Transmembrane Sodium Gradient Research Articles

Blocking Aδ- and C-fiber neural transmission by sub-kilohertz peripheral nerve stimulation.

We recently showed that sub-kilohertz electrical stimulation of the afferent somata in the dorsal root ganglia (DRG) reversibly blocks afferent transmission. Here, we further investigated whether similar conduction block can be achieved by stimulating the nerve trunk with electrical peripheral nerve stimulation (ePNS). We explored the mechanisms and parameters of conduction block by ePNS via ex vivo single-fiber recordings from two somatic (sciatic and saphenous) and one autonomic (vagal) nerves harvested from mice. Action potentials were evoked on one end of the nerve and recorded on the other end from teased nerve filaments, i.e., single-fiber recordings. ePNS was delivered in the middle of the nerve trunk using a glass suction electrode at frequencies of 5, 10, 50, 100, 500, and 1000 Hz. Suprathreshold ePNS reversibly blocks axonal neural transmission of both thinly myelinated Aδ-fiber axons and unmyelinated C-fiber axons. ePNS leads to a progressive decrease in conduction velocity (CV) until transmission blockage, suggesting activity-dependent conduction slowing. The blocking efficiency is dependent on the axonal conduction velocity, with Aδ-fibers efficiently blocked by 50-1000 Hz stimulation and C-fibers blocked by 10-50 Hz. The corresponding NEURON simulation of action potential transmission indicates that the disrupted transmembrane sodium and potassium concentration gradients underly the transmission block by the ePNS. The current study provides direct evidence of reversible Aδ- and C-fiber transmission blockage by low-frequency (<100 Hz) electrical stimulation of the nerve trunk, a previously overlooked mechanism that can be harnessed to enhance the therapeutic effect of ePNS in treating neurological disorders.

Mutation in Drosophila concentrative nucleoside transporter 1 alters spermatid maturation and mating behavior.

Concentrative nucleoside transporters (Cnts) are unidirectional carriers that mediate the energy-costly influx of nucleosides driven by the transmembrane sodium gradient. Cnts are transmembrane proteins that share a common structural organization and are found in all phyla. Although there have been studies on Cnts from a biochemical perspective, no deep research has examined their role at the organismal level. Here, we investigated the role of the Drosophila melanogaster cnt1 gene, which is specifically expressed in the testes. We used the CRISPR/Cas9 system to generate a mutation in the cnt1 gene. The cnt1 mutants exhibited defects in the duration of copulation and spermatid maturation, which significantly impaired male fertility. The most striking effect of the cnt1 mutation in spermatid maturation was an abnormal structure of the sperm tail, in which the formation of major and minor mitochondrial derivatives was disrupted. Our results demonstrate the importance of cnt1 in male fertility and suggest that the observed defects in mating behavior and spermatogenesis are due to alterations in nucleoside transport and associated metabolic pathways.

Investigating the Mechanism of Sodium Binding to SERT Using Direct Simulations.

The serotonin transporter (SERT) terminates neurotransmission by transporting serotonin from the synapse into the pre-synaptic nerve terminal. Altered SERT function leads to several neurological diseases including depression, anxiety, mood disorders, and attention deficit hyperactivity disorders (ADHD). Accordingly SERT is the target for their pharmacological treatments, but also targeted by multiple drugs of abuse. Transport of serotonin by SERT is energized by the transmembrane electrochemical gradient of sodium. We used extensive molecular dynamics simulations to investigate the process of sodium binding to SERT, which is the first step in the transport cycle that leads to serotonin uptake. Comparing data from 51 independent simulations, we find a remarkably well-defined path for sodium entry and could identify two transient binding sites, while observing binding kinetics that are comparable to experimental data. Importantly, the structure and dynamics of the sodium binding sites indicate that sodium binding is accompanied by an induced-fit mechanism that leads to new conformations and reduces local dynamics.

It takes two—or more—to tango: Revisiting the role of dopamine transporter oligomerization

The dopamine transporter utilizes the transmembrane sodium gradient to mediate reuptake of dopamine from the extracellular space. The dopamine transporter can form dimers and possibly also higher order structures in the plasma membrane, and this oligomerization has been implicated in both trafficking and transport. However, we still do not fully understand its biological importance. A study by Sorkina et al. now describes a series of small molecules that link transporter conformation to oligomerization and endocytosis, providing an interesting step forward in an intricate dance.

