The delta-subunit of the epithelial sodium channel (ENaC) enhances channel activity and alters proteolytic ENaC activation.
The epithelial sodium channel (ENaC) is probably a heterotrimer with three well characterized subunits (alphabetagamma). In humans an additional delta-subunit (delta-hENaC) exists but little is known about its function. Using the Xenopus laevis oocyte expression system, we compared the functional properties of alphabetagamma- and deltabetagamma-hENaC and investigated whether deltabetagamma-hENaC can be proteolytically activated. The amiloride-sensitive ENaC whole-cell current (DeltaI(ami)) was about 11-fold larger in oocytes expressing deltabetagamma-hENaC than in oocytes expressing alphabetagamma-hENaC. The 2-fold larger single-channel Na(+) conductance of deltabetagamma-hENaC cannot explain this difference. Using a chemiluminescence assay, we demonstrated that an increased channel surface expression is also not the cause. Thus, overall channel activity of deltabetagamma-hENaC must be higher than that of alphabetagamma-hENaC. Experiments exploiting the properties of the known betaS520C mutant ENaC confirmed this conclusion. Moreover, chymotrypsin had a reduced stimulatory effect on deltabetagamma-hENaC whole-cell currents compared with its effect on alphabetagamma-hENaC whole-cell currents (2-fold versus 5-fold). This suggests that the cell surface pool of so-called near-silent channels that can be proteolytically activated is smaller for deltabetagamma-hENaC than for alphabetagamma-hENaC. Proteolytic activation of deltabetagamma-hENaC was associated with the appearance of a delta-hENaC cleavage product at the cell surface. Finally, we demonstrated that a short inhibitory 13-mer peptide corresponding to a region of the extracellular loop of human alpha-ENaC inhibited DeltaI(ami) in oocytes expressing alphabetagamma-hENaC but not in those expressing deltabetagamma-hENaC. We conclude that the delta-subunit of ENaC alters proteolytic channel activation and enhances base-line channel activity.
Haerteis
Silke
S
Institut für Zelluläre und Molekulare Physiologie, Friedrich-Alexander-Universität Erlangen-Nürnberg, 91054 Erlangen, Germany.
Krueger
Bettina
B
Korbmacher
Christoph
C
Rauh
Robert
R
D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D002918:Chymotrypsin / Q000494:pharmacology; D053503:Epithelial Sodium Channels / Q000378:metabolism*; D005786:Gene Expression Regulation; D006801:Humans; D049449:Luminescence; D008954:Models, Biological; D009865:Oocytes / Q000378:metabolism*; D018408:Patch-Clamp Techniques; D010455:Peptides / Q000737:chemistry; D017434:Protein Structure, Tertiary; D018075:RNA, Complementary / Q000378:metabolism; D012964:Sodium / Q000737:chemistry; D014982:Xenopus laevis
D053503:Epithelial Sodium Channels; D010455:Peptides; D018075:RNA, Complementary; D012964:Sodium; D002918:Chymotrypsin