Non-autonomous stomatal control by pavement cell turgor via the K+ channel subunit AtKC1.

Stomata optimize land plants' photosynthetic requirements and limit water vapor loss. So far, all of the molecular and electrical components identified as regulating stomatal aperture are produced, and operate, directly within the guard cells. However, a completely autonomous function of guard cells is inconsistent with anatomical and biophysical observations hinting at mechanical contributions of epidermal origins. Here, potassium (K+) assays, membrane potential measurements, microindentation, and plasmolysis experiments provide evidence that disruption of the Arabidopsis thaliana K+ channel subunit gene AtKC1 reduces pavement cell turgor, due to decreased K+ accumulation, without affecting guard cell turgor. This results in an impaired back pressure of pavement cells onto guard cells, leading to larger stomatal apertures. Poorly rectifying membrane conductances to K+ were consistently observed in pavement cells. This plasmalemma property is likely to play an essential role in K+ shuttling within the epidermis. Functional complementation reveals that restoration of the wild-type stomatal functioning requires the expression of the transgenic AtKC1 at least in the pavement cells and trichomes. Altogether, the data suggest that AtKC1 activity contributes to the building of the back pressure that pavement cells exert onto guard cells by tuning K+ distribution throughout the leaf epidermis.

Inhibition of SlSKOR by SlCIPK23-SlCBL1/9 uncovers CIPK-CBL-target network rewiring in land plants.

Transport of K+ to the xylem is a key process in the mineral nutrition of the shoots. Although CIPK-CBL complexes have been widely shown to regulate K+ uptake transport systems, no information is available about the xylem ones. Here, we studied the physiological roles of the voltage-gated K+ channel SlSKOR and its regulation by the SlCIPK23-SlCBL1/9 complexes in tomato plants. We phenotyped gene-edited slskor and slcipk23 tomato knockout mutants and carried out two-electrode voltage-clamp (TEVC) and BiFC assays in Xenopus oocytes as key approaches. SlSKOR was preferentially expressed in the root stele and was important not only for K+ transport to shoots but also, indirectly, for that of Ca2+ , Mg2+ , Na+ , NO3 - , and Cl- . Surprisingly, the SlCIPK23-SlCBL1/9 complexes turned out to be negative regulators of SlSKOR. Inhibition of SlSKOR by SlCIPK23-SlCBL1/9 was observed in Xenopus oocytes and tomato plants. Regulation of SKOR-like channels by CIPK23-CBL1 complexes was also present in Medicago, grapevine, and lettuce but not in Arabidopsis and saltwater cress. Our results provide a molecular framework for coordinating root K+ uptake and its translocation to the shoot by SlCIPK23-SlCBL1/9 in tomato plants. Moreover, they evidenced that CIPK-CBL-target networks have evolved differently in land plants.

Insights into the mechanisms of transport and regulation of the arabidopsis high-affinity K+ transporter HAK51.

The high-affinity K+ transporter HAK5 from Arabidopsis (Arabidopsis thaliana) is essential for K+ acquisition and plant growth at low micromolar K+ concentrations. Despite its functional relevance in plant nutrition, information about functional domains of HAK5 is scarce. Its activity is enhanced by phosphorylation via the AtCIPK23/AtCBL1-9 complex. Based on the recently published three-dimensionalstructure of the bacterial ortholog KimA from Bacillus subtilis, we have modeled AtHAK5 and, by a mutational approach, identified residues G67, Y70, G71, D72, D201, and E312 as essential for transporter function. According to the structural model, residues D72, D201, and E312 may bind K+, whereas residues G67, Y70, and G71 may shape the selective filter for K+, which resembles that of K+shaker-like channels. In addition, we show that phosphorylation of residue S35 by AtCIPK23 is required for reaching maximal transport activity. Serial deletions of the AtHAK5 C-terminus disclosed the presence of an autoinhibitory domain located between residues 571 and 633 together with an AtCIPK23-dependent activation domain downstream of position 633. Presumably, autoinhibition of AtHAK5 is counteracted by phosphorylation of S35 by AtCIPK23. Our results provide a molecular model for K+ transport and describe CIPK-CBL-mediated regulation of plant HAK transporters.

