CALCIUM HOMEOSTASIS IN YEAST-CELLS EXPOSED TO HIGH-CONCENTRATIONS OF CALCIUM - ROLES OF VACUOLAR H+-ATPASE AND CELLULAR ATP

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 0.1 muM-10 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 DELTAvma4 mutant strain, with no functional vacuolar H+-ATPase, had elevated levels of [Ca2+]i, reaching 1.8 muM 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.