The calcium-dependent chloride conductance mediator pCLCA1.
The regulatory behavior, inhibitor sensitivity, and 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, 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. Direct involvement of calcium in the activation of pCLCA1 was supported by the finding that treatment with the intracellular calcium chelator 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid-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 despite 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 12-myristate 13-acetate 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 detection of outwardly rectifying chloride current and inhibition of this current by chloride conductance inhibitors in a whole cell patch-clamp study.
Loewen
Matthew E
ME
Department of Veterinary Biomedical Sciences, Western College of Veterinary Medicine, University of Saskatchewan, Canada S7N 5B4.
Gabriel
Sherif E
SE
Forsyth
George W
GW
D016475:3T3 Cells; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid / Q000494:pharmacology; D000818:Animals; D000838:Anions / Q000378:metabolism; D002118:Calcium / Q000502:physiology; D002614:Chelating Agents / Q000494:pharmacology; D018118:Chloride Channels / Q000037:antagonists & inhibitors / Q000235:genetics / Q000502:physiology*; D002712:Chlorides / Q000037:antagonists & inhibitors / Q000502:physiology; D016384:Consensus Sequence; D004533:Egtazic Acid / Q000031:analogs & derivatives* / Q000494:pharmacology; D004553:Electric Conductivity; D015759:Ionomycin / Q000494:pharmacology; D007476:Ionophores / Q000494:pharmacology; D051379:Mice; D020033:Protein Isoforms / Q000037:antagonists & inhibitors / Q000502:physiology; D013552:Swine; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D000838:Anions; C492109:CLCA1 protein, human; D002614:Chelating Agents; D018118:Chloride Channels; D002712:Chlorides; C116464:Clca3a1 protein, mouse; D007476:Ionophores; D020033:Protein Isoforms; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; D004533:Egtazic Acid; D015759:Ionomycin; D013755:Tetradecanoylphorbol Acetate; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; D002118:Calcium