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

单糖和二糖以及环状和线性多元醇进出液泡的严格调节的运输对于高等植物的最佳生长和发育以及胁迫耐受性是必不可少的。在拟南芥中，至少有26个液泡膜（TP）定位的转运蛋白参与这些运输过程的催化。到目前为止，只有少数这些蛋白质的功能特征，令人惊讶的是，几乎没有什么是已知的机制，参与任何这些转运蛋白的TP排序。我们继续对TP定位的肌醇转运蛋白（INT 1）和几种SUC 4型蔗糖转运蛋白进行表征，并开始对两种TP定位的单糖转运蛋白：ERD 6和ERDL 3（同义词：ESL 1）进行表征。与研究单位的合作伙伴合作，我们可以功能上表征拟南芥SUC 4蛋白作为一种转运蛋白，从液泡中释放蔗糖，我们提供了一个解决方案，从不同物种的SUC 4蛋白的亚细胞定位的矛盾数据。此外，我们可以证明SUC 4通过衔接蛋白复合物3（AP-3）靶向TP。对于INT 1，我们确定并表征了一个液泡膜靶向序列，并揭示了这种转运蛋白使用第二个AP-3独立途径到达TP。交换靶向信号允许在ap-3突变体中通过INT 1途径TP靶向SUC 4。现在，我们的目标是解开这两个分类路线的TP详细。我们将确定SUC 4途径的分选基序和INT 1途径的衔接复合物，我们将确定蛋白磷酸化的可能作用，我们将研究研究单位分析的所有转运蛋白与这两种分选途径有关。同时，我们将继续并完成两个液泡单糖转运蛋白的表征。

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.