Regulation of the epithelial sodium channel by membrane trafficking

Abstract

The epithelial Na(+) channel (ENaC) is a major regulator of salt and water reabsorption in a number of epithelial tissues. Abnormalities in ENaC function have been directly linked to several human disease states including Liddle's syndrome, psuedohypoaldosteronism, and cystic fibrosis and may be implicated in states as diverse as salt-sensitive hypertension, nephrosis, and pulmonary edema. ENaC activity in epithelial cells is highly regulated both by open probability and number of channels. Open probability is regulated by a number of factors, including proteolytic processing, while ENaC number is regulated by cellular trafficking. This review discusses current understanding of apical membrane delivery, cell surface stability, endocytosis, retrieval, and recycling of ENaC and the molecular partners that have so far been shown to participate in these processes. We review known sites and mechanisms of hormonal regulation of trafficking by aldosterone, vasopressin, and insulin. While many details of the regulation of ENaC trafficking remain to be elucidated, knowledge of these mechanisms may provide further insights into ENaC activity in normal and disease states.

Fig. 1.

Schematic diagram outlining steps in epithelial sodium channel (ENaC) endocytosis and progress through early endosomes as it is recycled. Each of the steps in ENaC internalization is discussed in detail in the text. ENaC internalization is initiated by the binding of Nedd4-2 to the intracellular NH₂ terminus (NH₂) of each subunit and the addition of ubiquitin (Ub) moieties to the COOH tail (COOH) of the ENaC subunits (enlarged inset). Binding of 14-3-3 proteins to phosphorylated Nedd4-2 (yellow symbols on Nedd4-2) prevents Nedd4-2 interaction with the PY motifs on ENaC. Following ubiquitination, the channel is internalized by clathrin-dependant endocytosis through an interaction with epsin and passes through an early endosomal compartment (EEA1 and Hrs positive). If ENaC is deubiquitinated by deubiquitinating enzymes (Dubs), it will be transferred to the apical recycling endosome (ARE) for recycling to the apical membrane (see Fig. 2). If it remains ubiquitinated, it is likely trafficked via the late endosomes (LE) to the lysosomes (Lys) for degration. From studies on other transporters, it is likely that Rab proteins may be involved in rescuing ENaC from the LE back to the ARE or facilitating its movement to Lys for degradation.

Fig. 2.

Schematic diagram outlining the steps in ENaC recycling and delivery to the apical membrane. Following synthesis and assembly (yellow Golgi apparatus), channels are transported to the apical surface (yellow vesicle) and can be localized in lipid raft membrane domains (Rafts). It is also possible that ENaC enters the regulated recycling pathway directly (? and arrow), but no support for this idea has yet emerged. Those channels retrieved from the apical surface (see Fig. 1) and delivered to the ARE are trafficked back to the apical membrane through the involvement of Rab proteins (most likely Rab11, 27). Delivery of ENaC to the apical surface is regulated by RhoA and sorting nexin 3 (SN3) and vesicle fusion is mediated by SNARE proteins (SNAREs). Details of the mechanisms of ENaC delivery are provided in the text.

Fig. 3.

Schematic diagram indicating points of hormonal regulation (in red) in the ENaC trafficking pathways previously depicted in Figs. 1 and 2. The actions of aldosterone (Aldo) and vasopressin are highlighted. Binding of Aldo to the mineralocorticoid receptor results in the long-term synthesis of additional ENaC, and in the regulation of Nedd4-2 (as depicted in Fig. 1) by the serum and glucocorticoid kinase (SGK). SGK phosphorylation (P) prevents Nedd4-2 binding and ENaC ubiquitination (Fig. 1, inset). Direct regulation of ENaC by phosphorylation through ERK is prevented by the action of GilZ, another Aldo-induced protein. Both vasopressin and insulin induce the translocation of ENaC from an intracellular vesicle storage population to the apical membrane. For vasopressin, this is facilitated by SN3.