Kinase regulation of hENaC mediated through a region in the COOH-terminal portion of the alpha-subunit.
In an effort to gain insight into how kinases might regulate epithelial Na(+) channel (ENaC) activity, we expressed human ENaC (hENaC) in Xenopus oocytes and examined the effect of agents that modulate the activity of some kinases. Activation of protein kinase C (PKC) by phorbol ester increased the activity of ENaC, but only in oocytes with a baseline current of <2,000 nA. Inhibitors of protein kinases produced varying effects. Chelerythrine, an inhibitor of PKC, produced a significant inhibition of ENaC current, but calphostin C, another PKC inhibitor, had no effect. The PKA/protein kinase G inhibitor H-8 had no effect, whereas the p38 mitogen-activated protein kinase inhibitor, SB-203580 had a significant inhibitory effect. Staurosporine, a nonspecific kinase inhibitor, was the most potent tested. It inhibited ENaC currents in both oocytes and in M-1 cells, a model for the collecting duct. Site-directed mutagenesis revealed that the staurosporine effect did not require an intact COOH terminus of either the beta- or gamma-hENaC subunit. However, an intact COOH terminus of the alpha-subunit was required for this effect. These results suggest that an integrated kinase network regulates ENaC activity through an action that requires a portion of the alpha-subunit.
Volk
K A
KA
Department of Internal Medicine, University of Iowa and Department of Veterans Affairs Medical Center, Iowa City, Iowa 52242, USA.
Husted
R F
RF
Snyder
P M
PM
Stokes
J B
JB
D000818:Animals; D004791:Enzyme Inhibitors / Q000494:pharmacology; D053503:Epithelial Sodium Channels; D005260:Female; D006801:Humans; D066298:In Vitro Techniques; D016297:Mutagenesis, Site-Directed; D009865:Oocytes / Q000187:drug effects / Q000378:metabolism; D011493:Protein Kinase C / Q000378:metabolism; D047428:Protein Kinase Inhibitors; D011494:Protein Kinases / Q000378:metabolism*; D020836:Protein Structure, Quaternary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D015222:Sodium Channels / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D019311:Staurosporine / Q000494:pharmacology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014982:Xenopus laevis
D004791:Enzyme Inhibitors; D053503:Epithelial Sodium Channels; D047428:Protein Kinase Inhibitors; D011994:Recombinant Proteins; D015222:Sodium Channels; D011494:Protein Kinases; D011493:Protein Kinase C; D019311:Staurosporine; D013755:Tetradecanoylphorbol Acetate