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Abstract

The K+ uptake pathways in yeast mitochondria are still undefined. Nonetheless, the K+-mediated mitochondrial swelling observed in the absence of phosphate (PO4) and in the presence of a respiratory substrate has led to propose that large K+ movements occur in yeast mitochondria. Thus, the uptake of K+ by isolated yeast mitochondria was evaluated. Two parallel experiments were conducted to evaluate K+ transport; these were mitochondrial swelling and the uptake of the radioactive K+ analog 86Rb+. The opening of the yeast mitochondrial unspecific channel (YMUC) was regulated by different PO4 concentrations. The high protein concentrations used to measure 86Rb+ uptake resulted in a slight stabilization of the transmembrane potential at 0.4 mM PO4 but not at 0 or 4 mM PO4. At 4 mM PO4 swelling was inhibited while, in contrast, 86Rb+ uptake was still observed. The results suggest that an energy-dependent K+ uptake mechanism was unmasked when the YMUC was closed. To further analyze the properties of this K+ uptake system, the Mg2+ and quinine sensitivity of both swelling and 86Rb+ uptake were evaluated. Under the conditions where the unspecific pore was closed, K+ transport sensitivity to Mg2+ and quinine increased. In addition, when Zn2+ was added as an antiport inhibitor, uptake of 86Rb+ increased. It is suggested that in yeast mitochondria, the K+ concentration is highly regulated by the equilibrium of uptake and exit of this cation through two specific transporters.

Sensitivity of the Mitochondrial Unspecific Channel ofSaccharomyces cerevisiaeto Butane-1,4-Bisphosphate, a Competitive Inhibitor of Fructose-1,6-Bisphosphate-Aldolase.

“…Since B1,4BP couples oxidative phosphorylation and promotes mitochondrial swelling, it was decided to evaluate the effects of B1,4BP on the transmembrane potential (ΔΨ), which is strongly dependent on the state of Sc MUC,i. e. as Sc MUC opens, ΔΨ decreases . At 4 mM Pi a stable ΔΨ was established while in 0.1 mM Pi (open Sc MUC) ΔΨ was depleted (Figure ).…”

Characterization of the respiration‐induced yeast mitochondrial permeability transition pore

“…The rate of the K + /H + exchange may not keep pace with the increased proton pumping at high respiratory substrate concentrations; therefore, a pH gradient could form, the yPTP would then open, and swelling would occur. Alternatively the rate of the K + /H + exchange may not keep up with the rate of electrophoretic K + entry accompanied by Cl − entry through the anion channel at high respiratory substrate concentrations (Castrejon, et al, 2002), resulting in swelling without yPTP opening. Since the combination of both P i and decavanadate, which keep the yPTP closed, partially inhibited swelling, both of these mechanisms likely contribute to the total extent of swelling under these conditions.…”

“…They also identified and characterized K + uniport activities in yeast mitochondria (Manon and Guerin, 1993). Since K + uniport in both yeast and mammalian mitochondria is inhibited by quinine, Mg 2+ , and mersalyl, the process may have been conserved throughout evolution (Castrejon, et al, 1997; Castrejon, et al, 2002). In the presence of a physiological P i concentration, the addition of K + to respiring yeast mitochondria causes a protonophoric effect resulting from K + entering mitochondria electrophoretically and exiting by the K + /H + exchange (Castrejon, et al, 1997; Manon, et al, 1995).…”

Methods of Measuring Mitochondrial Potassium Channels: A Critical Assessment

“…We omit flux measurements that use rubidium cations as a tracer for K + . The use of radioactive 86Rb + and 87Rb + to study mitochondrial potassium channels is uncommon [42] despite the widespread use of 86Rb + to study plasma membrane and intracellular potassium channels [43]; -Biochemical techniques, i.e., the use of unique mitochondrial properties such as swelling, respiration, and high membrane potential to track potassium fluxes via inner membrane channel proteins.…”