Modeling study of human renal chloride channel (hCLC-5) mutations suggests a structural-functional relationship

Wu, Fiona; Roché, Philippe; Christie, Paul T.; Loh, Nellie Y.; Reed, Anita; Esnouf, Robert; Thakker, Rajesh V.

doi:10.1046/j.1523-1755.2003.00859.x

Cited by 52 publications

(59 citation statements)

References 31 publications

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“…Each subunit has its own pore and 17 α-helices inserted into the plasma membrane, with an anti-parallel orientation that brings together residues responsible for anion selectivity [21]. Based on these data, a model for human ClC-5 has been established, allowing analysis of the role of 15 non-truncating mutations (14 missense and one in-frame deletion) of CLCN5 [22]. This analysis revealed that none of these mutations involves the selectivity filter, but that most of them (12/15) are clustered at the interface of the two subunits of the homodimeric channel [22].…”

Section: Recent Developments and Perspectivesmentioning

confidence: 99%

“…Based on these data, a model for human ClC-5 has been established, allowing analysis of the role of 15 non-truncating mutations (14 missense and one in-frame deletion) of CLCN5 [22]. This analysis revealed that none of these mutations involves the selectivity filter, but that most of them (12/15) are clustered at the interface of the two subunits of the homodimeric channel [22]. The functional evaluation of other naturally occurring mutants is necessary in order to define the role of conserved domains for ClC-5 function and/or trafficking: a case in point is a C-terminus internalization motif, resembling the PY motif that is essential for the internalization and degradation of the epithelial sodium channel, ENaC [23].…”

Section: Recent Developments and Perspectivesmentioning

confidence: 99%

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Chloride Channels and Endocytosis: New Insights from Dent’s Disease and ClC-5 Knockout Mice

et al. 2005

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Dent’s disease is a hereditary renal tubular disorder characterized by low-molecular weight (LMW) proteinuria, hypercalciuria and nephrolithiasis. The disease is due to mutations of ClC-5, a member of the family of voltage-gated CLC chloride channels. ClC-5 is expressed in part in cells lining the proximal tubule (PT) of the kidney, where it colocalizes with albumin-containing endocytic vesicles belonging to the receptor-mediated endocytic pathway that ensures efficient reabsorption of ultrafiltrated LMW proteins. Since progression along the endocytic apparatus requires endosomal acidification, it has been suggested that dysfunction of ClC-5 in endosomes may lead to inefficient reabsorption of LMW proteins and dysfunction of PT cells. Analysis of a ClC-5 knockout (KO) mouse model, displaying all the characteristic renal tubular defects of Dent’s disease, showed evidence of a severe LMW proteinuria. Cytochemical studies with the endocytic tracer, peroxidase, showed poor transfer into early endocytic vesicles, suggesting that impairment of receptor-mediated endocytosis in PT cells is the basis for the defective uptake of LMW proteins in patients with Dent’s disease. Endocytosis and processing of LMW proteins involve the multiligand tandem receptors, megalin and cubilin, that are abundantly expressed at the brush border of PT cells. Characterization of the endocytic defect in ClC-5 KO mice revealed that ligands of both megalin and cubilin were affected. The total kidney content of megalin and especially cubilin at the protein level was decreased but, more importantly, using analytical subcellular fractionation and quantitative immunogold labelling we demonstrated a selective disappearance of megalin and cubilin at the brush border of PT cells. These observations allowed us to conclude that defective protein endocytosis linked to ClC-5 inactivation is due at least in part to a major and selective loss of megalin and cubilin at the brush border, reflecting a trafficking defect in renal PT cells. These results improve our understanding of Dent’s disease, taken as a paradigm for renal Fanconi syndrome and nephrolithiasis, and demonstrate multiple roles for ClC-5 in the kidney. These studies also provided insights into important functions such as apical endocytosis, handling of proteins by renal tubular cells, calcium metabolism, and urinary acidification.

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Section: Recent Developments and Perspectivesmentioning

confidence: 99%

Section: Recent Developments and Perspectivesmentioning

confidence: 99%

Chloride Channels and Endocytosis: New Insights from Dent’s Disease and ClC-5 Knockout Mice

et al. 2005

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“…DNA samples from 87 unrelated healthy Japanese individuals (68 females, 19 males) with 155 alleles of the CLCN5 gene did not reveal this nucleotide change, indicating that the mutation is not due to a common single nucleotide polymorphism. In the CLC-5 protein, this amino acid substitution is located in helix J, one of 18 α-helices [9]. Genomic analysis of the patient’s mother revealed that she was heterozygous for the mutation.…”

