Comparison of amphibian and human ClC-5: Similarity of functional properties and inhibition by external pH

L. Mo, H. L. Hellmich, P. Fong, T. Wood, J. Embesi, N. K. Wills

Research output: Contribution to journalArticle

40 Scopus citations

Abstract

Loss of function mutations of the renal chloride channel, ClC-5, have been implicated in Dent's disease, a genetic disorder characterized by low weight proteinuria, hypercalciuria, nephrolithasis and, in some cases, eventual renal failure. Recently, our laboratory used an RT-PCR/RACE cloning strategy to isolate an amphibian cDNA from the renal epithelial cell line A6 that had high homology to human ClC-5. We now report a full-length native ClC-5 clone (xClC-5, containing 5' and 3' untranslated regions) isolated by screening a cDNA library from A6 cells that was successfully expressed in Xenopus oocytes. In addition, we compared the properties of xClC-5 and hClC- 5 using isogenic constructs of xClC-5 and hClC-5 consisting of the open reading frame subcloned into an optimized Xenopus expression vector. Expression of the full-length 'native' xClC-5 clone resulted in large, strongly rectifying, outward currents that were not significantly affected by the chloride channel blockers DIDS, DPC, and 9AC. The anion conductivity sequence was NO3/- > Cl- = I- > HCO3/- >> glutamate for xClC-5 and NO3/- > Cl- > HCO3/-> I- >> glutamate for hClC-5. Reduction of the extracellular pH (pH(o)) from 7.5 to 5.7 inhibited outward ClC-5 currents by 27 ± 9% for xClC-5 and 39 ± 7% for hClC-5. The results indicate that amphibian and mammalian ClC-5 have highly similar functional properties. Unlike hClC-5 and most other ClC channels, expression of xClC-5 in oocytes does not require the removal of its untranslated 5' and 3' regions. Acidic solutions inhibited both amphibian and human ClC-5 currents, opposite to the stimulatory effects of low external pH on other ClC channels, suggesting a possibly distinct regulatory mechanism for ClC-5 channels.

Original languageEnglish (US)
Pages (from-to)253-264
Number of pages12
JournalJournal of Membrane Biology
Volume168
Issue number3
DOIs
StatePublished - Apr 24 1999

Keywords

  • Amphibian renal cell line
  • Chloride channel
  • ClC-5
  • Human
  • Outward rectification
  • Xenopus laevis oocytes
  • pH

ASJC Scopus subject areas

  • Biophysics
  • Physiology
  • Cell Biology

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