Conservation of pH sensitivity in the epithelial sodium channel (ENaC) with Liddle's syndrome mutation

Abstract

Gain-of-function mutations of the epithelial Na+ channel (ENaC) cause a rare form of hereditary hypertension, Liddle's syndrome. How these mutations lead to increased channel activity is not yet fully understood. Since wild-type ENaC (wt-ENaC) is highly pH-sensitive, we wondered whether an altered pH-sensitivity of ENaC might contribute to the hyperactivity of ENaC with Liddle's syndrome mutation (Liddle-ENaC). Using Xenopus laevis oocytes as an expression system, we compared the pH-sensitivity of wt-ENaC (alphabetagammarENaC) and Liddle-ENaC (alphabeta(R564stop)gammarENaC). Oocytes were assayed for an amiloride-sensitive (2 microM) inward current (deltaIami) at -60 mV holding potential and cytosolic pH was altered by changing the extracellular pH in the presence of 60 mM sodium acetate. Alternatively, cytosolic acidification was achieved by proton loading the cells using a proton-coupled oligopeptide transporter (PepT-1) co-expressed in the oocytes together with ENaC. Cytosolic but not extracellular acidification substantially reduced deltaIami while cytosolic alkalinisation had a stimulatory effect. This pH-sensitivity was largely preserved in oocytes expressing Liddle-ENaC. The inhibition of wt-ENaC and Liddle-ENaC by cytosolic acidification was independent of so-called sodium-feedback inhibition, since it was not associated with a concomitant increase in intracellular Na+ concentration estimated from the reversal potential of deltaIami. In addition C-terminal deletions in the alpha or gamma subunits or in all three subunits of ENaC did not abolish the inhibitory effect of cytosolic acidification. We conclude that ENaC's pH-sensitivity is not mediated by its cytoplasmic C-termini and that an altered pH-sensitivity of ENaC does not contribute to the pathophysiology of Liddle's syndrome.

Expression and localization of rat NBC4c in liver and renal uroepithelium

“…A similar mechanism has been reported in the tongue, where altered cellular Na ϩ flux in non-taste cells augments the depolarization of taste cells (38). ENaC channel activity and neuronal activity are known to be pH sensitive (32). The importance of pH i in modulating both Na ϩ channel activity and neurosensory responses underscores an additional potentially important role for NBC4c in preventing large changes in the pH of the microenvironment of mechanosensory nerve fibers in the pelvic wall.…”

Mechanisms of ENaC Regulation and Clinical Implications

“…Intracellular acidification also inhibits ␣-ENaC, and intracellular alkalinization stimulates ENaC activity through an increase in P o . 125,126 Hypoxia also has direct effects on expression of ENaC subunits and activity in rodent airway epithelial cells. 127,128 Further clinical correlations of these phenomena are warranted to ascertain their physiologic or pathophysiologic importance.…”

NADPH Oxidase-dependent Acid Production in Airway Epithelial Cells

“…Several antimicrobial factors have been isolated from airway secretions indicating the role of the airways in bacterial defense. Production of O 2 . by airway cultures by NADPH oxidase has been shown (24).…”

“…The phagocytic NADPH oxidase is a multisubunit enzyme consisting of two transmembrane subunits (the cytochrome gp91 phox and p22 phox ), three cytosolic subunits (p40 phox , p47 phox , and p67 phox ), and a small ubiquitously expressed GTP-binding protein p21 rac (19). In phagocytes, all subunits were shown to be necessary for a regulated release of O 2 . during the respiratory burst (20).…”