Section: Resultsmentioning

confidence: 99%

“…The human CLCN5 gene, which is located on chromosome Xp11.22, has a coding region of 2,238 bp that consists of 11 exons and encodes the 746 amino acid protein, CLC-5 [7]. Prokaryotic CLC-5 was found to form a homodimeric structure by X-ray analysis [8] and a similar structural model was suggested for the human counterpart in which each subunit consists of 18 α-helices [9]. Heterologous expression of wild-type (WT) CLC-5 in Xenopus oocytes revealed that CLC-5 conducts outwardly rectifying chloride currents and similar expression of disease-associated CLC-5 mutants demonstrated markedly reduced or absent currents [4].…”

Section: Introductionmentioning

confidence: 99%

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Functional Characterization of a Novel Missense <i>CLCN5</i> Mutation Causing Alterations in Proximal Tubular Endocytic Machinery in Dent’s Disease

et al. 2007

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Background/Aims: Mutations of the endosomal chloride/proton exchanger gene, CLCN5, cause Dent’s disease, an X-linked recessive proximal tubular disorder. The renal endocytic system was found to be affected in clcn5 knockout mice. However, the impaired endocytic machinery of Dent’s disease patients has not been thoroughly investigated. Methods: The CLCN5 gene was sequenced in a Japanese patient with Dent’s disease and his family. The loss-of-function phenotype of the missense CLCN5 mutation was investigated by gene expression in Xenopus oocytes and CHO cells. Immunohistochemical analysis was performed on kidney biopsy specimens for endocytic machinery proteins, megalin, cubilin, and disabled-2 (Dab2) in proximal tubules. Results: Genomic analysis revealed a novel G-to-A transition at the first nucleotide of the 333rd codon of CLCN5, causing a substitution of glycine with arginine. Inefficient expression of the mutant gene in Xenopus oocytes resulted in abolished chloride currents. Impaired N-glycosylation of the mutant protein was evident in the DNA-transfected CHO cells. Proximal tubular expression of megalin, cubilin, and Dab2 was markedly reduced and irregular staining in some portions was observed in the patient compared with controls. Conclusions: A novel G333R CLCN5 mutation caused defective expression of megalin, cubilin, and Dab2 in a patient with Dent’s disease.

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Cell Biology and Physiology of CLC Chloride Channels and Transporters

Stauber

Weinert

Jentsch

2012

Comprehensive Physiology

134

138

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Proteins of the CLC gene family assemble to homo‐ or sometimes heterodimers and either function as Cl – channels or as Cl – /H + ‐exchangers. CLC proteins are present in all phyla. Detailed structural information is available from crystal structures of bacterial and algal CLCs. Mammals express nine CLC genes, four of which encode Cl – channels and five 2Cl – /H + ‐exchangers. Two accessory β‐subunits are known: (1) barttin and (2) Ostm1. ClC‐Ka and ClC‐Kb Cl – channels need barttin, whereas Ostm1 is required for the function of the lysosomal ClC‐7 2Cl – /H + ‐exchanger. ClC‐1, ‐2, ‐Ka and ‐Kb Cl – channels reside in the plasma membrane and function in the control of electrical excitability of muscles or neurons, in extra‐ and intracellular ion homeostasis, and in transepithelial transport. The mainly endosomal/lysosomal Cl – /H + ‐exchangers ClC‐3 to ClC‐7 may facilitate vesicular acidification by shunting currents of proton pumps and increase vesicular Cl – concentration. ClC‐3 is also present on synaptic vesicles, whereas ClC‐4 and ‐5 can reach the plasma membrane to some extent. ClC‐7/Ostm1 is coinserted with the vesicular H + ‐ATPase into the acid‐secreting ruffled border membrane of osteoclasts. Mice or humans lacking ClC‐7 or Ostm1 display osteopetrosis and lysosomal storage disease. Disruption of the endosomal ClC‐5 Cl – /H + ‐exchanger leads to proteinuria and Dent's disease. Mouse models in which ClC‐5 or ClC‐7 is converted to uncoupled Cl – conductors suggest an important role of vesicular Cl – accumulation in these pathologies. The important functions of CLC Cl – channels were also revealed by human diseases and mouse models, with phenotypes including myotonia, renal loss of salt and water, deafness, blindness, leukodystrophy, and male infertility. © 2012 American Physiological Society. Compr Physiol 2:1701‐1744, 2012.

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Modeling study of human renal chloride channel (hCLC-5) mutations suggests a structural-functional relationship

Abstract: These results demonstrate a crucial role for the interaction between the two subunits at the interface of the homodimeric hCLC-5.

Cited by 52 publications

References 31 publications

Chloride Channels and Endocytosis: New Insights from Dent’s Disease and ClC-5 Knockout Mice

Chloride Channels and Endocytosis: New Insights from Dent’s Disease and ClC-5 Knockout Mice

Functional Characterization of a Novel Missense <i>CLCN5</i> Mutation Causing Alterations in Proximal Tubular Endocytic Machinery in Dent’s Disease

Cell Biology and Physiology of CLC Chloride Channels and Transporters

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