GLUT transporters family — physiological functions and role in development of malignancies

In order to perform their physiologic functions, polarized epithelial cells must target ion transport proteins to the appropriate domains of their plasma membranes. Molecular signals responsible for polarized sorting have been identified for several membrane proteins which span the bilayer once. Most ion transport proteins are polytopic, however, and little is known of the signals responsible for the targeting of this class of polypeptides. Members of the gamma-aminobutyric acid (GABA) transporter family are polytopic membrane proteins found endogenously in both epithelial cells and neurons. We have identified narrowly defined sequences which are required for the proper accumulation of two members of this transporter family in Madin-Darby canine kidney cells. The highly homologous GABA transporter isoforms, GAT-2 and GAT-3, localize to the basolateral and apical surfaces, respectively, when expressed stably in Madin-Darby canine kidney cells. We have generated deletion constructs and chimeric transporters composed of complimentary portions of GAT-2 and GAT-3. We find that information which directs their differential sorting is present in the C-terminal cytoplasmic tails of these two polypeptides. A sequence of 22 amino acids at the C terminus of GAT-2 is required for the transporter's basolateral distribution and is capable of directing GAT-3 to the basolateral surface when appended to the C terminus of this normally apical polypeptide. The deletion of 32 amino acids from the C terminus of GAT-3 causes this transporter to become mislocalized to both surfaces. Moreover, removal of the final three amino acids of GAT-3 (THF) similarly disrupts its apical sorting. The GAT-3 C-terminal sequence resembles motifs which interact with PDZ domains, raising the possibility that the steady state distribution of GAT-3 at the apical plasmalemmal surface requires a protein-protein interaction mediated by its extreme C-terminal cytoplasmic tail. These data provide the first characterization of a protein-based signal required for the apical distribution of a membrane protein.

Glycans are known to be fundamental for many cellular and physiological functions. Congenital disorders of glycosylation (CDG) currently encompassing over 160 subtypes, are characterized by glycan synthesis and/or processing defects. Despite the increasing number of CDG patients, therapeutic options remain very limited as our knowledge on glycan synthesis is fragmented. The emergence of CDG resulting from defects in ER/ Golgi homeostasis makes this even more difficult. SLC10A7 belongs to the SLC10 protein family, known as bile acid and steroid transport family, exhibiting a unique structure. It shows a ubiquitous expression and is linked to negative calcium regulation in cells. The mechanisms by which SLC10A7 deficiency leads to Golgi glycosylation abnormalities are unknown. The present study identifies major O-glycosylation defects in both SLC10A7 KO HAP1 cells and SLC10A7-CDG patient fibroblasts and reveals an increased ER and Golgi calcium contents. We also show that the abundance of COSMC and C1GALT1 is altered in SLC10A7-CDG patient cells, as well as the subcellular Golgi localization of the Ca2+-binding Cab45 protein. Finally, we demonstrate that supraphysiological manganese supplementation suppresses the deficient electrophoretic mobility of TGN46 by an aberrant transfer of GalNAc residues, and reveal COSMC Mn2+ sensitivity. These findings provide novel insights into the mechanisms of Golgi glycosylation defects in SLC10A7-deficient cells. They show that SLC10A7 is a key Golgi transmembrane protein maintaining the tight regulation of Ca2+ homeostasis in the ER and Golgi compartments, both essential for glycosylation.

Molecular cloning of an orphan transporter A new member of the neurotransmitter transporter family

Transporters in human platelets: physiologic function and impact for pharmacotherapy

