Expression of ENaC and other Transport Proteins in Xenopus Oocytes is Modulated by Intracellular Na+

Abstract

The expression of the epithelial Na⁺ channel (ENaC) is tissue-specific and dependent on a variety of mediators and interacting proteins. Here we examined the role of intracellular Na⁺ ([Na⁺]_i) as a modulator of the expression of rat ENaC in Xenopus laevis oocytes. We manipulated [Na⁺]_i of ENaC-expressing oocytes in the range of 0-20 mM by incubating in extracellular solutions of different [Na⁺]_o. Electrophysiological, protein biochemical and fluorescence optical methods were used to determine the effects of different [Na⁺]_i on ENaC expression and membrane abundance. In voltage-clamp experiments we found that amiloride-sensitive ENaC current (I_ami) and conductance (G_ami) peak at a [Na⁺]_i of ∼10 mM Na⁺, but were significantly reduced in 5 mM and 20 mM [Na⁺]_i. Fluorescence intensity of EGFP-ENaC-expressing oocytes also followed a bell-shaped curve with a maximum at ∼ 10 mM [Na⁺]_i. In Western blot experiments with specific anti-ENaC antibodies the highest protein expression was found in ENaC-expressing oocytes with [Na⁺]_i of 10-15 mM. Since ENaC is also highly permeable for Li⁺, we incubated ENaC-expressing oocytes in different Li⁺ concentrations and found a peak of I_ami and G_ami with 5 mM Li⁺. The influence of [Na⁺]_i on the expression is not ENaC-specific, since expression of a Cl^- channel (CFTR) and a Na⁺/glucose cotransporter (SGLT1) showed the same dependence on [Na⁺]_i. We conclude that specific concentrations of Na⁺ and Li⁺ influence the expression and abundance of ENaC and other transport proteins in the plasma membrane in Xenopus laevis oocytes. Furthermore, we suggest the existence of a general mechanism dependent on monovalent cations that optimizes the expression of membrane proteins.