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Thierry Pourcher - One of the best experts on this subject based on the ideXlab platform.
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the sodium iodide symporter state of the art of its molecular characterization
Biochimica et Biophysica Acta, 2014Co-Authors: Elisabeth Darrouzet, Sabine Lindenthal, Didier Marcellin, Jeanluc Pellequer, Thierry PourcherAbstract:The sodium/iodide symporter (NIS or SLC5A5) is an intrinsic membrane protein implicated in iodide uptake into thyroid follicular cells. It plays a crucial role in iodine metabolism and thyroid regulation and its function is widely exploited in the diagnosis and treatment of benign and malignant thyroid diseases. A great effort is currently being made to develop a NIS-based gene therapy also allowing the radiotreatment of nonthyroidal tumors. NIS is also expressed in other tissues, such as salivary gland, stomach and mammary gland during lactation, where its physiological role remains unclear. The molecular identity of the thyroid iodide transporter was elucidated approximately fifteen years ago. It belongs to the superfamily of sodium/solute Symporters, SSS (and to the human transporter family, SLC5), and is composed of 13 transmembrane helices and 643 amino acid residues in humans. Knowledge concerning NIS structure/function relationship has been obtained by taking advantage of the high resolution structure of one member of the SSS family, the Vibrio parahaemolyticus sodium/galactose symporter (vSGLT), and from studies of gene mutations leading to congenital iodine transport defects (ITD). This review will summarize current knowledge regarding the molecular characterization of NIS.
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The sodium/iodide symporter: State of the art of its molecular characterization
Biochimica et Biophysica Acta:Biomembranes, 2014Co-Authors: Elisabeth Darrouzet, Sabine Lindenthal, Didier Marcellin, Jeanluc Pellequer, Thierry PourcherAbstract:The sodium/iodide symporter (NIS or SLC5A5) is an intrinsic membrane protein implicated in iodide uptake into thyroid follicular cells. It plays a crucial role in iodine metabolism and thyroid regulation and its function is widely exploited in the diagnosis and treatment of benign and malignant thyroid diseases. A great effort is currently being made to develop a NIS-based gene therapy also allowing the radiotreatment of nonthyroidal tumors. NIS is also expressed in other tissues, such as salivary gland, stomach and mammary gland during lactation, where its physiological role remains unclear. The molecular identity of the thyroid iodide transporter was elucidated approximately fifteen years ago. It belongs to the superfamily of sodium/solute Symporters, SSS (and to the human transporter family, SLC5), and is composed of 13 transmembrane helices and 643 amino acid residues in humans. Knowledge concerning NIS structure/function relationship has been obtained by taking advantage of the high resolution structure of one member of the SSS family, the Vibrio parahaemolyticus sodium/galactose symporter (vSGLT), and from studies of gene mutations leading to congenital iodine transport defects (ITD). This review will summarize current knowledge regarding the molecular characterization of NIS.
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distribution and dynamics of 99mtc pertechnetate uptake in the thyroid and other organs assessed by single photon emission computed tomography in living mice
Thyroid, 2010Co-Authors: Philippe R. Franken, Julien Guglielmi, Malick Koulibaly, Michel Defrise, Jacques Darcourt, Christian Vanhove, Thierry PourcherAbstract:Background: 99mTc pertechnetate is a well-known anion, used for clinical imaging of thyroid function. This gamma emitter is transported by the sodium iodide symporter but is not incorporated into thyroglobulin. Scintigraphy using 99mTc pertechnetate or 123iodide represents a powerful tool for the study of sodium iodide symporter activity in different organs of living animal models. However, in many studies that have been performed in mice, the thyroid could not be distinguished from the salivary glands. In this work, we have evaluated the use of a clinically dedicated single-photon emission computed tomography (SPECT) camera for thyroid imaging and assessed what improvements are necessary for the development of this technique. Methods: SPECT of the mouse neck region, with pinhole collimation and geometric calibration, was used for the individual measurement of 99mTc pertechnetate uptake in the thyroid and the salivary glands. Uptake in the stomach was studied by planar whole-body imaging. Uptake kinetics ...
