Abstract
Canonical epithelial sodium channels (ENaCs) are heterotrimers formed by α, β, and γ ENaC subunits in vertebrates and belong to the Degenerin/ENaC family of proteins. Proteins from this family form mechanosensitive channels throughout the animal kingdom. Activity of canonical ENaC is regulated by shear force (SF) mediating Na+ absorption in the kidney and vascular tone of arteries. Expression analysis suggests that non-canonical ENaC, formed by single or only two subunits, exist in certain tissues, but it is unknown if these channels respond to SF. α, β, γ, and δ ENaC subunits were expressed either alone or in combinations of two subunits in Xenopus oocytes. Amiloride-sensitive currents and the responses to SF were assessed using two-electrode voltage clamp recordings. With the exception of γ ENaC, all homomeric channels provided amiloride-sensitive currents and responded to SF applied via a fluid stream directed onto the oocytes. Channels containing two subunits were also activated by SF. Here, the presence of the γ ENaC subunit when co-expressed with α or δ augmented the SF response in comparison to the αβγ/δβγ ENaC. Overall, we provide evidence that non-canonical ENaC can form channels that respond to SF. This supports a potential function of non-canonical ENaC as mechanosensors in epithelial, vascular, and sensory cells.
Highlights
The epithelial sodium channel (ENaC) proteins belong to the degenerin/ENaC (DEG/ENaC) superfamily of proteins (Mano and Driscoll, 1999)
The aim of this study was to determine whether individual ENaC subunits can form functional channels that respond to shear force (SF)
To determine whether or not individual ENaC subunits can form functional homomeric ion channels, preliminary experiments were performed in Xenopus oocytes injected with mRNA encoding one subunit
Summary
The epithelial sodium channel (ENaC) proteins belong to the degenerin/ENaC (DEG/ENaC) superfamily of proteins (Mano and Driscoll, 1999). Canonical ENaC consists of three homologous subunits, known as α, β, and γ, that form a functional heteromultimeric channel (Canessa et al, 1994a). ENaC assembles with a 1:1:1 stoichiometry of α:β:γ subunits arranged in a counter-clockwise manner as revealed by cryo-electron microscopy (Noreng et al, 2018). Another subunit known as δ has been identified in humans (Waldmann et al, 1995), primates (Giraldez et al, 2007), and Xenopus (Wichmann et al, 2018) but is not expressed in mice or rats (Giraldez et al, 2012). Each individual ENaC subunit is made of 650–700 amino acids with a molecular weight of around 70–100 kDa, two transmembrane domains, short intracellular Nand C-termini, and a large extracellular-domain that consists of70% of the protein (Canessa et al, 1994a; Snyder et al, 1994; Noreng et al, 2018)
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