Publications

• SLC26A7 is a Cl- channel regulated by intracellular pH.

  Kim K.H., Shcheynikov N., Wang Y., Muallem S.

  Members of the SLC26 transporter family play an essential role in several epithelial functions, as revealed by diseases associated with mutations in members of the family. Several members were shown to function as Cl(-) and HCO(3)(-) transporters that likely play an important role in epithelial Cl ... >> More

  Members of the SLC26 transporter family play an essential role in several epithelial functions, as revealed by diseases associated with mutations in members of the family. Several members were shown to function as Cl(-) and HCO(3)(-) transporters that likely play an important role in epithelial Cl(-) absorption and HCO(3)(-) secretion. However, the mechanism of most transporters is not well understood. SLC26A7 is a member of the SLC26 transporter family reported to be expressed in the basolateral membrane of the cortical collecting duct and parietal cells and functions as a coupled Cl(-)/HCO(3)(-) exchanger. In the present work we examined the transport properties of SLC26A7 to determine its transport characteristics and electrogenicity. We found that when expressed in Xenopus oocytes or HEK293 cells SLC26A7 functions as a pH(i)-regulated Cl(-) channel with minimal OH(-)/HCO(3)(-) permeability. Expression of SLC26A7 in oocytes or HEK293 cells generated a Cl(-) current with linear I/V and an instantaneous current that was voltage- and time-independent. Based on measurement of reversal potential the selectivity of SLC26A7 is NO(3)(-)>>Cl(-)=Br(-)=I(-)>SO(4)(2-)=Glu(-), although I(-) partially inhibited the current. Incubating the cells with HCO(3)(-) or butyrate acidified the cytosol and increased the selectivity of SLC26A7 for Cl(-). Measurement of membrane potential and pH(i) showed minimal OH(-) and HCO(3)(-) transport by SLC26A7 when the cells were incubated in Cl(-)-containing or Cl(-)-free media. The activity of SLC26A7 was inhibited by all inhibitors of anion transporters tested, 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid, diphenylamine-2-carboxylic acid, and glybenclamide. These findings reveal that SLC26A7 functions as a unique Cl(-) channel that is regulated by intracellular H(+). << Less

  J. Biol. Chem. 280:6463-6470(2005) [PubMed] [EuropePMC]

  This publication is cited by 5 other entries.

• Identification of LRRC8 heteromers as an essential component of the volume-regulated anion channel VRAC.

  Voss F.K., Ullrich F., Muench J., Lazarow K., Lutter D., Mah N., Andrade-Navarro M.A., von Kries J.P., Stauber T., Jentsch T.J.

  Regulation of cell volume is critical for many cellular and organismal functions, yet the molecular identity of a key player, the volume-regulated anion channel VRAC, has remained unknown. A genome-wide small interfering RNA screen in mammalian cells identified LRRC8A as a VRAC component. LRRC8A f ... >> More

  Regulation of cell volume is critical for many cellular and organismal functions, yet the molecular identity of a key player, the volume-regulated anion channel VRAC, has remained unknown. A genome-wide small interfering RNA screen in mammalian cells identified LRRC8A as a VRAC component. LRRC8A formed heteromers with other LRRC8 multispan membrane proteins. Genomic disruption of LRRC8A ablated VRAC currents. Cells with disruption of all five LRRC8 genes required LRRC8A cotransfection with other LRRC8 isoforms to reconstitute VRAC currents. The isoform combination determined VRAC inactivation kinetics. Taurine flux and regulatory volume decrease also depended on LRRC8 proteins. Our work shows that VRAC defines a class of anion channels, suggests that VRAC is identical to the volume-sensitive organic osmolyte/anion channel VSOAC, and explains the heterogeneity of native VRAC currents. << Less

  Science 344:634-638(2014) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.

• Interactions of bromide, iodide, and fluoride with the pathways of chloride transport and diffusion in human neutrophils.

  Simchowitz L.