Molecular Mechanisms of the Redox Regulation of the Na,K-ATPase

This review considers the molecular mechanisms involved in the redox regulation of the Na,K-ATPase. The enzyme creates a transmembrane gradient of sodium and potassium ions, which is necessary for the vital activity of all animal cells, and acts as a receptor of cardiotonic steroids (CTSs), which regulate cell proliferation and apoptosis. The function of the Na,K-ATPase depends on the cell’s redox status. Although oxidative stress was initially found to inhibit the enzyme, it is clear now that the redox regulation of the Na,K-ATPase activity is a complex process that cannot be explained only by oxidative damage to the protein. Na,K-ATPase activity is maximal at physiological oxygen concentrations and decreases by both hypoxia and hyperoxia, as well as due to decrease or increase of intracellular glutathione concentrations. Thus, a specific range of redox conditions provides maximal activity of the Na,K-ATPase. Now it is obvious that a disturbance of the Na,K-ATPase activity in a number of pathologies such as hypoxia, ischemia, diabetes, Alzheimer’s disease is associated with a change in redox status in the cells. The receptor function of the Na,K-ATPase also depends on the cell redox status and it isshould be taken into account when studying the effects of cardiotonic steroids on cells and tissues. The very special point of this review is the redox modifications of thiol groups in Na,K-ATPase subunits and the regulatory processes in which they are involved in normal and pathological conditions. Insight into the molecular mechanisms of redox regulation provides a better understanding of what is necessary for preventing Na,K-ATPase dysfunction in pathological conditions and thus reducing cell damage.

SLC6 transporter oligomerization

Transporters of the solute carrier 6 (SLC6) family mediate the reuptake of neurotransmitters such as dopamine, norepinephrine, serotonin, GABA, and glycine. SLC6 family members are 12 transmembrane helix‐spanning proteins that operate using the transmembrane sodium gradient for transport. These transporters assume various quaternary arrangements ranging from monomers to complex stoichiometries with multiple subunits. Dopamine and serotonin transporter oligomerization has been implicated in trafficking of newly formed proteins from the endoplasmic reticulum to the plasma membrane with a pre‐fixed assembly. Once at the plasma membrane, oligomers are kept fixed in their quaternary assembly by interaction with phosphoinositides. While it remains unclear how oligomer formation precisely affects physiological transporter function, it has been shown that oligomerization supports the activity of release‐type psychostimulants. Most recently, single molecule microscopy experiments unveiled that the stoichiometry differs between individual members of the SLC6 family. The present overview summarizes our understanding of the influence of plasma membrane constituents on transporter oligomerization, describes the known interfaces between protomers and discusses open questions.

Ouabain affects cell migration via Na,K-ATPase-p130cas and via nucleus-centrosome association.

Na,K-ATPase is a membrane protein that catalyzes ATP to maintain transmembrane sodium and potassium gradients. In addition, Na,K-ATPase also acts as a signal-transducing receptor for cardiotonic steroids such as ouabain and activates a number of signalling pathways. Several studies report that ouabain affects cell migration. Here we used ouabain at concentrations far below those required to block Na,K-ATPase pump activity and show that it significantly reduced RPE cell migration through two mechanisms. It causes dephosphorylation of a 130 kD protein, which we identify as p130cas. Src is involved, because Src inhibitors, but not inhibitors of other kinases tested, caused a similar reduction in p130cas phosphorylation and ouabain increased the association of Na,K-ATPase and Src. Knockdown of p130cas by siRNA reduced cell migration. Unexpectedly, ouabain induced separation of nucleus and centrosome, also leading to a block in cell migration. Inhibitor and siRNA experiments show that this effect is mediated by ERK1,2. This is the first report showing that ouabain can regulate cell migration by affecting nucleus-centrosome association.

