyte Assay.

The Xenopus oocyte is a powerful system for the exogenous expression and functional characterization of plant membrane transport proteins. Until now, a number of potassium transporters and channels have been identified in oocytes expression system by the two-electrode voltage clamp technology. It is difficult to characterize K+/H+ anti-transporters, especially, electroneutral transporter. The K+ efflux assay system enables easy, fast, large-scale measurement of the transporters activity without two-electrode voltage clamp technology. This protocol describes a technique to measure the efflux activity of potassium transporter in oocytes expressing system.

Contribution of KUPs to potassium and cesium accumulation appears complementary in Arabidopsis.

Cesium has no known beneficial effects on plants and while plants have the ability to absorb it through the root system, plant growth is retarded at high concentrations. Recently, we have shown that potassium influx through a potassium channel complex AKT1-KC1 is inhibited by cesium in Arabidopsis thaliana and the resultant reduction in potassium accumulation in the plant is the primary cause of retarded growth. By contrast, a major potassium transporter, HAK5 whose function is crucial under potassium deficiency, was found to be either not affected or complementary under cesium stress in the low affinity potassium range. Here we show the effects of insertional mutation on other members of KUP/HAK/KT gene family in response to cesium stress. Potassium and cesium concentrations in each mutant line demonstrated that disruption of a single KUP/HAK/KT gene was not sufficient to significantly reduce potassium/cesium accumulation, suggesting a complementary effect among these KUP (K+ UPTAKE PERMEASE) transporters.

Association Analysis and Identification of ZmHKT1;5 Variation With Salt-Stress Tolerance.

The high-affinity potassium transporter (HKT) genes are essential for plant salt stress tolerance. However, there were limited studies on HKTs in maize (Zea mays), and it is basically unknown whether natural sequence variations in these genes are associated with the phenotypic variability of salt tolerance. Here, the characterization of ZmHKT1;5 was reported. Under salt stress, ZmHKT1;5 expression increased strongly in salt-tolerant inbred lines, which accompanied a better-balanced Na+/K+ ratio and preferable plant growth. The association between sequence variations in ZmHKT1;5 and salt tolerance was evaluated in a diverse population comprising 54 maize varieties from different maize production regions of China. Two SNPs (A134G and A511G) in the coding region of ZmHKT1;5 were significantly associated with different salt tolerance levels in maize varieties. In addition, the favorable allele of ZmHKT1; 5 identified in salt tolerant maize varieties effectively endowed plant salt tolerance. Transgenic tobacco plants of overexpressing the favorable allele displayed enhanced tolerance to salt stress better than overexpressing the wild type ZmHKT1;5. Our research showed that ZmHKT1;5 expression could effectively enhance salt tolerance by maintaining an optimal Na+/K+ balance and increasing the antioxidant activity that keeps reactive oxygen species (ROS) at a low accumulation level. Especially, the two SNPs in ZmHKT1;5 might be related with new amino acid residues to confer salt tolerance in maize. Key Message: Two SNPs of ZmHKT1;5 related with salt tolerance were identified by association analysis. Overexpressing ZmHKT1;5 in tobaccos showed that the SNPs might enhance its ability to regulating Na+/K+ homeostasis.