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Am J Physiol Cell Physiol (April 10, 2002). doi:10.1152/ajpcell.00477.2001
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Articles in PresS, published online ahead of print April 10, 2002
Am J Physiol Cell Physiol, 10.1152/ajpcell.00477.2001
Submitted on October 9, 2001
Accepted on March 27, 2002

The calcium-dependent chloride conductance mediator pCLCA1

Matthew E Loewen1, Sherif E Gabriel2, and George W Forsyth1*

1 Veterinary Biomedical Sciences, University of Saskatchewan, Saskatoon, SK, Canada
2 Pediatrics, University of North Carolina, Chapel Hill, NC, USA

* To whom correspondence should be addressed. E-mail: george.forsyth{at}usask.ca.

The regulatory behavior, inhibitor sensitivity and the properties of the whole cell chloride conductance observed in cells expressing the cDNA coding for a chloride conductance mediator isoform of the CLCA gene family, called pCLCA1, have been studied. Common C-kinase consensus phosphorylation sites between pCLCA1 and the closely related human isoform hCLCA1 are consistent with a role for calcium in channel activation. Both channels are activated rapidly on exposure to the calcium ionophore, ionomycin. The direct involvement of calcium in the activation of pCLCA1 was supported by the finding that treatment with the intracellular calcium chelator, BAPTA-AM, reduced the rate of chloride efflux from NIH 3T3 cells expressing the pCLCA1 channel. No combination of A-kinase activators used was effective in activating chloride efflux via this channel in spite of the presence of a unique strong A-kinase consensus site in pCLCA1. Notable differences of pCLCA1 from the reported properties of CLCA family members include the failure of phorbol myristoylacetate to activate chloride efflux in cells expressing pCLCA1, and a lack of inhibition of chloride efflux from these cells after treatment with DIDS or dithiothreitol. However, selected inhibitors of anionic conductance inhibited pCLCA1-dependent anion efflux. The electrogenic nature of the ionomycin-dependent efflux of chloride from cells expressing pCLCA1 was confirmed by the detection of outwardly rectifying chloride current and inhibition of this current by chloride conductance inhibitors in a whole-cell patch clamp study.




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