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a 96 well automated radioiodide uptake assay for sodium iodide symporter inhibitors
Assay and Drug Development Technologies, 2007Co-Authors: Nathalie Lecatguillet, Thierry Pourcher, Goulven Merer, Roman Lopez, Bernard Rousseau, Yves AmbroiseAbstract:A high-throughput screening method based on radioiodide uptake in human embryonic kidney 293 cells expressing the human sodium/iodide symporter was developed. Central to assay development was a homogeneous cell culture in the 96-well microplate coupled with the use of scintillation proximity technology. The assay is fast and highly reproducible with a Z′ greater than 0.8. The automated procedure allows the screening of 4,000 compounds per day. Using this methodology, several known substrates of the sodium/iodide symporter were evaluated in a single day. Inhibition of iodide uptake was shown to follow the series PF6− > ClO4− > BF4− > SCN− >> NO3− > IO4− > N3− >> Br−, in accord with the literature. This method represents an initial approach to the search for inhibitors of iodide transport mediated by the sodium/iodide symporter.
Christine Ziegler - One of the best experts on this subject based on the ideXlab platform.
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substrate bound outward open state of the betaine transporter betp provides insights into na coupling
Nature Communications, 2014Co-Authors: Camilo Perez, Christine Ziegler, Belinda Faust, Ahmad Reza Mehdipour, Kevin A. FrancesconiAbstract:The Na(+)-coupled betaine symporter BetP shares a highly conserved fold with other sequence unrelated secondary transporters, for example, with neurotransmitter Symporters. Recently, we obtained atomic structures of BetP in distinct conformational states, which elucidated parts of its alternating-access mechanism. Here, we report a structure of BetP in a new outward-open state in complex with an anomalous scattering substrate, adding a fundamental piece to an unprecedented set of structural snapshots for a secondary transporter. In combination with molecular dynamics simulations these structural data highlight important features of the sequential formation of the substrate and sodium-binding sites, in which coordinating water molecules play a crucial role. We observe a strictly interdependent binding of betaine and sodium ions during the coupling process. All three sites undergo progressive reshaping and dehydration during the alternating-access cycle, with the most optimal coordination of all substrates found in the closed state.
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role of bundle helices in a regulatory crosstalk in the trimeric betaine transporter betp
Journal of Molecular Biology, 2011Co-Authors: Rebecca M Gartner, Camilo Perez, Caroline Koshy, Christine ZieglerAbstract:The Na(+)-coupled betaine symporter BetP regulates transport activity in response to hyperosmotic stress only in its trimeric state, suggesting a regulatory crosstalk between individual protomers. BetP shares the overall fold of two inverted structurally related five-transmembrane (TM) helix repeats with the sequence-unrelated Na(+)-coupled Symporters LeuT, vSGLT, and Mhp1, which are neither trimeric nor regulated in transport activity. Conformational changes characteristic for this transporter fold involve the two first helices of each repeat, which form a four-TM-helix bundle. Here, we identify two ionic networks in BetP located on both sides of the membrane that might be responsible for BetP's unique regulatory behavior by restricting the conformational flexibility of the four-TM-helix bundle. The cytoplasmic ionic interaction network links both first helices of each repeat in one protomer to the osmosensing C-terminal domain of the adjacent protomer. Moreover, the periplasmic ionic interaction network conformationally locks the four-TM-helix bundle between the same neighbor protomers. By a combination of site-directed mutagenesis, cross-linking, and betaine uptake measurements, we demonstrate how conformational changes in individual bundle helices are transduced to the entire bundle by specific inter-helical interactions. We suggest that one purpose of bundle networking is to assist crosstalk between protomers during transport regulation by specifically modulating the transition from outward-facing to inward-facing state.