An amphipathic α-helix directs palmitoylation of the large intracellular loop of the sodium/calcium exchanger

The electrogenic sodium/calcium exchanger (NCX) mediates bidirectional calcium transport controlled by the transmembrane sodium gradient. NCX inactivation occurs in the absence of phosphatidylinositol 4,5-bisphosphate and is facilitated by palmitoylation of a single cysteine at position 739 within the large intracellular loop of NCX. The aim of this investigation was to identify the structural determinants of NCX1 palmitoylation. Full-length NCX1 (FL-NCX1) and a YFP fusion protein of the NCX1 large intracellular loop (YFP-NCX1) were expressed in HEK cells. Single amino acid changes around Cys-739 in FL-NCX1 and deletions on the N-terminal side of Cys-739 in YFP-NCX1 did not affect NCX1 palmitoylation, with the exception of the rare human polymorphism S738F, which enhanced FL-NCX1 palmitoylation, and D741A, which modestly reduced it. In contrast, deletion of a 21-amino acid segment enriched in aromatic amino acids on the C-terminal side of Cys-739 abolished YFP-NCX1 palmitoylation. We hypothesized that this segment forms an amphipathic α-helix whose properties facilitate Cys-739 palmitoylation. Introduction of negatively charged amino acids to the hydrophobic face or of helix-breaking prolines impaired palmitoylation of both YFP-NCX1 and FL-NCX1. Alanine mutations on the hydrophilic face of the helix significantly reduced FL-NCX1 palmitoylation. Of note, when the helix-containing segment was introduced adjacent to cysteines that are not normally palmitoylated, they became palmitoylation sites. In conclusion, we have identified an amphipathic α-helix in the NCX1 large intracellular loop that controls NCX1 palmitoylation. NCX1 palmitoylation is governed by a distal secondary structure element rather than by local primary sequence.

The NCLX-type Na+/Ca2+ Exchanger NCX-9 Is Required for Patterning of Neural Circuits in Caenorhabditis elegans

NCLX is a Na+/Ca2+ exchanger that uses energy stored in the transmembrane sodium gradient to facilitate the exchange of sodium ions for ionic calcium. Mammals have a single NCLX, which has been shown to function primarily at the mitochondrion and is an important regulator of neuronal physiology by contributing to neurotransmission and synaptic plasticity. The role of NCLX in developmental cell patterning (e.g. in neural circuits) is largely unknown. Here we describe a novel role for the Caenorhabditis elegans NCLX-type protein, NCX-9, in neural circuit formation. NCX-9 functions in hypodermal seam cells that secrete the axon guidance cue UNC-129/BMP, and our data revealed that ncx-9-/- mutant animals exhibit development defects in stereotyped left/right axon guidance choices within the GABAergic motor neuron circuit. Our data also implicate NCX-9 in a LON-2/heparan sulfate and UNC-6/netrin-mediated, RAC-dependent signaling pathway to guide left/right patterning within this circuit. Finally, we also provide in vitro physiology data supporting the role for NCX-9 in handling calcium exchange at the mitochondrion. Taken together, our work reveals the specificity by which the handling by NCLX of calcium exchange can map to neural circuit patterning and axon guidance decisions during development.

Origin and evolution of the sodium -pumping NADH: ubiquinone oxidoreductase.

The sodium -pumping NADH: ubiquinone oxidoreductase (Na+-NQR) is the main ion pump and the primary entry site for electrons into the respiratory chain of many different types of pathogenic bacteria. This enzymatic complex creates a transmembrane gradient of sodium that is used by the cell to sustain ionic homeostasis, nutrient transport, ATP synthesis, flagellum rotation and other essential processes. Comparative genomics data demonstrate that the nqr operon, which encodes all Na+-NQR subunits, is found in a large variety of bacterial lineages with different habitats and metabolic strategies. Here we studied the distribution, origin and evolution of this enzymatic complex. The molecular phylogenetic analyses and the organizations of the nqr operon indicate that Na+-NQR evolved within the Chlorobi/Bacteroidetes group, after the duplication and subsequent neofunctionalization of the operon that encodes the homolog RNF complex. Subsequently, the nqr operon dispersed through multiple horizontal transfer events to other bacterial lineages such as Chlamydiae, Planctomyces and α, β, γ and δ -proteobacteria. Considering the biochemical properties of the Na+-NQR complex and its physiological role in different bacteria, we propose a detailed scenario to explain the molecular mechanisms that gave rise to its novel redox- dependent sodium -pumping activity. Our model postulates that the evolution of the Na+-NQR complex involved a functional divergence from its RNF homolog, following the duplication of the rnf operon, the loss of the rnfB gene and the recruitment of the reductase subunit of an aromatic monooxygenase.

