Abstract
Sucrose transporters in the SUT family are important for phloem loading and sucrose uptake into sink tissues. The recent localization of type III SUTs AtSUT4 and HvSUT2 to the vacuole membrane suggests that SUTs also function in vacuolar sucrose transport. The transport mechanism of type III SUTs has not been analyzed in detail. LjSUT4, a type III sucrose transporter homolog from Lotus japonicus, is expressed in nodules and its transport activity has not been previously investigated. In this report, LjSUT4 was expressed in Xenopus oocytes and its transport activity assayed by two-electrode voltage clamping. LjSUT4 transported a range of glucosides including sucrose, salicin, helicin, maltose, sucralose and both α- and β-linked synthetic phenyl glucosides. In contrast to other sucrose transporters, LjSUT4 did not transport the plant glucosides arbutin, fraxin and esculin. LjSUT4 showed a low affinity for sucrose (K 0.5 = 16 mM at pH 5.3). In addition to inward currents induced by sucrose, other evidence also indicated that LjSUT4 is a proton-coupled symporter: 14C-sucrose uptake into LjSUT4-expressing oocytes was inhibited by CCCP and sucrose induced membrane depolarization in LjSUT4-expressing oocytes. A GFP-fusion of LjSUT4 localized to the vacuole membrane in Arabidopsis thaliana and in the roots and nodules of Medicago truncatula. Based on these results we propose that LjSUT4 functions in the proton-coupled uptake of sucrose and possibly other glucosides into the cytoplasm from the vacuole.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Aoki N, Hirose T, Scofield GN et al (2003) The sucrose transporter gene family in rice. Plant Cell Physiol 44:223–232. doi:10.1093/pcp/pcg030
Auriac MC, Timmers AC (2007) Nodulation studies in the model legume Medicago truncatula: advantages of using the constitutive EF1alpha promoter and limitations in detecting fluorescent reporter proteins in nodule tissues. Mol Plant Microbe Interact 20:1040–1047. doi:10.1094/MPMI-20-9-1040
Boisson-Dernier A, Chabaud M, Garcia F et al (2001) Agrobacterium rhizogenes-transformed roots of Medicago truncatula for the study of nitrogen-fixing and endomycorrhizal symbiotic associations. Mol Plant Microbe Interact 14:695–700. doi:10.1094/MPMI.2001.14.6.695
Boorer KJ, Loo DD, Frommer WB, Wright EM (1996) Transport mechanism of the cloned potato H+/sucrose cotransporter StSUT1. J Biol Chem 271:25139–25144. doi:10.1074/jbc.271.41.25139
Briskin DP, Thornley WR, Wyse RE (1985) Membrane transport in isolated vesicles from sugarbeet taproot: II. Evidence for a Sucrose/H+-antiport. Plant Physiol 78:871–875
Carpaneto A, Geiger D, Bamberg E et al (2005) Phloem-localized, proton-coupled sucrose carrier ZmSUT1 mediates sucrose efflux under the control of the sucrose gradient and the proton motive force. J Biol Chem 280:21437–21443. doi:10.1074/jbc.M501785200
Chandran D, Reinders A, Ward JM (2003) Substrate specificity of the Arabidopsis thaliana sucrose transporter AtSUC2. J Biol Chem 278:44320–44325. doi:10.1074/jbc.M308490200
Chang AB, Lin R, Keith Studley W et al (2004) Phylogeny as a guide to structure and function of membrane transport proteins. Mol Membr Biol 21:171–181. doi:10.1080/09687680410001720830
Chincinska IA, Liesche J, Krugel U et al (2008) Sucrose transporter StSUT4 from potato affects flowering, tuberization, and shade avoidance response. Plant Physiol 146:515–528. doi:10.1104/pp.107.112334
Curtis MD, Grossniklaus U (2003) A gateway cloning vector set for high-throughput functional analysis of genes in planta. Plant Physiol 133:462–469. doi:10.1104/pp.103.027979
