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

Abstract

Cytosolic Ca 2+ concentrations ([Ca 2+] i) were determined in haploid and diploid cells of Saccharomyces cerevisiae, loaded with indo-1 and exposed to media containing a range of Ca 2+ concentrations. [Ca 2+] 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 Ca 2+ in the medium. Slightly higher levels of [Ca 2+] i were determined in cells exposed to 50 mM Ca 2+. Pre-incubation with metabolic inhibitors instead of glucose caused a reduction in cellular ATP levels and an impaired [Ca 2+] i homeostasis; [Ca 2+] i reached 800 nM in cells exposed to 10 mM CaCl 2. Cells of the Δ vma4 mutant strain, with no functional vacuolar H 1-ATPase, had elevated levels of [Ca 2+] i, reaching 1.8 μM when pre-incubated with glucose and exposed to 10 mM CaCl 2. Higher levels of [Ca 2+] 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 [Ca 2+] i-homeostasis and suggests the presence of an additional non-vacuolar ATP-requiring mechanism which contributes to keeping [Ca 2+] i at low levels.