Functional characterization and physiological roles of the single Shaker outward K+ channel in Medicago truncatula.

The model legume Medicago truncatula possesses a single outward Shaker K+ channel, whereas Arabidopsis thaliana possesses two channels of this type, named AtSKOR and AtGORK, with AtSKOR having been shown to play a major role in K+ secretion into the xylem sap in the root vasculature and with AtGORK being shown to mediate the efflux of K+ across the guard cell membrane, leading to stomatal closure. Here we show that the expression pattern of the single M. truncatula outward Shaker channel, which has been named MtGORK, includes the root vasculature, guard cells and root hairs. As shown by patch-clamp experiments on root hair protoplasts, besides the Shaker-type slowly activating outwardly rectifying K+ conductance encoded by MtGORK, a second K+ -permeable conductance, displaying fast activation and weak rectification, can be expressed by M. truncatula. A knock-out (KO) mutation resulting in an absence of MtGORK activity is shown to weakly reduce K+ translocation to shoots, and only in plants engaged in rhizobial symbiosis, but to strongly affect the control of stomatal aperture and transpirational water loss. In legumes, the early electrical signaling pathway triggered by Nod-factor perception is known to comprise a short transient depolarization of the root hair plasma membrane. In the absence of the functional expression of MtGORK, the rate of the membrane repolarization is found to be decreased by a factor of approximately two. This defect was without any consequence on infection thread development and nodule production in plants grown in vitro, but a decrease in nodule production was observed in plants grown in soil.

The protein kinase SlCIPK23 boosts K+ and Na+ uptake in tomato plants.

Regulation of root transport systems is essential under fluctuating nutrient supply. In the case of potassium (K+ ), HAK/KUP/KT K+ transporters and voltage-gated K+ channels ensure root K+ uptake in a wide range of K+ concentrations. In Arabidopsis, the CIPK23/CBL1-9 complex regulates both transporter- and channel-mediated root K+ uptake. However, research about K+ homeostasis in crops is in demand due to species-specific mechanisms. In the present manuscript, we studied the contribution of the voltage-gated K+ channel LKT1 and the protein kinase SlCIPK23 to K+ uptake in tomato plants by analysing gene-edited knockout tomato mutant lines, together with two-electrode voltage-clamp experiments in Xenopus oocytes and protein-protein interaction analyses. It is shown that LKT1 is a crucial player in tomato K+ nutrition by contributing approximately 50% to root K+ uptake under K+ -sufficient conditions. Moreover, SlCIPK23 was responsible for approximately 100% of LKT1 and approximately 40% of the SlHAK5 K+ transporter activity in planta. Mg+2 and Na+ compensated for K+ deficit in tomato roots to a large extent, and the accumulation of Na+ was strongly dependent on SlCIPK23 function. The role of CIPK23 in Na+ accumulation in tomato roots was not conserved in Arabidopsis, which expands the current set of CIPK23-like protein functions in plants.

Genome-wide identification and expression analysis of two component system genes in Cicer arietinum.

The two-component system (TCS) plays an important role in signal transduction pathways, cytokinin signaling and stress resistance of prokaryotes and eukaryotes. It is comprised of three types of proteins in plants; histidine kinases (HKs), histidine phosphotransfer proteins (HPs) and response regulators (RRs). Chickpea (Cicer arietinum L.) is one of the most important legume crops worldwide with special economic value in semi-arid tropics. Availability of complete genome sequence of chickpea presents a valuable resource for comparative analysis among angiosperms. In current study, Arabidopsis thaliana and Oryza sativa were used as reference plant species for comparative genomics analysis with C. arietinum. A genome-wide computational survey enabled us to identify putative members of TCS protein family including 18HKs, 26 RRs (7 type-A, 7 type-B, 2 type C and 10 pseudo) and 7 HPs (5 true and 2pseudo) genes in chickpea. The predicted TCS genes displayed family specific intron/exon organization and were randomly distributed across all the eight chromosomes. Comparative phylogenetic and evolutionary analysis suggested a variable conservation of TCS genes in relation to mono/dicot model plants and segmental duplication was the principal route of expansion for this family in chickpea. The promoter regions of TCS genes exhibited several abiotic stress-related cis-elements indicating their involvement in abiotic stress response. The expression analysis of TCS genes demonstrated stress (drought, heat, osmotic and salt) specific differential expression. Current study provides insight into TCS genes in C. arietinum, which will be helpful for further functional analysis of these genes in response to different abiotic stresses.

