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Molecular identification of a volume-regulated chloride channel

Abstract

A volume-regulated chloride current ( I Cl.vol) is ubiquitously present in mammalian cells, and is required for the regulation of electrical activity, cell volume, intracellular pH, immunological responses, cell proliferation and differentiation. However, the molecule responsible for I Cl.vol has yet to be determined1,2,3. Although three putative chloride channel proteins expressed from cloned genes (P-glycoprotein4, p I Cln (ref. 5) and ClC-2 (ref. 6)) have been proposed to be the molecular equivalent of I Cl.vol, neither P-glycoprotein nor p I Cln is thought to be a chloride channel or part thereof7,8, and the properties of expressed ClC-2 channels differ from native I Cl.vol (refs. 3, 6). Here we report that functional expression in NIH/3T3 cells of a cardiac clone of another member of the ClC family, ClC-3, results in a large basally active chloride conductance, which is strongly modulated by cell volume and exhibits many properties identical to those of I Cl.vol in native cells1,2,3,9,10,11,12,13. A mutation of asparagine to lysine at position 579 at the end of the transmembrane domains of ClC-3 abolishes the outward rectification and changes the anion selectivity from I > Cl to Cl > I but leaves swelling activation intact. Because ClC-3 is a channel protein belonging to a large gene family of chloride channels3,14, these results indicate that ClC-3 encodes I Cl.vol in many native mammalian cells.

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Figure 1: Expression of ClC-3 in guinea-pig heart.
Figure 2: Pharmacology and ion selectivity of I ClC-3 in NIH/3T3 cells A, B, PKC under isotonic (A) and hypotonic (B) conditions.
Figure 3: Single-channel properties of I ClC-3 in cell-attached patches from gpClC-3-transfected NIH/3T3 cells.
Figure 4: Properties of wild-type and mutant N579K gpClC-3 transiently expressedin NIH/3T3 cells.

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Acknowledgements

We thank N. Horowitz, F. Schmalz, L. Ye and T. Truong for technical support, and J.Yamazaki for assistance in experiments with pZeoSV-CFTR. This study was supported by the Medical Research Council of Canada (D.D.), the American Heart Association, Nevada Affiliate (C.W.), and the NIH (J.R.H. and B.H.).

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Correspondence to Joseph R. Hume or Burton Horowitz.

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Duan, D., Winter, C., Cowley, S. et al. Molecular identification of a volume-regulated chloride channel. Nature 390, 417–421 (1997). https://doi.org/10.1038/37151

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