Sodium-dependent transporters in health and disease—a special issue

Sodium-dependent transporters utilize the energy stored in the transmembrane sodium gradient to move ions or other solutes against their concentration gradient across the plasma membrane. This sodium gradient is established and maintained by the sodium pump or Na + –K + ATPase that directly hydrolysis ATP. The Na + –K + ATPase pumps three sodium ions out of the cell in concert with pumping two potassium ions into the cell, thus maintaining an intracellular environment that has low sodium and high potassium (the opposite of extracellular fluid). In most cells, 50 % of the total energy expended is used to run these ion pumps. In neurons, which repeatedly gain sodium during action potentials, two thirds of the ATP usage goes toward fueling this single pump. Sodium-dependent transporters belong to the class of cotransporters including symporters and antiporters. Some of which carry both solutes in the same direction (symport), while others transport sodium into the cell and anion or solute out of the cell (antiport). In general, cotransporters are one of three main classes of integral membrane proteins known as transporters that move solutes and ions across the plasma membrane. The cotransporters belong the solute carrier (SLC) group of membrane transport with nearly 400 members organized into 52 families [2, http://slc.bioparadigms.org/]. The SLC gene nomenclature system was originally proposed

Lack of the Sodium-Driven Chloride Bicarbonate Exchanger NCBE Impairs Visual Function in the Mouse Retina

Regulation of ion and pH homeostasis is essential for normal neuronal function. The sodium-driven chloride bicarbonate exchanger NCBE (Slc4a10), a member of the SLC4 family of bicarbonate transporters, uses the transmembrane gradient of sodium to drive cellular net uptake of bicarbonate and to extrude chloride, thereby modulating both intracellular pH (pHi) and chloride concentration ([Cl−]i) in neurons. Here we show that NCBE is strongly expressed in the retina. As GABAA receptors conduct both chloride and bicarbonate, we hypothesized that NCBE may be relevant for GABAergic transmission in the retina. Importantly, we found a differential expression of NCBE in bipolar cells: whereas NCBE was expressed on ON and OFF bipolar cell axon terminals, it only localized to dendrites of OFF bipolar cells. On these compartments, NCBE colocalized with the main neuronal chloride extruder KCC2, which renders GABA hyperpolarizing. NCBE was also expressed in starburst amacrine cells, but was absent from neurons known to depolarize in response to GABA, like horizontal cells. Mice lacking NCBE showed decreased visual acuity and contrast sensitivity in behavioral experiments and smaller b-wave amplitudes and longer latencies in electroretinograms. Ganglion cells from NCBE-deficient mice also showed altered temporal response properties. In summary, our data suggest that NCBE may serve to maintain intracellular chloride and bicarbonate concentration in retinal neurons. Consequently, lack of NCBE in the retina may result in changes in pHi regulation and chloride-dependent inhibition, leading to altered signal transmission and impaired visual function.

Cation-dependent nutrient transport in shrimp digestive tract

Purified epithelial brush border membrane vesicles (BBMV) were produced from the hepatopancreas of the Atlantic White shrimp, Litopeneaus setiferus, using standard methods originally developed for mammalian tissues and previously applied to other crustacean and echinoderm epithelia. These vesicles were used to study the cation dependency of sugar and amino acid transport across luminal membranes of hepatopancreatic epithelial cells. (3)H-D: -glucose uptake by BBMV against transient sugar concentration gradients occurred when either transmembrane sodium or potassium gradients were the only driving forces for sugar accumulation, suggesting the presence of a possible coupled transport system capable of using either cation. (3)H-L: -histidine transport was only stimulated by a transmembrane potassium gradient, while (3)H-L: -leucine uptake was enhanced by either a sodium or potassium gradient. These responses suggest the possible presence of a potassium-dependent transporter that accommodates either amino acid and a sodium-dependent system restricted only to L: -leucine. Uptake of (3)H-L: -leucine was significantly stimulated (P < 0.05) by several metallic cations (e.g., Zn(2+), Cu(2+), Mn(2+), Cd(2+), or Co(2+)) at external pH values of 7.0 or 5.0 (internal pH 7.0), suggesting a potential synergistic role of the cations in the transmembrane transfer of amino acids. (3)H-L: -histidine influxes (15 suptakes) were hyperbolic functions of external [zinc] or [manganese], following Michaelis-Menten kinetics. The apparent affinity constant (e.g., K (m)) for manganese was an order of magnitude smaller (K (m) = 0.22 μM Mn) than that for zinc (K (m) = 1.80 μM Zn), while no significant difference (P > 0.05) occurred between their maximal transport velocities (e.g., J (max)). These results suggest that a number of cation-dependent nutrient transport systems occur on the shrimp brush border membrane and aid in the absorption of these important dietary elements.

