Capacity and plasticity of potassium channels and high-affinity transporters in roots of barley and Arabidopsis

Abstract

The role of potassium (K(+)) transporters in high- and low-affinity K(+) uptake was examined in roots of intact barley (Hordeum vulgare) and Arabidopsis (Arabidopsis thaliana) plants by use of (42)K radiotracing, electrophysiology, pharmacology, and mutant analysis. Comparisons were made between results from barley and five genotypes of Arabidopsis, including single and double knockout mutants for the high-affinity transporter, AtHAK5, and the Shaker-type channel, AtAKT1. In Arabidopsis, steady-state K(+) influx at low external K(+) concentration ([K(+)]ext = 22.5 µm) was predominantly mediated by AtAKT1 when high-affinity transport was inhibited by ammonium, whereas in barley, by contrast, K(+) channels could not operate below 100 µm. Withdrawal of ammonium resulted in an immediate and dramatic stimulation of K(+) influx in barley, indicating a shift from active to passive K(+) uptake at low [K(+)]ext and yielding fluxes as high as 36 µmol g (root fresh weight)(-1) h(-1) at 5 mm [K(+)]ext, among the highest transporter-mediated K(+) fluxes hitherto reported. This ammonium-withdrawal effect was also established in all Arabidopsis lines (the wild types, atakt1, athak5, and athak5 atakt1) at low [K(+)]ext, revealing the concerted involvement of several transport systems. The ammonium-withdrawal effect coincided with a suppression of K(+) efflux and a significant hyperpolarization of the plasma membrane in all genotypes except athak5 atakt1, could be sustained over 24 h, and resulted in increased tissue K(+) accumulation. We discuss key differences and similarities in K(+) acquisition between two important model systems and reveal novel aspects of K(+) transport in planta.

Figure 1.

The effects of various pharmacological and nutritional treatments, targeting either Shaker-like K⁺ channels and HAK/KUP/KT transporters (A, C, and E) or NSCCs (B, D, and F), on steady-state K⁺ influx in intact roots of barley seedlings grown in full nutrient medium at low (A and B), intermediate (C and D), and high (E and F) [K⁺]_ext and 10 mm [NH₄⁺]_ext. Fluxes are indicated on a root fresh weight basis. Asterisks denote different levels of significance between control and treatment pairs (*0.01 < P < 0.05, **0.001 < P < 0.01, ***P < 0.001; one-way ANOVA with Dunnett’s multiple comparison post hoc test). Each treatment represents the mean of four to 69 replicates. Error bars indicate se.

Figure 2.

The effects of various pharmacological and nutritional treatments on steady-state K⁺ influx in intact roots of Arabidopsis Col-0 wild type (A), WS wild type (B), atakt1 (C), athak5 (D), and athak5 atakt1 (E) grown in full nutrient medium at low [K⁺]_ext and 2 mm [NH₄⁺]_ext. Fluxes are indicated on a root fresh weight basis. Asterisks denote different levels of significance between control and treatment pairs (*0.01 < P < 0.05, **0.001 < P < 0.01, ***P < 0.001; one-way ANOVA with Dunnett’s multiple comparison post hoc test). Each treatment represents the mean of three to 14 replicates. Error bars indicate se.

Figure 3.

Responses of ⁴²K⁺ efflux from roots of intact barley seedlings to sudden application (see arrows) of various pharmacological and nutritional treatments. Plants were grown in full nutrient medium at low (A), intermediate (B), and high (C) [K⁺]_ext and 10 mm [NH₄⁺]_ext. Insets show responses of K⁺ efflux to sudden (see arrows) withdrawal of external NH₄⁺ and/or alkalinity during radiotracer uptake and elution periods. Numbers in parentheses indicate percentages of treated points differing significantly from the control (Student’s t test; P < 0.05). In the insets, axis labels are as in the main figures. Each plot represents the mean of three to seven replicates. Error bars indicate se. FW, Fresh weight. [See online article for color version of this figure.]

