ENaC at the Cutting Edge: Regulation of Epithelial Sodium Channels by Proteases

Abstract

Epithelial Na ؉ channels facilitate the transport of Na ؉ across high resistance epithelia. Proteolytic cleavage has an important role in regulating the activity of these channels by increasing their open probability. Specific proteases have been shown to activate epithelial Na ؉ channels by cleaving channel subunits at defined sites within their extracellular domains. This minireview addresses the mechanisms by which proteases activate this channel and the question of why proteolysis has evolved as a mechanism of channel activation.Many ion channels are silent at rest and are activated in response to a variety of factors, including membrane potential, external ligands, and intracellular signaling processes. The ENaC 2 has evolved as a channel that is thought to reside primarily in an active state, facilitating the bulk movement of Na ϩ out of renal tubular or airway lumens. The regulated insertion and retrieval of channels at the plasma membrane have important roles in modulating ENaC-dependent Na ϩ transport (1). A number of factors also have a role in regulating ENaC activity via changes in channel P o or gating. In this regard, it has become increasingly apparent that proteolysis of ENaC subunits has a key role in this process (2). This minireview addresses several questions regarding the role of ENaC subunit proteolysis in regulating channel gating. (i) Where are ENaC subunits cleaved? (ii) Which proteases mediate ENaC cleavage? (iii) Why are channels activated by proteolysis? (iv) Is proteolysis responsible, in part, for the highly variable channel P o that has been noted for ENaC? (v) Why have ENaCs evolved as channels that require proteolysis for activation? Where Are ENaC Subunits Cleaved?Reports in the early 1980s that serine protease inhibitors reduced transepithelial Na ϩ transport across toad urinary bladder suggested that proteases have a role in activating ENaC (3). A series of studies over the past decade have confirmed that proteases activate ENaC and have begun to elucidate the mechanism by which this occurs. Following the observation that ENaC activity was significantly reduced in epithelial cells treated with aprotinin and that low concentrations of external trypsin rapidly activated ENaC in aprotinin-pretreated cells, a series of CAPs were identified that activated ENaC when coexpressed in heterologous expression systems (4 -6). Furthermore, channels with a very low P o responded to external trypsin with a dramatic increase in P o (7).What is the target of these proteases? ENaC is composed of three structurally related subunits (␣, ␤, and ␥) that have two membrane-spanning domains connected by a large extracellular loop composed of ϳ450 residues. Early reports suggested that ENaC subunits or closely associated proteins were the protease target (5). Subsequent studies demonstrated that the ␣ and ␥ subunits of ENaC were processed by proteases (8 -11). The presence of full-length forms as well as faster migrating forms of the ␣ and ␥ subunits on SDS-polyacrylamide gels, both in cell lysates and...

Short Forms of Ste20-related Proline/Alanine-rich Kinase (SPAK) in the Kidney Are Created by Aspartyl Aminopeptidase (Dnpep)-mediated Proteolytic Cleavage

“…been inserted in the plasma membrane of epithelial cells (46). ENaC activity increases severalfold upon proteolytic cleavage at the membrane, mediating reabsorption of Na ϩ in the collecting duct (47,48).…”

Inhibitory Tract Traps the Epithelial Na+ Channel in a Low Activity Conformation

“…This processing event transitions channels to a high open probability (P o ) state and tailors channel function to its biological role in bulk Na ϩ transport. We have suggested previously that proteolysis provided a mechanism that allowed ENaCs to evolve from other members of the ENaC/degenerin family that reside primarily in the closed state and transiently open in response to external factors (3). Cleavage likely occurs during channel maturation by proteases in the biosynthetic pathway, as well as at the cell surface (3).…”

“…Rare among ion channels, ENaC is activated by proteolysis (3). This processing event transitions channels to a high open probability (P o ) state and tailors channel function to its biological role in bulk Na ϩ transport.…”

“…We have suggested previously that proteolysis provided a mechanism that allowed ENaCs to evolve from other members of the ENaC/degenerin family that reside primarily in the closed state and transiently open in response to external factors (3). Cleavage likely occurs during channel maturation by proteases in the biosynthetic pathway, as well as at the cell surface (3). ENaC proteolysis has been suggested to play a role in disorders such as nephrotic syndrome and cystic fibrosis, where concentrations of specific protease are elevated in the extracellular milieu (4,5).…”

Af17 Deficiency Increases Sodium Excretion and Decreases Blood Pressure

“…Finally, ENaC is tightly regulated at multiple levels by aldo. Most studies concerning ENaC and aldo focus on posttranslational events (reviewed in 21 ), leaving a gap in knowledge between chromatin modifications and regulation of ENaC transcription, Na ϩ retention and BP.…”