Molecular and physiological characterization of vacuolar disaccharide and polyol transporters and of vacuolar monosaccharide transporter-like proteins

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The tightly regulated transport of mono- and disaccharides and of cyclic and linear polyols into and out of the vacuole is indispensable for optimal growth and development and for stress tolerance in higher plants. In Arabidopsis at least 26 tonoplast (TP)-localized transporters are involved in the catalysis of these transport processes. So far, only few of these proteins have been functionally characterized, and, surprisingly, next to nothing is known on the mechanisms involved in the sorting of any of these transporters to the TP. We continued the characterization of a TP-localized inositol transporter (INT1) and of several SUC4-type sucrose transporters and we initiated the characterization of two TP-localized monosaccharide transporters: ERD6 and ERDL3 (synonym: ESL1). In cooperation with partners of the Research Unit we could functionally characterize the Arabidopsis SUC4 protein as a transporter that releases sucrose from the vacuole, and we provided a solution to contradictory data on the subcellular localization of SUC4 proteins from different species. Moreover, we could demonstrate that SUC4 is targeted to the TP via the adapter protein complex 3 (AP-3). For INT1, we identified and characterized a tonoplast-targeting sequence and revealed that this transporter uses a second, AP-3-independent route to the TP. Exchanging targeting signals allowed TP-targeting of SUC4 via the INT1-route in an ap-3 mutant. Now we aim to unravel these two sorting routes to the TP in detail. We will identify the sorting motif for the SUC4 route and the adapter complex for the INT1 route, we will determine the possible role of protein phosphorylation, and we will study all transporters analyzed in the Research Unit with respect to these two sorting routes. In parallel we will continue and finish the characterizations of two vacuolar monosaccharide transporters.