Calcium homeostasis in yeast cells exposed to high concentrations of calcium Roles of vacuolar H+‐ATPase and cellular ATP

Abstract

Cytosolic Ca²⁺ concentrations ([Ca²⁺]_i) were determined in haploid and diploid cells of Saccharomyces cerevisiae, loaded with indo-1 and exposed to media containing a range of Ca²⁺ concentrations. [Ca²⁺]_i homeostasis was maintained at the 100–150 nM level in cells which were pre-incubated with glucose and exposed to O.1 μM–1O mM Ca²⁺ in the medium. Slightly higher levels of [Ca²⁺]_i were determined in cells exposed to 50 mM Ca²⁺. Pre-incubation with metabolic inhibitors instead of glucose caused a reduction in cellular ATP levels and an impaired [Ca²⁺]_i homeostasis; [Ca²⁺]_i reached 800 nM in cells exposed to 10 mM CaCl₂. Cells of the Δvma4 mutant strain, with no functional vacuolar H¹-ATPase, had elevated levels of [Ca²⁺]_i, reaching 1.8 μM when pre-incubated with glucose and exposed to 10 mM CaCl₂. Higher levels of [Ca²⁺]_i were measured in the mutant cells which were pre-incubated with metabolic inhibitors. This result indicates the central role of the vacuoles in maintaining [Ca²⁺]_i-homeostasis and suggests the presence of an additional non-vacuolar ATP-requiring mechanism which contributes to keeping [Ca²⁺]_i at low levels.