Regulation of Intracellular and Mitochondrial Sodium in Health and Disease

The transmembrane sodium gradient is essential for both excitability of the cardiac cell and the regulation of the cytoplasmic concentrations of Ca and protons. In addition, movements of Na across the mitochondrial membrane affect matrix protons and calcium. In the first part of the review, we discuss the most important pathways responsible for sarcolemmal and mitochondrial sodium movements. The bulk of the review considers the changes of intracellular Na concentration ([Na(+)](i)) that occur in disease, specifically, ischemia, reperfusion, and heart failure. We review evidence implicating the increase of intracellular sodium to either increased influx of sodium (via either sodium channels or sodium/hydrogen exchange) or, alternatively, to decreased efflux on the Na/K pump. Although much has been learned about sodium regulation in the heart, there are still many unanswered questions, particularly concerning mitochondrial Na regulation.

Conformational Cycle Of A Bacterial Homolog Of Human Neurotransmitter Sodium Symporters

Neurotransmitter:sodium symporters (NSS) control the magnitude and duration of synaptic signaling through active reuptake of specific neurotransmitters. While it is known that the transmembrane sodium gradient supplies the energy for transport, the coupling mechanism to substrate translocation from its primary binding site (S1) is poorly understood. Conformational rearrangements of extracellular and cytoplasmic regions of the protein are thought to regulate alternating access to S1, and substrate binding at a second site (S2) located near the extracellular region of the transporter, to act as a symport coupler (Shi et al 2008 Mol Cell 30, 667). We used spin labeling and EPR spectroscopy to investigate the conformational dynamics of a highly homologous bacterial member of the NSS family, the leucine transporter LeuT, for which the crystal structure is known (Yamashita et al 2005 Nature 437, 215). Changes in global and local structural constraints derived from the EPR analysis and induced by sodium and leucine binding were then correlated to conformational changes in the LeuT structure in proteoliposomes. Sodium binding was found to increase the distance between the probes as assessed from global rearrangements measured in the extracellular region of LeuT. In contrast, subsequent leucine binding in the presence of sodium was found to decrease the distance between the probes. Consistent with these observations, sodium binding increases spin label mobility and water accessibility at positions within the S2 site. Furthermore, leucine binding increases spin label order and decreases water accessibility. These results suggest a model in which sodium binding to LeuT primes the transporter for substrate binding in S1 by increasing the population of an “outward-facing” conformation, exposing the substrate permeation pathway. With a filled S1, the extracellular pathway constricts, consistent with the occluded state observed in the LeuT crystal structure.

A subset of GAF domains are evolutionarily conserved sodium sensors

Most organisms maintain a transmembrane sodium gradient for cell function. Despite the importance of Na(+) in physiology, no directly Na(+)-responsive signalling molecules are known. The CyaB1 and CyaB2 adenylyl cyclases of the cyanobacterium Anabaena PCC 7120 are inhibited by Na(+). A D360A mutation in the GAF-B domain of CyaB1 ablated cAMP-mediated autoregulation and Na(+) inhibition. Na(+) bound the isolated GAF domains of CyaB2. cAMP blocked Na(+) binding to GAF domains but Na(+) had no effect on cAMP binding. Na(+) altered GAF domain structure indicating a mechanism of inhibition independent of cAMP binding. DeltacyaB1 and DeltacyaB2 mutant strains did not grow below 0.6 mM Na(+) and DeltacyaB1 cells possessed defects in Na(+)/H(+) antiporter function. Replacement of the CyaB1 GAF domains with those of rat phosphodiesterase type 2 revealed that Na(+) inhibition has been conserved since the eukaryotic/bacterial divergence. CyaB1 and CyaB2 are the first identified directly Na(+)-responsive signalling molecules that function in sodium homeostasis and we propose a subset of GAF domains underpin an evolutionarily conserved Na(+) signalling mechanism.

