Accumulation of sucrose in vacuoles isolated from red beet tissue

Vacuoles were isolated from red beets (Beta vulgaris L.) by slicing the tissue and separated using a discontinuous dextran gradient centrifugation. The uptake of sucrose against a concentration gradient into the dextran-impermeable [(3)H]-H2O space of these organelles was studied using silicone layer filtering centrifugation on both fluorometric and (14)C-measurement of sucrose. The rate is 24 nmol sucrose (unit betacyanin)(-1) h(-1) and appears to be stimulated by ATP to an uptake rate of 34 nmol. Control experiments with slices cut from red beet tissue and incubated with [(14)C]sucrose gave comparable results. An ATPase activity dependent on both Mg(2+) and K(+) seems to be localized at the inner surface of the tonoplast. This activity is strongly inhibited by EDAC and tartrate and there is no effect of oligomycin, whereas a slight stimulation was caused by DCCD.