  Isolated human neutrophils possess three distinct pathways by which Cl-crosses the plasma membrane of steady state cells: anion exchange, active transport, and electrodiffusion. The purpose of the present work was to investigate the selectivity of each of these separate processes with respect to o ... >> More

  Isolated human neutrophils possess three distinct pathways by which Cl-crosses the plasma membrane of steady state cells: anion exchange, active transport, and electrodiffusion. The purpose of the present work was to investigate the selectivity of each of these separate processes with respect to other external halide ions. (a) The bulk of total anion movements represents transport through an electrically silent anion-exchange mechanism that is insensitive to disulfonic stilbenes, but which can be competitively inhibited by alpha-cyano-4-hydroxycinnamate (CHC; Ki approximately 0.3 mM). The affinity of the external translocation site of the carrier for each of the different anions was determined (i) from substrate competition between Cl- and either Br-, F-, or I-, (ii) from trans stimulation of 36Cl-efflux as a function of the external concentrations of these anions, (iii) from changes in the apparent Ki for CHC depending on the nature of the replacement anion in the bathing medium, and (iv) from activation of 82Br- and 125I-influxes by their respective ions. Each was bound and transported at roughly similar rates (Vmax values all 1.0-1.4 meq/liter cell water.min); the order of decreasing affinities is Cl- greater than Br- greater than F- greater than I-(true Km values of 5, 9, 23, and 44 mM, respectively). These anions undergo 1:1 countertransport for internal Cl-. (b) There is a minor component of total Cl-influx that constitutes an active inward transport system for the intracellular accumulation of Cl-[( Cl-]i approximately 80 meq/liter cell water), fourfold higher than expected for passive distribution. This uptake is sensitive to intracellular ATP depletion by 2-deoxy-D-glucose and can be inhibited by furosemide, ethacrynic acid, and CHC, which also blocks anion exchange. This active Cl-uptake process binds and transports other members of the halide series in the sequence Cl- greater than Br- greater than I- greater than F-(Km values of 5, 8, 15, and 41 mM, respectively). (c) Electrodiffusive fluxes are small. CHC-resistant 82Br- and 125I-influxes behave as passive leak fluxes through low-conductance ion channels: they are nonsaturable and strongly voltage dependent. These anions permeate the putative Cl- channel in the sequence I- greater than Br- greater than Cl-with relative permeability ratios of 2.2:1.4:1, respectively, where PCl approximately 5 X 10(-9) cm/s. << Less

  J Gen Physiol 91:835-860(1988) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.

• Neutral amino acid transporter ASCT2 displays substrate-induced Na+ exchange and a substrate-gated anion conductance.

  Broeer A., Wagner C., Lang F., Broeer S.

  The neutral amino acid transporter ASCT2 mediates electroneutral obligatory antiport but at the same time requires Na(+) for its function. To elucidate the mechanism, ASCT2 was expressed in Xenopus laevis oocytes and transport was analysed by flux studies and two-electrode voltage clamp recordings ... >> More

  The neutral amino acid transporter ASCT2 mediates electroneutral obligatory antiport but at the same time requires Na(+) for its function. To elucidate the mechanism, ASCT2 was expressed in Xenopus laevis oocytes and transport was analysed by flux studies and two-electrode voltage clamp recordings. Flux studies with (22)NaCl indicated that the uptake of one molecule of glutamine or alanine is accompanied by the uptake of four to seven Na(+) ions. Similarly to the transport of amino acids, the Na(+) uptake was mediated by an obligatory Na(+) exchange mechanism that depended on the presence of amino acids but was not stoichiometrically coupled to the amino acid transport. Other cations could not replace Na(+) in this transport mechanism. When NaCl was replaced by NaSCN in the transport buffer, the superfusion of oocytes with amino acid substrates resulted in large inward currents, indicating the presence of a substrate-gated anion channel in the ASCT2 transporter. The K(m) for glutamine derived from these experiments is in good agreement with the K(m) derived from flux studies; it varied between 40 and 90 microM at holding potentials of -60 and -20 mV respectively. The permeability of the substrate-gated anion conductance decreased in the order SCN(-)>>NO(3)(-)>I(-)>Cl(-) and also required the presence of Na(+). << Less

  Biochem. J. 346:705-710(2000) [PubMed] [EuropePMC]

  This publication is cited by 2 other entries.

• SWELL1, a plasma membrane protein, is an essential component of volume-regulated anion channel.

  Qiu Z., Dubin A.E., Mathur J., Tu B., Reddy K., Miraglia L.J., Reinhardt J., Orth A.P., Patapoutian A.

  Maintenance of a constant cell volume in response to extracellular or intracellular osmotic changes is critical for cellular homeostasis. Activation of a ubiquitous volume-regulated anion channel (VRAC) plays a key role in this process; however, its molecular identity in vertebrates remains unknow ... >> More

  Maintenance of a constant cell volume in response to extracellular or intracellular osmotic changes is critical for cellular homeostasis. Activation of a ubiquitous volume-regulated anion channel (VRAC) plays a key role in this process; however, its molecular identity in vertebrates remains unknown. Here, we used a cell-based fluorescence assay and performed a genome-wide RNAi screen to find components of VRAC. We identified SWELL1 (LRRC8A), a member of a four-transmembrane protein family with unknown function, as essential for hypotonicity-induced iodide influx. SWELL1 is localized to the plasma membrane, and its knockdown dramatically reduces endogenous VRAC currents and regulatory cell volume decrease in various cell types. Furthermore, point mutations in SWELL1 cause a significant change in VRAC anion selectivity, demonstrating that SWELL1 is an essential VRAC component. These findings enable further molecular characterization of the VRAC channel complex and genetic studies for understanding the function of VRAC in normal physiology and disease. << Less

  Cell 157:447-458(2014) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.