Phospholemman and the Cardiac Sodium Pump

See related article, pages 1376–1383 In excitable tissues, the activity of the plasmalemmal sodium/potassium ATPase (Na/K pump) is vital for the maintenance of normal electrical activity and ion gradients. In cardiac muscle, the transsarcolemmal sodium (Na) gradient established by the Na/K activity is essential not only for generating the rapid upstroke of the action potential but also for driving a number of ion exchange and transport processes that are crucial for normal cellular function, excitation contraction coupling, ion homeostasis and the control of cell volume. These Na-dependent membrane transporters include those responsible for the regulation of other ions (such as the sodium calcium exchanger (NCX), Na/H exchanger and Na-HCO3 cotransporter), as well as those involved in the movement of substrates and amino acids.1 By determining the set point for NCX, the Na/K pump controls the predominant mechanism of transmembrane calcium (Ca) flux, and hence indirectly controls intracellular Ca load and myocardial contractility. Interventions that influence either the activity of the Na/K pump, or indirectly the transmembrane sodium gradient, can therefore profoundly affect normal cardiac function. Phospholemman, a type 1 transmembrane protein, is the predominant quantitative site of phosphorylation by protein kinase A (PKA) and protein kinase C (PKC) in cardiac sarcolemma.2 PKA phosphorylates serine 68 and PKC phosphorylates serines 63 and 68 in phospholemman. For a long time following its cloning in 1991,3 the physiological role of phospholemman was unclear. Indeed, writing in this journal in 1998, researchers noted that “As a major target for hormone-stimulated phosphorylation in the heart, the physiological function of phospholemman is likely to be an important one”,4 however they were unable to suggest what this function may be. Various roles have now been proposed: 1) regulation of cell volume; …

The Transmembrane Sodium Gradient Influences Ambient GABA Concentration by Altering the Equilibrium of GABA Transporters

Tonic inhibition is widely believed to be caused solely by "spillover" of GABA that escapes the synaptic cleft and activates extrasynaptic GABA(A) receptors. However, an exclusively vesicular source is not consistent with the observation that tonic inhibition can still occur after blocking vesicular release. Here, we made patch-clamp recordings from neurons in rat hippocampal cultures and measured the tonic current that was blocked by bicuculline or gabazine. During perforated patch recordings, the tonic GABA current was decreased by the GAT1 antagonist SKF-89976a. Zero calcium solution did not change the amount of tonic current, despite a large reduction in vesicular GABA release. Perturbations that would be expected to alter the transmembrane sodium gradient influenced the tonic current. For example, in zero calcium Ringer, TTX (which can decrease cytosolic [Na(+)]) reduced tonic current, whereas veratridine (which can increase cytosolic [Na(+)]) increased tonic current. Likewise, removal of extracellular sodium led to a large increase in tonic current. The increases in tonic current induced by veratridine and sodium removal were completely blocked by SKF89976a. When these experiments were repeated in hippocampal slices, similar results were obtained except that a GAT1- and GAT3-independent nonvesicular source(s) of GABA was found to contribute to the tonic current. We conclude that multiple sources can contribute to ambient GABA, including spillover and GAT1 reversal. The source of GABA release may be conceptually less important in determining the amount of tonic inhibition than the factors that control the equilibrium of GABA transporters.

From Bacteria to Man: Archaic Proton-Dependent Peptide Transporters at Work

Uptake of nutrients into cells is essential to life and occurs in all organisms at the expense of energy. Whereas in most prokaryotic and simple eukaryotic cells electrochemical transmembrane proton gradients provide the central driving force for nutrient uptake, in higher eukaryotes it is more frequently coupled to sodium movement along the transmembrane sodium gradient, occurs via uniport mechanisms driven by the substrate gradient only, or is linked to the countertransport of a similar organic solute. With the cloning of a large number of mammalian nutrient transport proteins, it became obvious that a few "archaic'' transporters that utilize a transmembrane proton gradient for nutrient transport into cells can still be found in mammals. The present review focuses on the electrogenic peptide transporters as the best studied examples of proton-dependent nutrient transporters in mammals and summarizes the most recent findings on their physiological importance. Taking peptide transport as a general phenomenon found in nature, we also include peptide transport mechanisms in bacteria, yeast, invertebrates, and lower vertebrates, which are not that often addressed in physiology journals.