In plants, sugar transporters are involved not only in long-distance transport, but also in sugar accumulations in sink cells. To identify members of sugar transporter gene families and to analyze their function in fruit sugar accumulation, we conducted a phylogenetic analysis of the Malus domestica genome. Expression profiling was performed with shoot tips, mature leaves, and developed fruit of “Gala” apple. Genes for sugar alcohol [including 17 sorbitol transporters (SOTs)], sucrose, and monosaccharide transporters, plus SWEET genes, were selected as candidates in 31, 9, 50, and 27 loci, respectively, of the genome. The monosaccharide transporter family appears to include five subfamilies (30 MdHTs, 8 MdEDR6s, 5 MdTMTs, 3 MdvGTs, and 4 MdpGLTs). Phylogenetic analysis of the protein sequences indicated that orthologs exist among Malus, Vitis, and Arabidopsis. Investigations of transcripts revealed that 68 candidate transporters are expressed in apple, albeit to different extents. Here, we discuss their possible roles based on the relationship between their levels of expression and sugar concentrations. The high accumulation of fructose in apple fruit is possibly linked to the coordination and cooperation between MdTMT1/2 and MdEDR6. By contrast, these fruits show low MdSWEET4.1 expression and a high flux of fructose produced from sorbitol. Our study provides an exhaustive survey of sugar transporter genes and demonstrates that sugar transporter gene families in M. domestica are comparable to those in other species. Expression profiling of these transporters will likely contribute to improving our understanding of their physiological functions in fruit formation and the development of sweetness properties.

The brain is considered to be a glucose-obligatory organ, that is neuronal and glial cells from the central nervous system (CNS) are dependent on glucose as a sole energy source1. The ability of various anatomical areas of the brain to utilize glucose has been shown to have a major impact on the control of certain physiological and behavioral functions of the brain. This has resulted in an increased desire to elucidate the cellular and molecular mechanisms involved in the transport, metabolism, and sensitivity to the glucose in the brain. Recent identification and characterization of a facilitative glucose transporter family have set a pace for research involving glucose uptake systems present in the brain cells.

Intracellular organization of insulin signaling and GLUT4 translocation.

The transporter associated with antigen processing like (TAPL, ABCB9) is a member of the ATP-binding cassette (ABC) transporter family. Moreover, TAPL belongs to the TAP family due to its high sequence homology to TAP1 and TAP2. TAPL forms a homodimer which is localized in lysosomes with a minor fraction in the ER. It functions as an ATP-dependent peptide transporter which shows a broad peptide specificity ranging from 6-mer up to 59-mer peptides. In contrast to TAP, TAPL transports peptides with low affinity but high efficiency. This review will briefly summarize current knowledge about the structural organization and possible physiological function of TAPL in antigen processing and presentation.

NRT1 proteins are H+-coupling nitrate transporters which belong to the family of peptide transporters (PTRs) and facilitate low and high affinity nitrate transport systems in a model plant Arabidopsis thaliana. In this study, we present the first inventory of the Cucumis sativus NRT1 family together with the transcriptional profile of CsNRT1 genes suggesting the physiological function of the family members in cucumber. Semiquantitative RT- PCR was used to analyze the level and organ-distribution of expression of CsNRT1 genes. The response of those CsNRT1s, whose transcripts were clearly detectable in vegetative tissues to different level of nitrate supply was examined through real-time PCR assays. The newly identified cucumber NRT1s were given the designation according to their homology to A. thaliana AtNRT1s. The comparison of the Arabidopsis and cucumber NRT gene families, similarly to the previous comparison of NRT1s in Arabidopsis, poplar and grasses, reveals some striking differences in genes’ structure and quantity. The putative function of particular CsNRT1 proteins is discussed, considering the results obtained here as well as the already published studies on A. thaliana NRT1 transporters.

