Plant Physiology Preview
Published on March 31, 2006; 10.1104/pp.106.079533
Received February 22, 2006
Returned for revision March 16, 2006
Accepted March 16, 2006
Identification of a Vacuolar Sucrose Transporter in Hordeum vulgare and Arabidopsis thaliana Mesophyll Cells by a Tonoplast Proteomic Approach
Anne Endler , Stefan Meyer , Silvia Schelbert , Thomas Schneider , Winfriede Weschke , Shaun W. Peters , Felix Keller , Sacha Baginsky , Enrico Martinoia , and Ulrike G. Schmidt *
University of Zurich, Institute of Plant Biology, 8008 Zurich, Switzerland; Institute of Plant Genetics and Crop Plant Research, 06466 Gatersleben, Germany; and Eidgenössische Technische Hochschule (ETH) Zurich, Institute of Plant Science and Functional Genomics Center Zurich, Swiss Federal Institute of Technology, 8092 Zurich, Switzerland
* Corresponding author; email: ulrike.schmidt{at}botinst.unizh.ch.
The vacuole is the main cellular storage pool, where sucrose accumulates to high concentrations. While a limited number of vacuolar membrane proteins such as V-type H+-ATPases and H+-PPases are well characterized, the majority of vacuolar transporters are still unidentified, among them the transporter(s) responsible for vacuolar sucrose uptake and release. In search of novel tonoplast transporters we used a proteomic approach, analyzing the tonoplast fraction of highly purified mesophyll vacuoles of the crop plant Hordeum vulgare (Hv). We identified 101 proteins, including 88 vacuolar and putative vacuolar proteins. The sucrose transporter (SUT) HvSUT2 was discovered among the 40 vacuolar proteins, which were previously not reported in Arabidopsis vacuolar proteomic studies. To confirm the tonoplast localization of this sucrose transporter we constructed and expressed GFP fusion proteins with HvSUT2 and its closest Arabidopsis homologue AtSUT4. Transient expression of HvSUT2-GFP and AtSUT4-GFP in Arabidopsis leaves and onion epidermal cells resulted in green fluorescence at the tonoplast, indicating that these sucrose transporters are indeed located at the vacuolar membrane. Using a microcapillary we selected mesophyll protoplasts from a leaf protoplast preparation and demonstrated unequivocally that, in contrast to the companion cell-specific AtSUC2, HvSUT2 and AtSUT4 are expressed in mesophyll protoplasts, suggesting that HvSUT2 and AtSUT4 are involved in transport and vacuolar storage of photosynthetically derived sucrose.
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