SUT Sucrose and MST Monosaccharide Transporter Inventory of the Selaginella Genome

Most metazoa use hexose transporters to acquire hexoses from their diet and as a transport form for distributing carbon and energy within their bodies; insects use trehalose, and plants use sucrose as their major form for translocation. Plant genomes contain at least three families of mono- and disaccharide transporters: monosaccharide/polyol transporters that are evolutionary closely related to the yeast and human glucose transporters, sucrose transporters of the SUT family, which similar to the hexose transporters belong to the major facilitator superfamily, but share only minimal amino acid sequence homology with the hexose transporters, and the family of SWEET sugar transporters conserved between animals and plants. Recently, the genome sequence of the spikemoss Selaginella has been determined. In order to study the evolution of sugar transport in plants, we carefully annotated of the complement of sugar transporters in Selaginella. We review the current knowledge regarding sugar transport in spikemoss and provide phylogenetic analyses of the complement of MST and SUT homologs in Selaginella (and Physcomitrella)

Development of a fluorescent nanosensor for ribose

AbstractTo analyze ribose uptake and metabolism in living cells, nanosensors were engineered by flanking the Escherichia coli periplasmic ribose binding protein with two green fluorescent protein variants. Following binding of ribose, fluorescence resonance energy transfer decreased with increasing ribose concentration. Five affinity mutants were generated covering binding constants between 400 nM and 11.7 mM. Analysis of nanosensor response in COS-7 cells showed that free ribose accumulates in the cell and is slowly metabolized. Inhibitor studies suggest that uptake is mediated by a monosaccharide transporter of the GLUT family, however, ribose taken up into the cell was not or only slowly released, indicating irreversibility of uptake

Amino Acid Transporter Inventory of the Selaginella Genome

Amino acids play fundamental roles in a multitude of functions including protein synthesis, hormone metabolism, nerve transmission, cell growth, production of metabolic energy, nucleobase synthesis, nitrogen metabolism, and urea biosynthesis. Selaginella as a member of the lycophytes is part of an ancient lineage of vascular plants that had arisen ∼400 million years ago. In angiosperms, which have attracted most of the attention for nutrient transport so far, we have been able to identify many of the key transporters for nitrogen. Their role is not always fully clear, thus an analysis of Selaginella as a representative of an ancient vascular plant may help shed light on the evolution and function of these diverse transporters. Here we annotated and analyzed the genes encoding putative transporters involved in cellular uptake of amino acids present in the Selaginella genome

Interactions between co-expressed Arabidopsis sucrose transporters in the split-ubiquitin system

BACKGROUND: The Arabidopsis genome contains nine sucrose transporter paralogs falling into three clades: SUT1-like, SUT2 and SUT4. The carriers differ in their kinetic properties. Many transport proteins are known to exist as oligomers. The yeast-based split ubiquitin system can be used to analyze the ability of membrane proteins to interact. RESULTS: Promoter-GUS fusions were used to analyze the cellular expression of the three transporter genes in transgenic Arabidopsis plants. All three fusion genes are co-expressed in companion cells. Protein-protein interactions between Arabidopsis sucrose transporters were tested using the split ubiquitin system. Three paralogous sucrose transporters are capable of interacting as either homo- or heteromers. The interactions are specific, since a potassium channel and a glucose transporter did not show interaction with sucrose transporters. Also the biosynthetic and metabolizing enzymes, sucrose phosphate phosphatase and sucrose synthase, which were found to be at least in part bound to the plasma membrane, did not specifically interact with sucrose transporters. CONCLUSIONS: The split-ubiquitin system provides a powerful tool to detect potential interactions between plant membrane proteins by heterologous expression in yeast, and can be used to screen for interactions with membrane proteins as baits. Like other membrane proteins, the Arabidopsis sucrose transporters are able to form oligomers. The biochemical approaches are required to confirm the in planta interaction

Fusion to GFP blocks intercellular trafficking of the sucrose transporter SUT1 leading to accumulation in companion cells

