The G215R Mutation in the Cl−-H -Antiporter ClC-7 Found in ADO II Osteopetrosis Does Not Abolish Function but Causes a Severe Trafficking DefectReportar como inadecuado




The G215R Mutation in the Cl−-H -Antiporter ClC-7 Found in ADO II Osteopetrosis Does Not Abolish Function but Causes a Severe Trafficking Defect - Descarga este documento en PDF. Documentación en PDF para descargar gratis. Disponible también para leer online.

Background

ClC-7 is a ubiquitous transporter which is broadly expressed in mammalian tissues. It is implied in the pathogenesis of lysosomal storage disease and osteopetrosis. Because of its endosomal-lysosomal localization it is still poorly characterized.

Methodology-Principal Findings

An electrophysiological characterization of rat ClC-7 using solid-supported membrane-based electrophysiology is presented. The measured currents show the characteristics of ClC-7 and confirm its function as a Cl−-H+-antiporter. We have used rat ClC-7 in CHO cells as a model system to investigate the functionality and cellular localization of the wt transporter and its variant G213R ClC-7 which is the analogue of human G215R ClC-7 responsible for autosomal dominant osteopetrosis type II. Our study shows that rat G213R ClC-7 is functional but has a localization defect in CHO cells which prevents it from being correctly targeted to the lysosomal membrane. The electrophysiological assay is tested as a tool for drug discovery. The assay is validated with a number of drug candidates. It is shown that ClC-7 is inhibited by DIDS, NPPB and NS5818 at micromolar concentrations.

Conclusions-Significance

It is suggested that the scenario found in the CHO model system also applies to the human transporter and that mislocalization rather than impaired functionality of G215R ClC-7 is the primary cause of the related autosomal dominant osteopetrosis type II. Furthermore, the robust solid-supported membrane-based electrophysiological assay is proposed for rapid screening for potential ClC-7 inhibitors which are discussed for treatment of osteoporosis.



Autor: Patrick Schulz, Johannes Werner, Tobias Stauber, Kim Henriksen, Klaus Fendler

Fuente: http://plos.srce.hr/



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