Abstract

Potassium (K⁺) channels mediating important physiological functions are characterized by a common pore–forming (P) domain. We report the cloning and functional analysis of the first higher plant outward rectifying K⁺ channel (KCO1) from Arabidopsis thaliana. KCO1 belongs to a new class of 'two-pore' K⁺ channels recently described in human and yeast. KCO1 has four putative transmembrane segments and tandem calcium-binding EF-hand motifs. Heterologous expression of KCO1 in baculovirus-infected insect (Spodoptera frugiperda) cells resulted in outwardly rectifying, K⁺-selective currents elicited by depolarizing voltage pulses in whole-cell measurements. Activation of KCO1 was strongly dependent on the presence of nanomolar concentrations of cytosolic free Ca²⁺ [Ca²⁺]_cyt. No K⁺ currents were detected when [Ca²⁺]_cyt was adjusted to <150 nM. However, KCO1 strongly activated at increasing [Ca²⁺]_cyt, with a saturating activity observed at 300 nM [Ca²⁺]_cyt. KCO1 single channel analysis on excised membrane patches, resulting in a single channel conductance of 64 pS, confirmed outward rectification as well as Ca²⁺-dependent activation. These data suggest a direct link between calcium-mediated signaling processes and K⁺ ion transport in higher plants. The identification of KCO1 as the first plant K⁺ outward channel opens a new field of structure–function studies in plant ion channels.