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molecular basis of transport and regulation in the na betaine symporter betp
Nature, 2009Co-Authors: Susanne Ressl, Anke Terwisscha C Van Scheltinga, Clemens Vonrhein, Vera Ott, Christine ZieglerAbstract:Osmoregulated transporters sense intracellular osmotic pressure and respond to hyperosmotic stress by accumulation of osmolytes to restore normal hydration levels. Here we report the determination of the X-ray structure of a member of the family of betaine/choline/carnitine transporters, the Na+-coupled symporter BetP from Corynebacterium glutamicum, which is a highly effective osmoregulated uptake system for glycine betaine. Glycine betaine is bound in a tryptophan box occluded from both sides of the membrane with aromatic side chains lining the transport pathway. BetP has the same overall fold as three unrelated Na+-coupled Symporters. Whereas these are crystallized in either the outward-facing or the inward-facing conformation, the BetP structure reveals a unique intermediate conformation in the Na+-coupled transport cycle. The trimeric architecture of BetP and the break in three-fold symmetry by the osmosensing C-terminal helices suggest a regulatory mechanism of Na+-coupled osmolyte transport to counteract osmotic stress. Microorganisms need to adapt rapidly to extreme variations in salinity, temperature or osmolarity. Many of them counteract high osmolarity via the intracellular accumulation of osmolytes, highly polar, organic compounds that promote protein stability through unfavourable interactions with the unfolded state. High-affinity transporters are used to import osmolytes from extracellular sources and Ressl et al. report the determination of the X-ray structure of a member of the family of betaine/choline/carnitine transporters, the Na+-coupled symporter BetP, which is a highly effective osmoregulated uptake system for glycine betaine. The crystal structure shows BetP to have the same overall fold as other, unrelated Na+-coupled transporters, but it is captured in a unique intermediate conformation whereby the glycine betaine substrate is occluded from both sides of the membrane. The structure of BetP therefore provides new insights into both osmoregulation and the fundamental mechanisms of membrane transport. This paper presents the first X-ray structure of a member of the the betaine/choline/carnitine transporter family. This Na+-coupled symporter (BetP) has the same overall fold as other unrelated Na+-coupled transporters and is captured in an intermediate conformation, whereby the substrate (glycine betaine) is occluded from both sides of the membrane.
Tsuneo Saga - One of the best experts on this subject based on the ideXlab platform.
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single photon emission computed tomography of spontaneous liver metastasis from orthotopically implanted human colon cancer cell line stably expressing human sodium iodide symporter reporter gene
EJNMMI research, 2012Co-Authors: Masayuki Inubushi, Chika Murai, Hironobu Hata, Yoshimasa Kitagawa, Tsuneo SagaAbstract:Background We aimed to develop a mouse spontaneous liver metastasis model from an orthotopically implanted human colon cancer cell line stably expressing a human sodium/iodide symporter (NIS) reporter gene, which can be imaged with single-photon emission computed tomography (SPECT) using 99mTcO4−.
Jonathan A Javitch - One of the best experts on this subject based on the ideXlab platform.
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A partially-open inward-facing intermediate conformation of LeuT is associated with Na+ release and substrate transport
Nature Publishing Group, 2018Co-Authors: Daniel S Terry, Harel Weinstein, Jonathan A Javitch, Matthias Quick, Rachel A. Kolster, Michael V. Levine, George Khelashvili, Zhou Zhou, Scott C BlanchardAbstract:Neurotransmitter:sodium Symporters (NSS) modulate the duration and magnitude of signaling via the sodium-coupled reuptake of neurotransmitters. Here the authors describe quantitative single molecule imaging of ligand-induced, functional dynamics of both intracellular and extracellular surfaces of LeuT, further defining the mechanism for NSS transport
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a mechanism for intracellular release of na by neurotransmitter sodium Symporters
Nature Structural & Molecular Biology, 2014Co-Authors: Lina Malinauskaite, Jonathan A Javitch, Matthias Quick, Linda Reinhard, Joseph A Lyons, Hideaki Yano, Poul NissenAbstract:Crystal structures of MhsT, a bacterial member of the neurotransmitter/sodium symporter family, in an occluded, inward-facing state with bound sodium and substrate reveal conformational changes during the transport cycle that provide new insights into the mechanism of cytoplasmic sodium release.