• Anionic leak currents through the Na+/monocarboxylate cotransporter SMCT1.

  Coady M.J., Wallendorff B., Bourgeois F., Lapointe J.Y.

  SMCT1 is a Na-coupled cotransporter of short chain monocarboxylates, which is expressed in the apical membrane of diverse epithelia such as colon, renal cortex, and thyroid. We previously reported that SMCT1 cotransport was reduced by extracellular Cl(-) replacement with cyclamate(-) and that the ... >> More

  SMCT1 is a Na-coupled cotransporter of short chain monocarboxylates, which is expressed in the apical membrane of diverse epithelia such as colon, renal cortex, and thyroid. We previously reported that SMCT1 cotransport was reduced by extracellular Cl(-) replacement with cyclamate(-) and that the protein exhibited an ostensible anionic leak current. In this paper, we have revisited the interaction between small monovalent anions and SMCT cotransport and leak currents. We found that the apparent Cl(-) dependence of cotransport was due to inhibition of this protein by the replacement anion cyclamate, whereas several other replacement anions function as substrates for SMCT1; a suitable replacement anion (MES(-)) was identified. The observed outward leak currents represented anionic influx and favored larger anions (NO(3)(-)>I(-)>Br(-)>Cl(-)); currents in excess of 1 muA (at +50 mV) could be observed and exhibited a quasilinear relationship with anion concentrations up to 100 mM. Application of 25 mM bicarbonate did not produce measurable leak currents. The leak current displayed outward rectification, which disappeared when external Na(+) was replaced by N-methyl-d-glucamine(+). More precisely, external Na(+) blocked the leak current in both directions, but its K(i) value rose rapidly when membrane potential became positive. Thus SMCT1 possesses a anionic leak current that becomes significant whenever external Na(+) concentration is reduced. The presence of this leak current may represent a second function for SMCT1 in addition to cotransporting short chain fatty acids, and future experiments will determine whether this function serves a physiological role in tissues where SMCT1 is expressed. << Less

  Am. J. Physiol. 298:C124-C131(2010) [PubMed] [EuropePMC]

  This publication is cited by 2 other entries.

• CLN7 is an organellar chloride channel regulating lysosomal function.

  Wang Y., Zeng W., Lin B., Yao Y., Li C., Hu W., Wu H., Huang J., Zhang M., Xue T., Ren D., Qu L., Cang C.

  Neuronal ceroid lipofuscinoses (NCLs) are a group of autosomal recessive lysosomal storage diseases. One variant form of late-infantile NCL (vLINCL) is caused by mutations of a lysosomal membrane protein CLN7, the function of which has remained unknown. Here, we identified CLN7 as a novel endolyso ... >> More

  Neuronal ceroid lipofuscinoses (NCLs) are a group of autosomal recessive lysosomal storage diseases. One variant form of late-infantile NCL (vLINCL) is caused by mutations of a lysosomal membrane protein CLN7, the function of which has remained unknown. Here, we identified CLN7 as a novel endolysosomal chloride channel. Overexpression of CLN7 increases endolysosomal chloride currents and enlarges endolysosomes through a Ca²⁺/calmodulin-dependent way. Human CLN7 and its yeast homolog exhibit characteristics of chloride channels and are sensitive to chloride channel blockers. Moreover, CLN7 regulates lysosomal chloride conductance, luminal pH, and lysosomal membrane potential and promotes the release of lysosomal Ca²⁺ through transient receptor potential mucolipin 1 (TRPML1). Knocking out CLN7 causes pathological features that are similar to those of patients with vLINCL, including retinal degeneration and autofluorescent lipofuscin. The pathogenic mutations in CLN7 lead to a decrease in chloride permeability, suggesting that reconstitution of lysosomal Cl⁻ homeostasis may be an effective strategy for the treatment of vLINCL. << Less

  Sci. Adv. 7:eabj9608-eabj9608(2021) [PubMed] [EuropePMC]

  This publication is cited by 2 other entries.