Downregulation of epithelial sodium channel (ENaC) by CFTR co-expressed in Xenopus oocytes is independent of Cl- conductance.
Defective regulatory interactions between the cystic fibrosis conductance regulator (CFTR) and the epithelial sodium channel (ENaC) have been implicated in the elevated Na+ transport rates across cystic fibrosis airway epithelium. It has recently been proposed that ENaC downregulation by CFTR depends on the ability of CFTR to conduct Cl- into the cell and is negligible when Cl- flows out of the cell. To study the mechanisms of this downregulation we have measured amiloride-inhibitable Na+ current (Iamil) in oocytes co-expressing rat ENaC and human wild-type CFTR. In oocytes voltage-clamped to -60 mV, stimulating CFTR with 1 mm IBMX reduced Iamil by up to 80%, demonstrating that ENaC is inhibited when Cl- is conducted out of the cell. Decreasing the level of CFTR stimulation in a single oocyte, decreased both the degree of Iamil downregulation and the CFTR-mediated plasma membrane Cl- conductance, suggesting a direct correlation. However, Iamil downregulation was not affected when Cl- flux across oocyte membrane was minimized by holding the oocyte membrane potential near the Cl- reversal potential (67% +/- 10% inhibition at -20 mV compared to 79% +/- 4% at -60 mV) demonstrating that Iamil downregulation was independent of the amount of current flow through CFTR. Studies with the Ca2+-sensitive photoprotein aequorin showed that Ca2+ is not involved in Iamil downregulation by CFTR, although Ca2+ injection into the cytoplasm did inhibit Iamil. These results demonstrate that downregulation of ENaC by CFTR depends on the degree of CFTR stimulation, but does not involve Ca2+ and is independent of the direction and magnitude of Cl- transport across the plasma membrane.
Chabot
H
H
Centre de recherche, Centre hospitalier de l'Université de Montréal, Campus Hôtel-Dieu, 3850 Saint-Urbain, Montréal, Québec, Canada, H2W 1T8.
Vives
M F
MF
Dagenais
A
A
Grygorczyk
C
C
Berthiaume
Y
Y
Grygorczyk
R
R
D000818:Animals; D018118:Chloride Channels / Q000502:physiology; D002712:Chlorides / Q000502:physiology*; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000502:physiology*; D015536:Down-Regulation; D004847:Epithelial Cells / Q000502:physiology*; D006801:Humans; D018408:Patch-Clamp Techniques; D051381:Rats; D015222:Sodium Channels / Q000502:physiology*; D014981:Xenopus
C505032:CFTR protein, human; D018118:Chloride Channels; D002712:Chlorides; D015222:Sodium Channels; D019005:Cystic Fibrosis Transmembrane Conductance Regulator