Measuring ClC-Transporters on Solid Supported Membranes

The human multidrug resistance-associated protein (MRP) family currently has seven members. The ability of several of these membrane proteins to transport a wide range of anticancer drugs out of cells and their presence in many tumors make them prime suspects in unexplained cases of drug resistance, although proof that they contribute to clinical drug resistance is still lacking. Recent studies have begun to clarify the function of the MRP family members. MRPs are organic anion transporters; i.e., they transport anionic drugs, exemplified by methotrexate, and neutral drugs conjugated to acidic ligands, such as glutathione (GSH), glucuronate, or sulfate. However, MRP1, MRP2, and MRP3 can also cause resistance to neutral organic drugs that are not known to be conjugated to acidic ligands by transporting these drugs together with free GSH. MRP1 can even confer resistance to arsenite and MRP2 to cisplatin, again probably by transporting these compounds in complexes with GSH. MRP4 overexpression is associated with high-level resistance to the nucleoside analogues 9-(2-phosphonylmethoxyethyl) adenine and azidothymidine, both of which are used as anti-human immunodeficiency virus drugs. MRPs may, therefore, also have a role in resistance against nucleoside analogues used in cancer chemotherapy. Mice without Mrp1, a high-affinity leukotriene C(4) transporter, have an altered response to inflammatory stimuli but are otherwise healthy and fertile. MRP2 is the major transporter responsible for the secretion of bilirubin glucuronides into bile, and humans without MRP2 develop a mild liver disease known as the Dubin-Johnson syndrome. The physiologic functions of the other MRPs are not known. Whether long-term inhibition of MRPs in humans can be tolerated (assuming that suitable inhibitors will be found) remains to be determined.

The oligopeptide transporter (OPT) family contains nine members in Arabidopsis. While there is some evidence that AtOPTs mediate the uptake of tetra- and pentapeptides, OPT homologs in rice (Oryza sativa; OsGT1) and Indian mustard (Brassica juncea; BjGT1) have been described as transporters of glutathione derivatives. This study investigates the possibility that two members of the AtOPT family, AtOPT6 and AtOPT7, may also transport glutathione and its conjugates. Complementation of the hgt1met1 yeast double mutant by plant homologs of the yeast glutathione transporter HGT1 (AtOPT6, AtOPT7, OsGT1, BjGT1) did not restore the growth phenotype, unlike complementation by HGT1. By contrast, complementation by AtOPT6 restored growth of the hgt1 yeast mutant on a medium containing reduced (GSH) or oxidized glutathione as the sole sulfur source and induced uptake of [3H]GSH, whereas complementation by AtOPT7 did not. In these conditions, AtOPT6-dependent GSH uptake in yeast was mediated by a high affinity (Km = 400 microm) and a low affinity (Km = 5 mm) phase. It was strongly competed for by an excess oxidized glutathione and glutathione-N-ethylmaleimide conjugate. Growth assays of yeasts in the presence of cadmium (Cd) suggested that AtOPT6 may transport Cd and Cd/GSH conjugate. Reporter gene experiments showed that AtOPT6 is mainly expressed in dividing areas of the plant (cambium, areas of lateral root initiation). RNA blots on cell suspensions and real-time reverse transcription-PCR on Arabidopsis plants indicated that AtOPT6 expression is strongly induced by primisulfuron and, to a lesser extent, by abscisic acid but not by Cd. Altogether, the data show that the substrate specificity and the physiological functions of AtOPT members may be diverse. In addition to peptide transport, AtOPT6 is able to transport glutathione derivatives and metal complexes, and may be involved in stress resistance.

Glucose is the primary energy source for most mammalian cells and its transport is affected by a family of facilitative glucose transporters (GLUTs) encoded by the SLC2 gene. GLUT1 and GLUT3, highly expressed isoforms in the blood-brain barrier and neuronal membranes, respectively, are associated with multiple neurodevelopmental disorders including epilepsy, dyslexia, ADHD, and autism spectrum disorder (ASD). Dietary therapies, such as the ketogenic diet, are widely accepted treatments for patients with the GLUT1 deficiency syndrome, while ameliorating certain symptoms associated with GLUT3 deficiency in animal models. A ketogenic diet, high-fat diet, and calorie/energy restriction during prenatal and postnatal stages can also alter the placental and brain GLUTs expression with long-term consequences on neurobehavior. This review focuses primarily on the role of diet/energy perturbations upon GLUT isoform-mediated emergence of neurodevelopmental and neurodegenerative disorders.

Chapter 23 - Zinc Transporters in Health and Disease

Here we present two structures of the major facilitator (MFS) xylose transporter XylE from Escherichia coli in inward open and partially occluded inward open conformations. These structures provide key information about the transport cycle of XylE and the closely related human GLUT transporters. This is, to our knowledge, the first MFS transporter structure determined in more than one conformational state, which may establish XylE as an important MFS model protein.