BACKGROUND: Plant phloem consists of an interdependent cell pair, the sieve element / companion cell complex. Sucrose transporters are localized to enucleate sieve elements (SE), despite being transcribed in companion cells (CC). Due to the high turnover of SUT1, sucrose transporter mRNA or protein must traffic from CC to SE via the plasmodesmata. Localization of SUT mRNA at plasmodesmatal orifices connecting CC and SE suggests RNA transport, potentially mediated by RNA binding proteins. In many organisms, polar RNA transport is mediated through RNA binding proteins interacting with the 3'-UTR and controlling localized protein synthesis. To study mechanisms for trafficking of SUT1, GFP-fusions with and without 3'-UTR were expressed in transgenic plants. RESULTS: In contrast to plants expressing GFP from the strong SUC2 promoter, in RolC-controlled expression GFP is retained in companion cells. The 3'-UTR of SUT1 affected intracellular distribution of GFP but was insufficient for trafficking of SUT1, GFP or their fusions to SEs. Fusion of GFP to SUT1 did however lead to accumulation of SUT1-GFP in the CC, indicating that trafficking was blocked while translational inhibition of SUT1 mRNA was released in CCs. CONCLUSION: A fusion with GFP prevents targeting of the sucrose transporter SUT1 to the SE while leading to accumulation in the CC. The 3'-UTR of SUT1 is insufficient for mobilization of either the fusion or GFP alone. It is conceivable that SUT1-GFP protein transport through PD to SE was blocked due to the presence of GFP, resulting in retention in CC particles. Alternatively, SUT1 mRNA transport through the PD could have been blocked due to insertion of GFP between the SUT1 coding sequence and 3'-UTR

Exact Lagrangian submanifolds in simply-connected cotangent bundles

We consider exact Lagrangian submanifolds in cotangent bundles. Under certain additional restrictions (triviality of the fundamental group of the cotangent bundle, and of the Maslov class and second Stiefel-Whitney class of the Lagrangian submanifold) we prove such submanifolds are Floer-cohomologically indistinguishable from the zero-section. This implies strong restrictions on their topology. An essentially equivalent result was recently proved independently by Nadler, using a different approach.Comment: 28 pages, 3 figures. Version 2 -- derivation and discussion of the spectral sequence considerably expanded. Other minor change

The homotopy type of the loops on (n−1)(n-1)-connected (2n+1)(2n+1)-manifolds

For $n\geq 2$ we compute the homotopy groups of $(n-1)$-connected closed manifolds of dimension $(2n+1)$. Away from the finite set of primes dividing the order of the torsion subgroup in homology, the $p$-local homotopy groups of $M$ are determined by the rank of the free Abelian part of the homology. Moreover, we show that these $p$-local homotopy groups can be expressed as a direct sum of $p$-local homotopy groups of spheres. The integral homotopy type of the loop space is also computed and shown to depend only on the rank of the free Abelian part and the torsion subgroup.Comment: Trends in Algebraic Topology and Related Topics, Trends Math., Birkhauser/Springer, 2018. arXiv admin note: text overlap with arXiv:1510.0519

Animal tissue-based quantitative comparison of dual-energy CT to SPR conversion methods using high-resolution gel dosimetry

Dual-energy computed tomography (DECT) has been shown to allow for more accurate ion therapy treatment planning by improving the estimation of tissue stopping power ratio (SPR) relative to water, among other tissue properties. In this study, we measured and compared the accuracy of SPR values derived using both dual- and single-energy CT (SECT) based on different published conversion algorithms. For this purpose, a phantom setup containing either fresh animal soft tissue samples (beef, pork) and a water reference or tissue equivalent plastic materials was designed and irradiated in a clinical proton therapy facility. Dosimetric polymer gel was positioned downstream of the samples to obtain a three-dimensional proton range distribution with high spatial resolution. The mean proton range in gel for each tissue relative to the water sample was converted to a SPR value. Additionally, the homogeneous samples were probed with a variable water column encompassed by two ionization chambers to benchmark the SPR accuracy of the gel dosimetry. The SPR values measured with both methods were consistent with a mean deviation of 0.2%, but the gel dosimetry captured range variations up to 5 mm within individual samples. Across all fresh tissue samples the SECT approach yielded significantly greater mean absolute deviations from the SPR deduced using gel range measurements, with an average difference of 1.2%, compared to just 0.3% for the most accurate DECT-based algorithm. These results show a significant advantage of DECT over SECT for stopping power prediction in a realistic setting, and for the first time allow to compare a large set of methods under the same conditions