Figure 4.

The effects of various pharmacological and nutritional treatments on K⁺ influx stimulated due to NH₄⁺ withdrawal in intact roots of barley seedlings grown in full nutrient medium at low (A), intermediate (B), and high (C) [K⁺]_ext and 10 mm [NH₄⁺]_ext. Fluxes are indicated on a root fresh weight basis. Asterisks denote different levels of significance between –NH₄⁺ and treatment pairs (*0.01 < P < 0.05, **0.001 < P < 0.01, ***P < 0.001; one-way ANOVA with Dunnett’s multiple comparison post hoc test). Asterisks in parentheses denote levels of significance between control and –NH₄⁺ pairs (Student’s t test). Each treatment represents the mean of four to 69 replicates. Error bars indicate se.

Figure 5.

The effects of various pharmacological and nutritional treatments on K⁺ influx stimulated due to NH₄⁺ withdrawal in intact roots of Arabidopsis Col-0 wild type (A), WS wild type (B), atakt1 (C), athak5 (D), and athak5 atakt1 (E) grown in full nutrient medium at low [K⁺]_ext and 2 mm [NH₄⁺]_ext. Fluxes are indicated on a root fresh weight basis. Asterisks denote different levels of significance between –NH₄⁺ and treatment pairs (*0.01 < P < 0.05, **0.001 < P < 0.01, ***P < 0.001; one-way ANOVA with Dunnett’s multiple comparison post hoc test). Asterisks in parentheses denote levels of significance between control and –NH₄⁺ pairs (Student’s t test). Each treatment represents the mean of four to 14 replicates. Error bars indicate se.

Figure 6.

Responses of K⁺ influx to the duration of NH₄⁺ withdrawal in roots of intact barley seedlings grown under full nutrient medium, various [K⁺]_ext levels, and 10 mm [NH₄⁺]_ext. Fluxes are indicated on a root fresh weight basis. Each data point represents the mean of four replicates. Error bars indicate se. [See online article for color version of this figure.]

Figure 7.

Responses of tissue K⁺ content to the duration of NH₄⁺ withdrawal in barley seedlings grown under full nutrient medium at low (A), intermediate (B), and high (C) [K⁺]_ext and 10 mm [NH₄⁺]_ext. Content measurements are indicated on a root fresh weight basis. Each data point represents the mean of four replicates. Error bars indicate se. [See online article for color version of this figure.]

Figure 8.

Schematic overview of K⁺ uptake in plant roots under steady-state conditions (i.e. in the presence of high [millimolar] [NH₄⁺]_ext). Under low (0.0225 mm) [K⁺]_ext, K⁺ uptake is predominantly mediated by K⁺ channels (AtAKT1) in Arabidopsis (Col-0 wild type; A), whereas in barley (B), K⁺ channels do not operate and uptake is likely mediated by high-affinity transporters (HvHAK1), albeit at a residual capacity due to NH₄⁺-induced inhibition. Above intermediate K⁺ levels ([K⁺]_ext = 0.1125 mm), K⁺ channels do operate in barley, with further Cl⁻- and NO₃⁻-induced stimulations of K⁺ uptake observed at high K⁺ ([K⁺]_ext = 5 mm). In barley, K⁺ efflux is likely channel mediated at low and intermediate K⁺, although with varying sensitivities to Cs⁺, whereas K⁺ efflux is likely inoperative at high K⁺. K⁺ efflux in Arabidopsis is also likely channel mediated (Maathuis and Sanders, 1993). [K⁺]_cyt and resting ΔΨ_m values, when measured, are also listed. Asterisks refer to references as follows: *see Maathuis and Sanders (1993) and Halperin and Lynch (2003); **see Leigh and Wyn-Jones (1984), Walker et al. (1996), and Kronzucker et al. (2006). [See online article for color version of this figure.]