Single Nucleotide Variant Increases the open Probability of Human ENaC

Abstract

The epithelial Na+ channel (ENaC) mediates Na+ entry into specific epithelial cells and has an essential role in the regulation of blood pressure and airway surface liquid volume. Rare ENaC mutations have been reported in inherited disorders associated with hypertension or hypotension, and ENaC gene variants have been associated with several human disorders. Recent human genome sequencing projects have revealed a large number of ENaC gene variations in both exons and introns. However, the functional consequences of most variants are unknown. We examined the functional properties of the human ENaC variant γL511Q in the Xenopus oocyte expression system. Oocytes expressing αβγL511Q exhibited five-fold greater amiloride-sensitive currents than cells expressing WT channels. Mutant and WT channels had similar levels of surface expression. Single channel recordings with a cell-attached patch revealed that mutant channels had a four-fold higher open probability than WT, but similar unitary currents. The mutant had a significantly reduced Na+ self-inhibition response, reflecting less reduction of ENaC open probability by extracellular Na+. Interestingly, the mutant diminished the activating effect of external Zn2+ and essentially converted Zn2+ from a high-affinity ENaC activator to a low-affinity inhibitor. Furthermore, γL511Q exhibited blunted activation by chymotrypsin. We conclude that γL511Q is a gain-of-function human ENaC variant that is characterized by increased open probability and suppressed responses to extracellular Na+, Zn2+ and chymotrypsin.