Ehrhardt DW, Atkinson EM, Long SR (1992) Depolarization of alfalfa root hair membrane potential by Rhizobium meliloti nod factors. Science 256:998–1000. doi:10.1126/science.10744524
Endler A, Meyer S, Schelbert S et al (2006) Identification of a vacuolar sucrose transporter in barley and Arabidopsis mesophyll cells by a tonoplast proteomic approach. Plant Physiol 141:196–207. doi:10.1104/pp.106.079533
Farag MA, Huhman DV, Dixon RA, Sumner LW (2008) Metabolomics reveals novel pathways and differential mechanistic and elicitor-specific responses in phenylpropanoid and isoflavonoid biosynthesis in Medicago truncatula cell cultures. Plant Physiol 146:387–402. doi:10.1104/pp.107.108431
Flemetakis E, Dimou M, Cotzur D et al (2003) A sucrose transporter, LjSUT4, is up-regulated during Lotus japonicus nodule development. J Exp Bot 54:1789–1791. doi:10.1093/jxb/erg179
Gottwald JR, Krysan PJ, Young JC et al (2000) Genetic evidence for the in planta role of phloem-specific plasma membrane sucrose transporters. Proc Natl Acad Sci USA 97:13979–13984. doi:10.1073/pnas.250473797
Greutert H, Keller F (1993) Further evidence for stachyose and Sucrose/H+ antiporters on the tonoplast of Japanese artichoke (Stachys sieboldii) tubers. Plant Physiol 101:1317–1322
Hayashi S, Yagi K, Ishikawa T et al (2004) Emission of 2-phenylethanol from its β-D-glucopyranoside and the biogenesis of these compounds from [2H8]l-phenylalanine in rose flowers. Tetrahedron 60:7005–7013. doi:10.1016/j.tet.2003.10.130
Journet EP, El-Gachtouli N, Vernoud V et al (2001) Medicago truncatula ENOD11: a novel RPRP-encoding early nodulin gene expressed during mycorrhization in arbuscule-containing cells. Mol Plant Microbe Interact 14:737–748. doi:10.1094/MPMI.2001.14.6.737
Lopez M, Herrera-Cervera JA, Iribarne C et al (2008) Growth and nitrogen fixation in Lotus japonicus and Medicago truncatula under NaCl stress: nodule carbon metabolism. J Plant Physiol 165:641–650. doi:10.1016/j.jplph.2007.05.009
Ludewig U, von Wiren N, Frommer WB (2002) Uniport of NH4+ by the root hair plasma membrane ammonium transporter LeAMT1;1. J Biol Chem 277:13548–13555. doi:10.1074/jbc.M200739200
Marinova K, Pourcel L, Weder B et al (2007) The Arabidopsis MATE transporter TT12 acts as a vacuolar flavonoid/H+-antiporter active in proanthocyanidin-accumulating cells of the seed coat. Plant Cell 19:2023–2038. doi:10.1105/tpc.106.046029
Maurel C, Reizer J, Schroeder JI, Chrispeels MJ (1993) The vacuolar membrane protein gamma-TIP creates water specific channels in Xenopus oocytes. EMBO J 12:2241–2247
Nadwodnik J, Lohaus G (2008) Subcellular concentrations of sugar alcohols and sugars in relation to phloem translocation in Plantago major, Plantago maritima, Prunus persica, and Apium graveolens. Planta 227:1079–1089. doi:10.1007/s00425-007-0682-0
Neuhaus HE (2007) Transport of primary metabolites across the plant vacuolar membrane. FEBS Lett 581:2223–2226. doi:10.1016/j.febslet.2007.02.003
Peragine A, Yoshikawa M, Wu G et al (2004) SGS3 and SGS2/SDE1/RDR6 are required for juvenile development and the production of trans-acting siRNAs in Arabidopsis. Genes Dev 18:2368–2379. doi:10.1101/gad.1231804
Quick M, Loo DD, Wright EM (2001) Neutralization of a conserved amino acid residue in the human Na+/glucose transporter (hSGLT1) generates a glucose-gated H+ channel. J Biol Chem 276:1728–1734. doi:10.1074/jbc.M005521200
