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Expression and regulation of ClC-5 chloride channels: effects of antisense and oxidants

T X Weng1, L Mo, H L Hellmich

  • 1Department of Physiology and Biophysics, University of Texas Medical Branch, Galveston, TX 77555, USA.

American Journal of Physiology. Cell Physiology
|May 15, 2001
PubMed

Insights

Genetic mutations in ClC-5 channels cause Dent's disease. This study found ClC-5 channel properties are consistent across cell types and may be regulated by protein kinase A and reactive oxygen species.

Area of Science:

  • Molecular Biology
  • Cell Biology
  • Physiology

Background:

  • Genetic mutations in the chloride channel ClC-5 are linked to Dent's disease in humans.
  • An amphibian ortholog, Xenopus ClC-5 (xClC-5), was previously cloned from the A6 cell line.

Purpose of the Study:

  • To compare the properties and regulation of ClC-5 currents expressed in mammalian (COS-7) cells and Xenopus oocytes.
  • To investigate the functional conservation of ClC-5 across different expression systems.

Main Methods:

  • Whole-cell patch-clamp electrophysiology was used to record ClC-5 currents in COS-7 cells and Xenopus oocytes.
  • Transfection of COS-7 cells with xClC-5 cDNA and injection of antisense xClC-5 cRNA into oocytes were performed.
  • The effects of pH, anion selectivity, and various signaling molecules (cAMP activators, protein kinase C inhibitors, H-89, hydrogen peroxide) on ClC-5 currents were assessed.

Main Results:

  • ClC-5 currents expressed in COS-7 cells exhibited strong outward rectification, chloride over iodide anion sensitivity, and inhibition at low pH, mirroring observations in oocytes.
  • Antisense xClC-5 cRNA injection in oocytes did not affect endogenous currents or human ClC-5 expression.
  • While cAMP and protein kinase C modulators had no significant effect, H-89 (a protein kinase A inhibitor) and hydrogen peroxide reduced ClC-5 currents.

Conclusions:

  • The fundamental properties of ClC-5 currents are conserved and independent of the host cell type (mammalian vs. amphibian).
  • ClC-5 channels are potentially modulated by protein kinase A (PKA) signaling and reactive oxygen species (ROS).
  • These findings contribute to understanding the molecular basis of Dent's disease and ClC-5 channel function.

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