Doing 'business as usual' comes with a cost: evaluating energy cost of maintaining plant intracellular K+ homeostasis under saline conditions.

Salinization of agricultural lands is a major threat to agriculture. Many different factors affect and determine plant salt tolerance. Nonetheless, there is a consensus on the relevance of maintaining an optimal cytosolic potassium : sodium ion (K+  : Na+ ) ratio for salinity tolerance in plants. This ratio depends on the operation of plasma membrane and tonoplast transporters. In the present review we focus on some aspects related to the energetic cost of maintaining that K+  : Na+ ratio. One of the factors that affect the cost of the first step of K+ acquisition - root K+ uptake through High Affinity K+ transporter and Arabidopsis K+ transport system 1 transport systems - is the value of the plasma membrane potential of root cells, a parameter that may differ amongst plant species. In addition to its role in nutrition, cytosolic K+ also is important for signalling, and K+ efflux through gated outward-rectifying K+ and nonselective cation channels can be regarded as a switch to redirect energy towards defence reactions. In maintaining cytosolic K+ , the great buffer capacity of the vacuole should be considered. The possible role of high-affinity K+ transporters (HKT)2s in mediating K+ uptake under saline conditions and the importance of cycling of K+ throughout the plant also are discussed.

Root high-affinity K+ and Cs+ uptake and plant fertility in tomato plants are dependent on the activity of the high-affinity K+ transporter SlHAK5.

Root K+ acquisition is a key process for plant growth and development, extensively studied in the model plant Arabidopsis thaliana. Because important differences may exist among species, translational research supported by specific studies is needed in crops such as tomato. Here we present a reverse genetics study to demonstrate the role of the SlHAK5 K+ transporter in tomato K+ nutrition, Cs+ accumulation and its fertility. slhak5 KO lines, generated by CRISPR-Cas edition, were characterized in growth experiments, Rb+ and Cs+ uptake tests and root cells K+ -induced plasma membrane depolarizations. Pollen viability and its K+ accumulation capacity were estimated by using the K+ -sensitive dye Ion Potassium Green 4. SlHAK5 is the major system for high-affinity root K+ uptake required for plant growth at low K+ , even in the presence of salinity. It also constitutes a pathway for Cs+ entry in tomato plants with a strong impact on fruit Cs+ accumulation. SlHAK5 also contributes to pollen K+ uptake and viability and its absence produces almost seedless fruits. Knowledge gained into SlHAK5 can serve as a model for other crops with fleshy fruits and it can help to generate tools to develop low Cs+ or seedless fruits crops.

Arabidopsis K+ transporter HAK5-mediated high-affinity root K+ uptake is regulated by protein kinases CIPK1 and CIPK9.

The high-affinity K+ transporter HAK5 is the major contributor to root K+ uptake from dilute solutions in K+-starved Arabidopsis plants. Its functionality is tightly regulated and its activity is enhanced under K+ starvation by the transcriptional induction of the AtHAK5 gene, and by the activation of the transporter via the AtCBL1-AtCIPK23 complex. In the present study, the 26 members of the Arabidopsis CIPK protein kinase family were screened in yeast for their capacity to activate HAK5-mediated K+ uptake. Among them, AtCIPK1 was the most efficient activator of AtHAK5. In addition, AtCIPK9, previously reported to participate in K+ homeostasis, also activated the transporter. In roots, the genes encoding AtCIPK1 and AtCIPK9 were induced by K+ deprivation and atcipk1 and atcipk9 Arabidopsis KO mutants showed a reduced AtHAK5-mediated Rb+ uptake. Activation of AtHAK5 by AtCIPK1 did not occur under hyperosmotic stress conditions, where AtCIPK1 function has been shown to be required to maintain plant growth. Taken together, our data contribute to the identification of the complex regulatory networks that control the high-affinity K+ transporter AtHAK5 and root K+ uptake.