Reinders A, Ward JM (2001) Functional characterization of the alpha-glucoside transporter Sut1p from Schizosaccharomyces pombe, the first fungal homologue of plant sucrose transporters. Mol Microbiol 39:445–454. doi:10.1046/j.1365-2958.2001.02237.x
Reinders A, Sivitz AB, Hsi A et al (2006) Sugarcane ShSUT1: analysis of sucrose transport activity and inhibition by sucralose. Plant Cell Environ 29:1871–1880. doi:10.1111/j.1365-3040.2006.01563.x
Rosendahl L, Vance CP, Pedersen WB (1990) Products of dark CO2 fixation in pea root nodules support bacteroid metabolism. Plant Physiol 93:12–19
Schneider S, Beyhl D, Hedrich R, Sauer N (2008) Functional and physiological characterization of Arabidopsis INOSITOL TRANSPORTER1, a novel tonoplast-localized transporter for myo-inositol. Plant Cell 20:1073–1087. doi:10.1105/tpc.107.055632
Sivitz AB, Reinders A, Ward JM (2005) Analysis of the transport activity of barley sucrose transporter HvSUT1. Plant Cell Physiol 46:1666–1673. doi:10.1093/pcp/pci182
Sivitz AB, Reinders A, Johnson ME et al (2007) Arabidopsis sucrose transporter AtSUC9. High-affinity transport activity, intragenic control of expression, and early flowering mutant phenotype. Plant Physiol 143:188–198. doi:10.1104/pp.106.089003
Sivitz AB, Reinders A, Ward JM (2008) Arabidopsis sucrose transporter AtSUC1 is important for pollen germination and sucrose-induced anthocyanin accumulation. Plant Physiol 147:92–100. doi:10.1104/pp.108.118992
Tieman DM, Loucas HM, Kim JY et al (2007) Tomato phenylacetaldehyde reductases catalyze the last step in the synthesis of the aroma volatile 2-phenylethanol. Phytochemistry 68:2660–2669. doi:10.1016/j.phytochem.2007.06.005
Weise A, Barker L, Kuhn C et al (2000) A new subfamily of sucrose transporters, SUT4, with low affinity/high capacity localized in enucleate sieve elements of plants. Plant Cell 12:1345–1355
Weschke W, Panitz R, Sauer N et al (2000) Sucrose transport into barley seeds: molecular characterization of two transporters and implications for seed development and starch accumulation. Plant J 21:455–467. doi:10.1046/j.1365-313x.2000.00695.x
Zhou J, Theodoulou F, Sauer N et al (1997) A kinetic model with ordered cytoplasmic dissociation for SUC1, an Arabidopsis H+/sucrose cotransporter expressed in Xenopus oocytes. J Membr Biol 159:113–125. doi:10.1007/s002329900275
Zhou Y, Qu H, Dibley KE et al (2007) A suite of sucrose transporters expressed in coats of developing legume seeds includes novel pH-independent facilitators. Plant J 49:750–764. doi:10.1111/j.1365-313X.2006.03000.x
Acknowledgements
The LjSUT4 cDNA clone in vector pSPORT was provided by Panagiotis Katinakis and Emmanouil Flemetakis, Agricultural University of Athens, Greece. We are grateful to Mark A. Sanders and Tracy E. Anderson of the College of Biological Sciences Imaging Center at the University of Minnesota for their help with confocal microscopy. We thank Elison Blancaflor (Noble Foundation) and Michael Knoblauch (Washington State University) for their helpful suggestions on microscopy. This project was supported by Department of Energy Grant No. DE-FG02-07ER15886 (to JMW) and NSF Grant No. DBI-0421676 (to JSG). ABS gratefully acknowledges funding from the Doctoral Dissertation Fellowship (DDF) program of the University of Minnesota.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Reinders, A., Sivitz, A.B., Starker, C.G. et al. Functional analysis of LjSUT4, a vacuolar sucrose transporter from Lotus japonicus . Plant Mol Biol 68, 289–299 (2008). https://doi.org/10.1007/s11103-008-9370-0
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s11103-008-9370-0