Proteolytic channel activation is a unique feature of the epithelial sodium channel (ENaC) and is thought to play an important role in channel regulation under (patho-)physiological conditions. ENaC is a heteromultimeric channel composed of three homologous subunits (α, β, γ). A fourth subunit, δENaC, has been identified in human tissues but is absent in mouse and rat. The δ-subunit may replace the α-subunit in certain tissues but its physiological role is not yet known. Proteolytic ENaC activation involves cleavage at specific sites in the extracellular domains of the α- and γ-subunits. In humans, cleavage of δENaC may also contribute to channel activation (1). Cleavage results in the release of inhibitory peptides and probably activates the channel by changing its conformation (2). At present the precise molecular mechanisms and the (patho-)physiologically relevant proteases involved in proteolytic ENaC activation remain to be determined.

In nephrotic syndrome filtered plasminogen is converted to the serine protease plasmin by tubular urokinase. We have shown that plasmin activates ENaC which may contribute to renal sodium retention in nephrotic syndrome (3). Recently, we identified two cleavage sites (prostasin site γRKRK178 and plasmin site γK189) in the γ-subunit of human ENaC with functional importance for its proteolytic activation by plasmin. Moreover, we identified for the first time a preferential cleavage site for the serine protease chymotrypsin (γFF174). In a time course experiment we demonstrated that activation of ENaC currents correlates nicely with the appearance of ENaC cleavage fragments at the cell surface (4).

In addition to serine proteases, cathepsin proteases from the group of cysteine proteases may activate ENaC. Cathepsin proteases play a pathophysiological role in inflammatory diseases. We showed that cathepsin-S (Cat-S) activates ENaC and that this stimulation was associated with the appearance of a γENaC cleavage fragment at the plasma membrane. Interestingly, mutating two valine residues (V182;V193) in the critical region of γENaC prevented proteolytic activation of ENaC by Cat-S (5).

In summary, we identified cleavage sites in a region critical for proteolytic channel activation in the γ-subunit of human ENaC with functional importance for its proteolytic activation by plasmin, chymotrypsin and Cat-S. The fact that γENaC contains several different cleavage sites in this region is likely to have physiological implications. It may provide a mechanism for differential ENaC regulation by tissue-specific proteases.

(1) Haerteis et al. (2009) J Biol Chem 284, 29024-29030.

(2) Haerteis et al. (2012) Cell Physiol Biochem 29, 761-774.

(3) Svenningsen et al. (2009) J Am Soc Nephrol 20, 299-310.

(4) Haerteis et al. (2012) J Gen Physiol 140, 375-389.

(5) Haerteis et al. (2012) Pflügers Archiv 464, 353-365.