The process of sucrose transport was investigated in sealed putative tonoplast vesicles isolated from sugarbeet (Beta vulgaris L.) The partitioning of photosynthetic products represents a complex interaction between the source supply of assimilates and their uptake by competing sink regions (7 and references therein). The overall importance of the sink region in determining the partitioning ofphotosynthate is well documented (7,8); however, the factors which determine the competitive ability of various sinks are not well understood. A thorough knowledge of this process will be required in order to identify those factors amenable to either chemical or genetic modification for increased partitioning of photosynthates into the economically important part of the crop plant.In sugarbeet, sucrose produced in the leaves is translocated to the taproot sink and stored in the vacuole ofthe root parenchyma '

Sealed membrane vesicles were isolated from homogenates of sugarbeet (Beta valgaris L.) taproot by a combination of differential centrifugation, extraction with KI, and dextran gradient centrifugation. Relative to the KI-extracted microsomes, the content of plasma membranes, mitochondrial membranes, and Golgi membranes was much reduced in the final vesicle fraction. A component of ATPase activity that was inhibited by nitrate co-enriched with the capacity of the vesicles to form a steady state pH gradient during the purification procedure. This suggests that the nitrate-sensitive ATPase may be involved in driving H'-transport, and this is consistent with the observation that Hf-transport, in the final vesicle fraction was inhibited by nitrate. Proton transport in the sugarbeet vesicles was substrate specific for ATP, insensitive to sodium vanadate and oligomycin but was inhibited by diethylstilbestrol and N,N'-dicyclohexylcarbodiimide. The formation of a pH gradient in the vesicles was enhanced by halide ions in the sequence I-> Br-> Cl1 while P was inhibitory. These stimulatory effects occur from both a direct stimulation of the ATPase by anions and a reduction in the vesicle membrane potential. In the presence of Cl1, alkali cations reduce the pH gradient relative to that observed with bis-tris-propane, possibly by H'/ alkali cation exchange. Based upon the properties of the HW-transporting vesicles, it is proposed that they are most likely derived from the tonoplast so that this vesicle preparation would represent a convenient system for studying the mechanism of transport at this membrane boundary.Energy dependent, primary proton transport appears to be a ubiquitous property of higher plant cells (21 and references therein). The energy conserved in the electrochemical potential gradient of protons, the proton motive force, can then provide the driving force for the transport of other solutes such as sugars according to the chemiosmotic hypothesis (18). The latter process has been termed 'secondary' transport since the coupling to metabolic energy is indirect via an ion gradient (11 and references therein). Studies mary proton transport can occur both at the plasma membrane (cytoplasm to cell exterior) and at the tonoplast (cytoplasm to vacuole lumen) (27 and references therein). When membrane fractions enriched with plasma membrane (16) or tonoplast (15) are prepared, they are found to contain ATP hydrolytic activity which has been postulated to reflect the presence of ATP-fueled proton pumps responsible for carrying out these primary transport events (16,21,27).A significant advance to the study of membrane transport in higher plants came with the development of the methodology to isolate sealed membrane vesicles by Sze (25). The sealed vesicle system allowed the demonstration that these membrane-associated ATPases isolated from higher plant cells could directly transport protons (27). Several laboratories have since characterized ATPase-mediated proton transport in sealed vesicles thought to be d...