Topology of the human skeletal muscle chloride channel hClC-1 probed with hydrophilic epitope insertion

Kuchenbecker, Maya; Schu, Bernhard; Kürz, Lothar L.; Rüdel, Reinhardt

doi:10.1007/s004240100688

Cited by 3 publications

(2 citation statements)

References 41 publications

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“…The transmembrane topology of CLC proteins had been explored using a variety of techniques, correctly identifying the orientation of several membrane helices and intra/extracellular localization of loops (228,303,415,437,489,544,750). However, in retrospect, given the complex architecture revealed by CLC crystal structures, a full determination of the membrane topology was extremely challenging.…”

Section: A the Double-barreled Torpedo Clc-0 Channelmentioning

confidence: 99%

CLC Chloride Channels and Transporters: Structure, Function, Physiology, and Disease

Jentsch

Pusch

2018

Physiological Reviews

368

421

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CLC anion transporters are found in all phyla and form a gene family of eight members in mammals. Two CLC proteins, each of which completely contains an ion translocation parthway, assemble to homo- or heteromeric dimers that sometimes require accessory β-subunits for function. CLC proteins come in two flavors: anion channels and anion/proton exchangers. Structures of these two CLC protein classes are surprisingly similar. Extensive structure-function analysis identified residues involved in ion permeation, anion-proton coupling and gating and led to attractive biophysical models. In mammals, ClC-1, -2, -Ka/-Kb are plasma membrane Cl channels, whereas ClC-3 through ClC-7 are 2Cl/H-exchangers in endolysosomal membranes. Biological roles of CLCs were mostly studied in mammals, but also in plants and model organisms like yeast and Caenorhabditis elegans. CLC Cl channels have roles in the control of electrical excitability, extra- and intracellular ion homeostasis, and transepithelial transport, whereas anion/proton exchangers influence vesicular ion composition and impinge on endocytosis and lysosomal function. The surprisingly diverse roles of CLCs are highlighted by human and mouse disorders elicited by mutations in their genes. These pathologies include neurodegeneration, leukodystrophy, mental retardation, deafness, blindness, myotonia, hyperaldosteronism, renal salt loss, proteinuria, kidney stones, male infertility, and osteopetrosis. In this review, emphasis is laid on biophysical structure-function analysis and on the cell biological and organismal roles of mammalian CLCs and their role in disease.

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Section: A the Double-barreled Torpedo Clc-0 Channelmentioning

confidence: 99%

CLC Chloride Channels and Transporters: Structure, Function, Physiology, and Disease

Jentsch

Pusch

2018

Physiological Reviews

368

421

View full text Add to dashboard Cite

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“…1D) in tsA201 cells, applied anti-FLAG antibodies to nonpermeabilized cells, and individually measured GFP and antibody fluorescence levels. Because the engineered FLAG epitope is located extracellularly (13,14), a plot of the antibody fluorescence levels versus the GFP fluorescence ( Fig. 1 J and K) provides information about the fraction of channels inserted into the plasma membrane.…”

Section: Barttin Promotes Surface Membrane Insertion Of Rclc-k1 and Hmentioning

confidence: 99%

Barttin modulates trafficking and function of ClC-K channels

Scholl

Hebeisen²,

Janssen³

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

104

161

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Barttin is an accessory subunit of a subgroup of ClC-type chloride channels expressed in renal and inner ear epithelia. In this study, we examined the effects of barttin on two ClC-K channel isoforms, rat ClC-K1 and human ClC-Kb, using heterologous expression, patch clamping, confocal imaging, and flow cytometry. In the absence of barttin, only a small percentage of rClC-K1 and hClC-Kb channels are inserted into the plasma membrane. Coexpression of barttin enhances surface membrane insertion and furthermore modifies permeation and gating of ClC-K channels. hClC-Kb channels are nonfunctional without barttin and require the coexpressed accessory subunit to become anion conducting. In contrast, rClC-K1 channels are active without barttin, but at the cost of reduced unitary conductance as well as altered voltage dependence of activation. We mapped the separate functions of barttin to structural domains by a deletion analysis. Whereas the transmembrane core is necessary and sufficient to promote ClC-K channel exit from the endoplasmic reticulum, a short cytoplasmic segment following the second transmembrane helix modifies the unitary conductance. The entire cytoplasmic carboxyl terminus affects the open probability of ClC-K channels. The multiple functions of barttin might be necessary for a tight adjustment of epithelial Cl ؊ conductances to ensure a precise regulation of body salt content and endocochlear potential.chloride channels ͉ gating ͉ kidney ͉ ClC family ͉ accessory subunit

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Plasma Membrane Expression of T-type Calcium Channel α1 Subunits Is Modulated by High Voltage-activated Auxiliary Subunits

Dübel

Altier

Chaumont

et al. 2004

Journal of Biological Chemistry

113

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It has been suggested that the auxiliary subunits of high voltage-activated (HVA) calcium channels modulate T-type, low voltage-activated (LVA) calcium channels. Such a regulation has yet to be documented, especially because there has been no biochemical characterization of T-channels. To monitor total protein levels and plasma membrane expression of T-channels in living cells, external epitopes (hemagglutinin, FLAG) were introduced into human recombinant Ca V 3 channels that were also N-terminally fused to green fluorescent protein. Utilizing Western blot techniques, fluorescence flow cytometry, immunofluorescence, luminometry, and electrophysiology, we describe here that ␤ 1b and ␣ 2 -␦ 1 subunits enhance the level of Ca V 3 proteins as well as their plasma membrane expression in various expression systems. We also report that, in both Xenopus oocytes and mammalian cells, the ␣ 2 -␦ 1 and ␤ 1b subunits increase by at least 2-fold the current density of Ca V 3 channels with no change in the electrophysiological properties. Altogether, these data indicate that HVA auxiliary subunits modulate Ca V 3 channel surface expression, suggesting that the membrane targeting of HVA and LVA ␣ 1 subunits is regulated dynamically through the expression of a common set of regulatory subunits.

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Topology of the human skeletal muscle chloride channel hClC-1 probed with hydrophilic epitope insertion

Cited by 3 publications

References 41 publications

CLC Chloride Channels and Transporters: Structure, Function, Physiology, and Disease

CLC Chloride Channels and Transporters: Structure, Function, Physiology, and Disease

Barttin modulates trafficking and function of ClC-K channels

Plasma Membrane Expression of T-type Calcium Channel α1 Subunits Is Modulated by High Voltage-activated Auxiliary Subunits

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