Clinical review 132: The sodium iodide symporter and its potential role in cancer therapy.

Active transport of iodide into the thyroid gland is a crucial and rate-limiting step in the biosynthesis of thyroid hormones that play an important role in the metabolism, growth, and maturation of a variety of organ systems, in particular the nervous system (1). Although it has been known for decades that iodide transport into the thyroid gland is mediated by a specific sodium-dependent iodide transporter located at the basolateral membrane of thyroid follicular cells, the sodium iodide symporter (NIS) gene was cloned just 4 yr ago (2, 3). After cloning of the rat sodium iodide symporter (rNIS) from a Fisher rat thyroid line (FRTL-5)derived complementary DNA (cDNA) library (2), the human sodium iodide symporter (hNIS) was cloned from a human thyroid cDNA library in 1996 (3). The hNIS gene is localized on chromosome 19p12–13.2 and encodes a glycoprotein of 643 amino acids (aa) with a molecular mass of approximately 70–90 kDa. The coding region of hNIS contains 15 exons interrupted by 14 introns and codes for a 3.9-kb messenger ribonucleic acid (mRNA) transcript (4). As a member of the sodium-dependent transporter family, NIS is an intrinsic membrane protein with 13 putative transmembrane domains, an extracellular amino-terminus, and an intracellular carboxyl-terminus. The NIS protein has three potential Nlinked glycosylation sites; 1 is located in the fourth extracellular (seventh extramembranous domain), and 2 are located in the last extracellular (13th extramembranous domain) loop (5) (Fig. 1). NIS cotransports two sodium ions along with one iodide ion, with the transmembrane sodium gradient serving as the driving force for iodide uptake. The sodium gradient providing the energy for this transfer is generated by the ouabain-sensitive Na/K-adenosine triphosphatase (Na/ K-ATPase). NIS-mediated iodide transport is, therefore, inhibited by the Na/K-ATPase inhibitor ouabain as well as by the competitive inhibitors thiocyanate and perchlorate (1) (Fig. 2). After active transport across the basolateral membrane of thyroid follicular cells, iodide is translocated across the apical membrane by pendrin, the Pendred syndrome gene product, which is a chloride/iodide transporter (6–10) (Fig. 2). Other apical anion transporters may also be involved. At the cell/colloid interface iodide is organified in a complex reaction involving oxidation catalyzed by thyroid peroxidase (TPO) and incorporation into tyrosyl residues along the thyroglobulin (Tg) backbone. The thyroid hormones T3 and T4 are synthesized by coupling of two iodotyrosine residues and are stored in the colloid (Fig. 2). The iodide organification step can be inhibited by propylthiouracil and methimazole, which are TPO enzyme inhibitors. All of these steps are stimulated through pituitary-derived TSH, which interacts with the TSH receptor at the basolateral membrane of thyroidal cells (1). It has been known for many years that TSH stimulates iodide transport into the thyroid gland via the adenylate cyclase-cAMP pathway (1). After NIS was cloned, several studies in FRTL-5 cells and cultured human thyroid cells showed that treatment with TSH stimulates iodide transport activity as well as NIS gene and protein expression (11, 12). Forskolin and dibutyryl cAMP are able to mimic this stimulatory effect on both iodide transport activity as well as NIS gene and protein expression, suggesting that TSH regulates NIS expression through the cAMP signal transduction pathway (11) (Fig. 2). In addition to its key role in thyroid physiology, NISmediated iodide accumulation in the thyroid gland is a crucial prerequisite for diagnostic scintigraphic imaging as well as for the highly efficient radioiodine therapy of benign and malignant thyroid diseases. The purpose of this review is to summarize and discuss the current knowledge of NIS and its diagnostic and therapeutic implications in thyroidal and nonthyroidal cancer.