Characterization of the grapevine Shaker K+ channel VvK3.1 supports its function in massive potassium fluxes necessary for berry potassium loading and pulvinus-actuated leaf movements.

In grapevine, climate changes lead to increased berry potassium (K+ ) contents that result in must with low acidity. Consequently, wines are becoming 'flat' to the taste, with poor organoleptic properties and low potential aging, resulting in significant economic loss. Precise investigation into the molecular determinants controlling berry K+ accumulation during its development are only now emerging. Here, we report functional characterization by electrophysiology of a new grapevine Shaker-type K+ channel, VvK3.1. The analysis of VvK3.1 expression patterns was performed by qPCR and in situ hybridization. We found that VvK3.1 belongs to the AKT2 channel phylogenetic branch and is a weakly rectifying channel, mediating both inward and outward K+ currents. We showed that VvK3.1 is highly expressed in the phloem and in a unique structure located at the two ends of the petiole, identified as a pulvinus. From the onset of fruit ripening, all data support the role of the VvK3.1 channel in the massive K+ fluxes from the phloem cell cytosol to the berry apoplast during berry K+ loading. Moreover, the high amount of VvK3.1 transcripts detected in the pulvinus strongly suggests a role for this Shaker in the swelling and shrinking of motor cells involved in paraheliotropic leaf movements.

Modulation of K+ translocation by AKT1 and AtHAK5 in Arabidopsis plants.

Root cells take up K+ from the soil solution, and a fraction of the absorbed K+ is translocated to the shoot after being loaded into xylem vessels. K+ uptake and translocation are spatially separated processes. K+ uptake occurs in the cortex and epidermis whereas K+ translocation starts at the stele. Both uptake and translocation processes are expected to be linked, but the connection between them is not well characterized. Here, we studied K+ uptake and translocation using Rb+ as a tracer in wild-type Arabidopsis thaliana and in T-DNA insertion mutants in the K+ uptake or translocation systems. The relative amount of translocated Rb+ to the shoot was positively correlated with net Rb+ uptake rates, and the akt1 athak5 T-DNA mutant plants were more efficient in their allocation of Rb+ to shoots. Moreover, a mutation of SKOR and a reduced plant transpiration prevented the full upregulation of AtHAK5 gene expression and Rb+ uptake in K+ -starved plants. Lastly, Rb+ was found to be retrieved from root xylem vessels, with AKT1 playing a significant role in K+ -sufficient plants. Overall, our results suggest that K+ uptake and translocation are tightly coordinated via signals that regulate the expression of K+ transport systems.

The combination of K+ deficiency with other environmental stresses: What is the outcome?

Potassium (K+ ) is a macronutrient known for its high mobility and positive charge, which allows efficient and fast control of the electrical balance and osmotic potential in plant cells. Such features allow K+ to remarkably contribute to plant stress adaptation. Some agricultural lands are deficient in K+ , imposing a stress that reduces crop yield and makes fertilization a common practice. However, individual stress conditions in the field are rare, and crops usually face a combination of different stresses. As plant response to a stress combination cannot always be deduced from individual stress action, it is necessary to gain insights into the specific mechanisms that connect K+ homeostasis with other stress effects to improve plant performance in the context of climate change. Surprisingly, plant responses to environmental stresses under a K+ -limiting scenario are poorly understood. In the present review, we summarize current knowledge and find substantial gaps regarding specific outcomes of K+ deficiency in addition to other environmental stresses. In this regard, combined nutrient deficiencies of K+ and other macronutrients are covered in the first part of the review and interactions arising from K+ deficiency with salinity, drought and biotic factors in the second part. Information available so far suggests a prominent role of potassium and nitrate transport systems and their regulatory proteins in the response of plants to several stress combinations. Thus, such molecular pathways, which are located at the crossroad between K+ homeostasis and environmental stresses, could be considered biotechnological targets in future studies.

