Effect of ABA on cytosolic pH and of ABA-like conditions on K⁺ channels of Arabidopsis bundle sheath cells

In their vein-to-mesophyll radial passage in the leaf, solutes and water encounter a selective barrier: a tight layer of bundle sheath cells (BSCs). We characterized the voltage-gated K⁺ channels {K_OUT (K⁺-release channels, SKOR, GORK), K_IN (K⁺-influx channels, KAT1, KAT2, AKT1) and K_IN-WEAK (AKT2)} of Arabidopsis thaliana BSCs under “drought-stress-like conditions” modeled on guard cells experiencing water limitation and signals from the drought-stress hormone ABA. We applied electrophysiology (patch clamp in a whole-cell configuration) to BSCs protoplasts and varied the external and intracellular (cytosolic) pH and free cytosolic Ca²⁺ concentration (pH_EXT, pH_CYT and [Ca²⁺]_CYT, respectively). Among the most prevalent K⁺ channels, the K_OUT and K_IN-WEAK channels, ABA-effect-like conditions (pH_EXT 7.5, pH_CYT 8, [Ca²⁺]_CYT 500 nM) increased the K_OUT channel activity relative to the “resting” conditions (pH_EXT 5.6, pH_CYT 7.5 and [Ca²⁺]_CYT 100 nM).

The effect of ABA (3 uM) on the pH_CYT of the BSCs was monitored using the fluorescent ratiometric pH probe SNARF1. ABA-induced pH_CYT elevation was abolished by the vacuolar H⁺-ATPase (VHA)-specific inhibitor, bafilomycin A1, indicating the underlying mechanism as ABA stimulation of the VHA.

Interestingly, among whole-plant K⁺ channels knockout mutants experiencing water shortage in a greenhouse, skor showed stress signs earlier than WT or the mutants kat1 and akt2, suggesting an involvement of the K_OUT channels in whole plant stress response. We hypothesize this behavior of the plants to reflect an underlying osmo-protective mechanism against drought-induced embolism: drought-->BSCs perception of drought signal-->activation of K_OUT channels-->K⁺ release-->xylem sap enrichment with K⁺-->water retention in xylem.