Several recent publications reported different subcellular localization of the sucrose transporters belonging to the SUT4 subfamily. The physiological function of the SUT4 sucrose transporters requires clarification, because down-regulation of the members of the SUT4 clade had different effects in rice, poplar, and potato. Here, we provide new data for the localization and function of the Solanaceous StSUT4 protein, further elucidating involvement in the onset of flowering, tuberization and in the shade avoidance syndrome of potato plants. Induction of an early flowering and a tuberization in the SUT4-inhibited potato plants correlates with increased sucrose export from leaves and increased sucrose and starch accumulation in terminal sink organs, such as developing tubers. SUT4 affects expression of the enzymes involved in gibberellin and ethylene biosynthesis, as well as the rate of ethylene biosynthesis in potato. In the SUT4-inhibited plants, the ethylene production no longer follows a diurnal rhythm. Thus it was concluded that StSUT4 controls circadian gene expression, potentially by regulating sucrose export from leaves. Furthermore, SUT4 expression affects clock-regulated genes such as StFT, StSOC1, and StCO, which might be also involved in a photoperiod-dependent tuberization. A model is proposed in which StSUT4 controls a phloem-mobile signaling molecule generated in leaves, which together with enhanced sucrose export affects developmental switches in apical meristems. SUT4 seems to link photoreceptor-perceived information about the light quality and day length with phytohormone biosynthesis and the expression of circadian-regulated genes.

Organization of proteins into complexes is crucial for many cellular functions. Recently, the SUT1 protein was shown to form homodimeric complexes, to be associated with lipid raft-like microdomains in yeast as well as in plants and to undergo endocytosis in response to brefeldin A. We therefore aimed to identify SUT1-interacting proteins that might be involved in dimerization, endocytosis, or targeting of SUT1 to raft-like microdomains. Therefore, we identified potato membrane proteins, which are associated with the detergent-resistant membrane (DRM) fraction. Among the proteins identified, we clearly confirmed StSUT1 as part of DRM in potato source leaves. We used the yeast two-hybrid split ubiquitin system (SUS) to systematically screen for interaction between the sucrose transporter StSUT1 and other membrane-associated or soluble proteins in vivo. The SUS screen was followed by immunoprecipitation using affinity-purified StSUT1-specific peptide antibodies and mass spectrometric analysis of co-precipitated proteins. A large overlap was observed between the StSUT1-interacting proteins identified in the co-immunoprecipitation and the detergent-resistant membrane fraction. One of the SUT1-interacting proteins, a protein disulfide isomerase (PDI), interacts also with other sucrose transporter proteins. A potential role of the PDI as escort protein is discussed.

Sucrose accumulation in leaves in response to various abiotic stresses suggests a specific role of this disaccharide for stress tolerance and adaptation. The high-affinity transporter StSUT1 undergoes substrate-induced endocytosis presenting the question as to whether altered sucrose accumulation in leaves in response to stresses is also related to enhanced endocytosis or altered activity of the sucrose transporter. StSUT1 is known to interact with several stress-inducible proteins; here we investigated whether one of the interacting candidates, StPDI1, affects its subcellular localization in response to stress: StPDI1 expression is induced by ER-stress and salt. Both proteins, StSUT1 and StPDI1, were found in the detergent resistant membrane (DRM) fraction, and this might affect internalization. Knockdown of StPDI1 expression severely affects abiotic stress tolerance of transgenic potato plants. Analysis of these plants does not reveal modified subcellular localization or endocytosis of StSUT1, but rather a disturbed redox homeostasis, reduced detoxification of reactive oxygen species and effects on primary metabolism. Parallel observations with other StSUT1-interacting proteins are discussed. The redox status in leaves seems to be linked to the sugar status in response to various stress stimuli and to play a role in stress tolerance.

The fluorescence-based multi-analyte chip platform for the analysis of estrogenic and anti-estrogenic substances is a new in vitro tool for the high throughput screening of environmental samples. In contrast to existing tools, the chip investigates the complex action of xenoestrogens in a human cell model by characterizing protein expression. It allows for the quantification of 10 proteins secreted by MCF-7 cells, representing various biological and pathological endpoints of endocrine action and distinguishing between estrogen- and anti-estrogen-dependent secretion of proteins. Distinct protein secretion patterns of the cancer cell line after exposure to known estrogen receptor agonists ß-estradiol, bisphenol A, genistein, and nonylphenol as well as antagonists fulvestrant and tamoxifen demonstrate the potential of the chip. Stimulation of cells with Interleukin-1ß shifts concentrations of low abundant biomarkers towards the working range of the chip. In the non-stimulated cell culture, Matrix Metalloproteinase 9 (MMP-9) and Vascular Endothelial Growth Factor (VEGF) show differences upon treatment with antagonists and agonists of the estrogen receptor. In stimulated MCF-7 cells challenged with receptor agonists secretion of Monocyte Chemoattractant Protein (MCP-1), Interleukin-6 (IL-6), Rantes, and Interleukin-8 (IL-8) significantly decreases. In parallel, the proliferating effect of endocrine-disrupting substances in MCF-7 cells is assessed in a proliferation assay based on resazurin. Using ethanol as a solvent for test substances increases the background of proliferation and secretion experiments, while using dimethyl sulfoxide (DMSO) does not show any adverse effects. The role of the selected biomarkers in different physiological processes such as cell development, reproduction, cancer, and metabolic syndrome makes the chip an excellent tool for either indicating endocrine-disrupting effects in food and environmental samples, or for screening the effect of xenoestrogens on a cellular and molecular level.

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