High External K+ Concentrations Impair Pi Nutrition, Induce the Phosphate Starvation Response, and Reduce Arsenic Toxicity in Arabidopsis Plants.

Potassium (K+) and phosphorous (Pi) are two of the most important nutrients required by plants and there is an interest in studying how they are acquired. Most studies have focused on the characterization of the mechanisms involved in K+ and Pi uptake and their distribution within the plants, as well as the regulatory mechanisms involved. Evidence is emerging which points to interactions in the nutrition of different nutrients and to the existence of crosstalk in the signaling cascades regulating their acquisition. However, the interaction between K+ and Pi has been scarcely studied. Here we show that high concentrations of K+ in the external solution inhibit Pi uptake and impair Pi nutrition in Arabidopsis plants, resulting in the induction of phosphate starvation response (PSR) and the upregulation of genes encoding root phosphate uptake systems. The high K+-induced PSR depends on the PHR1 and PHL1 transcription factors that are key pieces of Pi signaling in Arabidopsis. Importantly, high K+ reduces arsenic accumulation in plants and its toxic effects. The results presented may help to design strategies to reduce Pi deficiency as well as the accumulation of arsenic in crops.

Production of low-Cs+ rice plants by inactivation of the K+ transporter OsHAK1 with the CRISPR-Cas system.

The occurrence of radiocesium in food has raised sharp health concerns after nuclear accidents. Despite being present at low concentrations in contaminated soils (below µm), cesium (Cs+ ) can be taken up by crops and transported to their edible parts. This plant capacity to take up Cs+ from low concentrations has notably affected the production of rice (Oryza sativa L.) in Japan after the nuclear accident at Fukushima in 2011. Several strategies have been put into practice to reduce Cs+ content in this crop species such as contaminated soil removal or adaptation of agricultural practices, including dedicated fertilizer management, with limited impact or pernicious side-effects. Conversely, the development of biotechnological approaches aimed at reducing Cs+ accumulation in rice remain challenging. Here, we show that inactivation of the Cs+ -permeable K+ transporter OsHAK1 with the CRISPR-Cas system dramatically reduced Cs+ uptake by rice plants. Cs+ uptake in rice roots and in transformed yeast cells that expressed OsHAK1 displayed very similar kinetics parameters. In rice, Cs+ uptake is dependent on two functional properties of OsHAK1: (i) a poor capacity of this system to discriminate between Cs+ and K+ ; and (ii) a high capacity to transport Cs+ from very low external concentrations that is likely to involve an active transport mechanism. In an experiment with a Fukushima soil highly contaminated with 137 Cs+ , plants lacking OsHAK1 function displayed strikingly reduced levels of 137 Cs+ in roots and shoots. These results open stimulating perspectives to smartly produce safe food in regions contaminated by nuclear accidents.

Pharmacological and gene regulation properties point to the SlHAK5 K+ transporter as a system for high-affinity Cs+ uptake in tomato plants.

Potassium (K+ ) and cesium (Cs+ ) are chemically similar but while K+ is an essential nutrient, Cs+ can be toxic for living organisms, plants included. Two different situations could lead to problems derived from the presence of Cs+ in agricultural systems: (1) presence of Cs+ at high concentrations that could produce toxic effects on plants, (2) presence of micromolar concentrations of radiocesium, which can be accumulated in the plant and affect animal and human health through the food chain. While K+ uptake has been well described in tomato plants, information on molecular mechanisms involved in Cs+ accumulation in this species is absent. Here, we show that in tomato plants, high concentrations of Cs+ produce deficiency of K+ but do not induce high-affinity K+ uptake or the gene encoding the high-affinity K+ transporter SlHAK5. At these concentrations, Cs+ uptake takes place through a Ca2+ -sensitive pathway, probably a non-selective cation channel. At micromolar concentrations, Cs+ is accumulated by a high-affinity uptake system upregulated in K+ -starved plants. This high-affinity Cs+ uptake shares features with high-affinity K+ uptake. It is sensitive to NH4+ and insensitive to Ba2+ and Ca2+ and its presence parallels the pattern of SlHAK5 expression. Moreover, blockers of reactive oxygen species and ethylene action repress SlHAK5 and negatively regulate both high-affinity K+ and Cs+ uptake. Thus, we propose that SlHAK5 contributes to Cs+ uptake from micromolar concentrations in tomato plants and can constitute a pathway for radiocesium transfer from contaminated areas to the food chain.