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chloride binding site of neurotransmitter sodium Symporters
Proceedings of the National Academy of Sciences of the United States of America, 2013Co-Authors: Adriana Krassimirova Kantcheva, Harel Weinstein, Jonathan A Javitch, Matthias Quick, Annemarie Lund Winther, Sebastian Stolzenberg, Poul NissenAbstract:Neurotransmitter:sodium Symporters (NSSs) play a critical role in signaling by reuptake of neurotransmitters. Eukaryotic NSSs are chloride-dependent, whereas prokaryotic NSS homologs like LeuT are chloride-independent but contain an acidic residue (Glu290 in LeuT) at a site where eukaryotic NSSs have a serine. The LeuT-E290S mutant displays chloride-dependent activity. We show that, in LeuT-E290S cocrystallized with bromide or chloride, the anion is coordinated by side chain hydroxyls from Tyr47, Ser290, and Thr254 and the side chain amide of Gln250. The bound anion and the nearby sodium ion in the Na1 site organize a connection between their coordinating residues and the extracellular gate of LeuT through a continuous H-bond network. The specific insights from the structures, combined with results from substrate binding studies and molecular dynamics simulations, reveal an anion-dependent occlusion mechanism for NSS and shed light on the functional role of chloride binding.
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ion substrate dependent conformational dynamics of a bacterial homolog of neurotransmitter sodium Symporters
Nature Structural & Molecular Biology, 2010Co-Authors: Derek P Claxton, Harel Weinstein, Jonathan A Javitch, Matthias Quick, Fernanda Delmondes De Carvalho, Hassane S MchaourabAbstract:LeuT is a bacterial sodium/leucine symporter, related to neurotransmitter transporters targeted by antidepressant drugs. Now spin labeling and EPR analysis on the dynamics in the extracellular vestibule of LeuT reveal the conformational changes caused by Na+ and Leu binding that drive the transport cycle.
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single molecule dynamics of gating in a neurotransmitter transporter homologue
Nature, 2010Co-Authors: Yongfang Zhao, Harel Weinstein, Daniel S Terry, Scott C Blanchard, Jonathan A JavitchAbstract:Neurotransmitter:Na+ Symporters (NSS) remove neurotransmitters from the synapse in a reuptake process that is driven by the Na+ gradient. Drugs that interfere with this reuptake mechanism, such as cocaine and antidepressants, profoundly influence behaviour and mood. To probe the nature of the conformational changes that are associated with substrate binding and transport, we have developed a single-molecule fluorescence imaging assay and combined it with functional and computational studies of the prokaryotic NSS homologue LeuT. Here we show molecular details of the modulation of intracellular gating of LeuT by substrates and inhibitors, as well as by mutations that alter binding, transport or both. Our direct observations of single-molecule transitions, reflecting structural dynamics of the intracellular region of the transporter that might be masked by ensemble averaging or suppressed under crystallographic conditions, are interpreted in the context of an allosteric mechanism that couples ion and substrate binding to transport. The neurotransmitter:Na+ symporter (NSS) family stops cellular signalling by recapturing released neurotransmitters from the synapse in a reuptake process driven by the Na+ gradient. Drugs that interfere with this mechanism, such as cocaine and antidepressants, profoundly influence behaviour and mood. The conformational changes associated with substrate binding have been studied in the prokaryotic NSS homologue LeuT using single-molecule fluorescence imaging experiments and molecular dynamics simulations. The observed single-molecule transitions reflect the structural dynamics of the intracellular region of the transporter, and can be interpreted in the context of an allosteric mechanism coupling ion and substrate binding to transport. Neurotransmitter:Na+ Symporters (NSS) remove neurotransmitters from the synapse in a reuptake process that is driven by the Na+ gradient. Here, single-molecule fluorescence imaging assays have been combined with molecular dynamics simulations to probe the conformational changes that are associated with substrate binding and transport by a prokaryotic NSS homologue, LeuT. The findings are interpreted in the context of an allosteric mechanism that couples ion and substrate binding to transport.
Jurgen Seidel - One of the best experts on this subject based on the ideXlab platform.
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ectopic expression of the sodium iodide symporter enables imaging of transplanted cardiac stem cells in vivo by single photon emission computed tomography or positron emission tomography
Journal of the American College of Cardiology, 2008Co-Authors: John Terrovitis, Keng Fai Kwok, Riikka Lautamaki, James M Engles, Andreas S Barth, Eddy Kizana, Junichiro Miake, Michelle K Leppo, James Fox, Jurgen SeidelAbstract:Objectives: We examined the sodium-iodide symporter (NIS), which promotes in vivo cellular uptake of technetium 99m (99mTc) or iodine 124 (124I), as a reporter gene for cell tracking by single-phot...
