Calcium- and voltage-dependent ion channels inSaccharomyces cerevisiae

Abstract

Ion channels in both the tonoplast and the plasma membrane ofSaccharomyces cerevisiaehave been characterized at the single channel level by patch-clamp techniques. The predominant tonoplast channel is cation selective, has an open-channel conductance of 120 pS in 100 mM KCl, and conducts Na+or K+equally well, and Ca2+to a lesser extent. Its open probability (P„) is voltage-dependent, peaking at about — 80 mV (cytoplasm negative), and falling to near zero at + 8 0 mV. Elevated cytoplasmic Ca2+, alkaline cytoplasmic pH, and reducing agents activate the channel. The predominant plasma membrane channel is highly selective for K+over anions and other cations, and shows strong outward rectification of the time-averaged current-voltage curves in cell-attached experiments. In isolated inside-out patches with micromolar cytoplasmic Ca2+, this channel is activated by positive going membrane voltages: meanPois zero at negative membrane voltages and near unity at 100 mV. At moderate positive membrane voltages (20-40 mV), elevating cytoplasmic Ca2+activates the channel to open in bursts of several hundred milliseconds duration. At higher positive membrane voltages, however, elevating cytoplasmic Ca2+blocks the channel in a voltage-dependent fashion for periods of 2-3 ms. The frequency of these blocking events depends on cytoplasmic Ca2+and membrane voltage according to second-order kinetics. Alternative cations, such as Mg2+of Na+, block the yeast plasma-membrane K+channel in a similar but less pronounced manner.