Tolerance to Stress Combination in Tomato Plants: New Insights in the Protective Role of Melatonin.

Abiotic stresses such as drought, heat or salinity are major causes of yield loss worldwide. Recent studies have revealed that the acclimation of plants to a combination of different environmental stresses is unique and therefore cannot be directly deduced from studying the response of plants to each of the different stresses applied individually. The efficient detoxification of reactive oxygen species (ROS) is thought to play a key role in enhancing the tolerance of plants to abiotic stresses. Here, we report on the role of melatonin in the protection of the photosynthetic apparatus through the increase in ROS detoxification in tomato plants grown under the combination of salinity and heat, two of the most common abiotic stresses known to act jointly. Plants treated with exogenous melatonin showed a different modulation in the expression on some antioxidant-related genes and their related enzymes. More specifically, ascorbate peroxidase, glutathione reductase, glutathione peroxidase and phospholipid hydroperoxide glutathione peroxidase (APX, GR, GPX and Ph-GPX, resepctively) showed an antagonistic regulation as compared to plants that did not receive melatonin. This translated into a better antioxidant capacity and to a lesser ROS accumulation under stress combination. The performance of the photosynthesis parameters and the photosystems was also increased in plants treated with exogenous melatonin under the combination of salinity and heat. In accordance with these findings, tomato plants treated with melatonin were found to grow better under stress combination that the non-treated ones. Our study highlights the important role that exogenous melatonin plays in the acclimation of plants to a combination of two different abiotic stresses, and how this compound can specifically regulate oxidative stress-related genes and enzymes to increase plant tolerance.

The CBL-Interacting Protein Kinase CIPK23 Regulates HAK5-Mediated High-Affinity K+ Uptake in Arabidopsis Roots.

Plant growth and development requires efficient acquisition of essential elements. Potassium (K(+)) is an important macronutrient present in the soil solution at a wide range of concentrations. Regulation of the K(+) uptake systems in the roots is essential to secure K(+) supply. It has been shown in Arabidopsis (Arabidopsis thaliana) that when the external K(+) concentration is very low (<10 µm), K(+) nutrition depends exclusively on the high-affinity K(+) transporter5 (HAK5). Low-K(+)-induced transcriptional activation of the gene encoding HAK5 has been previously reported. Here, we show the posttranscriptional regulation of HAK5 transport activity by phosphorylation. Expression in a heterologous system showed that the Ca(2+) sensors calcineurin B-like (CBL1), CBL8, CBL9, and CBL10, together with CBL-interacting protein kinase23 (CIPK23), activated HAK5 in vivo. This activation produced an increase in the affinity and the Vmax of K(+) transport. In vitro experiments show that the N terminus of HAK5 is phosphorylated by CIPK23. This supports the idea that phosphorylation of HAK5 induces a conformational change that increases its affinity for K(+). Experiments of K(+) (Rb(+)) uptake and growth measurements in low-K(+) medium with Arabidopsis single mutants hak5, akt1, and cipk23, double mutants hak5 akt1, hak5 cipk23, and akt1 cipk23, and the triple mutant hak5 akt1 cipk23 confirmed the regulatory role of CIPK23 in planta.

Distinct amino acids in the C-linker domain of the Arabidopsis K+ channel KAT2 determine its subcellular localization and activity at the plasma membrane.

Shaker K(+) channels form the major K(+) conductance of the plasma membrane in plants. They are composed of four subunits arranged around a central ion-conducting pore. The intracellular carboxy-terminal region of each subunit contains several regulatory elements, including a C-linker region and a cyclic nucleotide-binding domain (CNBD). The C-linker is the first domain present downstream of the sixth transmembrane segment and connects the CNBD to the transmembrane core. With the aim of identifying the role of the C-linker in the Shaker channel properties, we performed subdomain swapping between the C-linker of two Arabidopsis (Arabidopsis thaliana) Shaker subunits, K(+) channel in Arabidopsis thaliana2 (KAT2) and Arabidopsis thaliana K(+) rectifying channel1 (AtKC1). These two subunits contribute to K(+) transport in planta by forming heteromeric channels with other Shaker subunits. However, they display contrasting behavior when expressed in tobacco mesophyll protoplasts: KAT2 forms homotetrameric channels active at the plasma membrane, whereas AtKC1 is retained in the endoplasmic reticulum when expressed alone. The resulting chimeric/mutated constructs were analyzed for subcellular localization and functionally characterized. We identified two contiguous amino acids, valine-381 and serine-382, located in the C-linker carboxy-terminal end, which prevent KAT2 surface expression when mutated into the equivalent residues from AtKC1. Moreover, we demonstrated that the nine-amino acid stretch 312TVRAASEFA320 that composes the first C-linker α-helix located just below the pore is a crucial determinant of KAT2 channel activity. A KAT2 C-linker/CNBD three-dimensional model, based on animal HCN (for Hyperpolarization-activated, cyclic nucleotide-gated K(+)) channels as structure templates, has been built and used to discuss the role of the C-linker in plant Shaker inward channel structure and function.

A low K+ signal is required for functional high-affinity K+ uptake through HAK5 transporters.

The high-affinity K(+) transporter HAK5 is a key system for root K(+) uptake and, under very low external K(+), the only one capable of supplying K(+) to the plant. Functional HAK5-mediated K(+) uptake should be tightly regulated for plant adaptation to different environmental conditions. Thus, it has been described that the gene encoding the transporter is transcriptionally regulated, being highly induced under K(+) limitation. Here we show that environmental conditions, such as the lack of K(+), NO(3)(-) or P, that induced a hyperpolarization of the plasma membrane of root cells, induce HAK5 transcription. However, only the deprivation of K(+) produces functional HAK5-mediated K(+) uptake in the root. These results suggest on the one hand the existence of a posttranscriptional regulation of HAK5 elicited by the low K(+) signal and on the other that HAK5 may be involved in yet-unknown functions related to NO(3)(-) and P deficiencies. These results have been obtained here with Solanum lycopersicum (cv. Micro-Tom) as well as Arabidopsis thaliana plants, suggesting that the posttranscriptional regulation of high-affinity HAK transporters take place in all plant species.

Relevance of the SlCIPK23 kinase in Na+ uptake and root morphology in K+-starved tomato plants.

The beneficial effects of Na+ as a substitute for K+ have been well-documented at the physiological level. However, the transport systems and regulatory mechanisms that allow Na+ acquisition under K+ deficiency remain poorly understood in the majority of land plants. In tomato, SlCIPK23 kinase was involved in Na+ accumulation in K+-starved plants, in addition to activating the LKT1 K+ channel and the K+ transporter SlHAK5. We used the central role of SlCIPK23 in K+ and Na+ acquisition to study which molecular entities mediate Na+ uptake with knockout tomato mutants and expression in heterologous systems. Two main pathways for Na+ uptake were deduced in tomato plants: an NH4+-sensitive pathway dependent on SlCIPK23, and a second one sensitive to Ba2+, Ca2+, La3+, and Li+. The addition of Na+ (10 mM) to lkt1, slhak5, or slcipk23 mutant KO lines produced interesting changes in root morphology. In particular, the roots of slcipk23 plants were longer and lighter than those of the WT under K+-deficient conditions and this effect was reversed by the addition of 10 mM Na+. These results provide a stimulating perspective for the study of the beneficial effects of Na+ in crops.