Endocytosis: A Turnover Mechanism Controlling Ion Channel Function

Abstract

Ion channels (IChs) are transmembrane proteins that selectively drive ions across membranes. The function of IChs partially relies on their abundance and proper location in the cell, fine-tuned by the delicate balance between secretory, endocytic, and degradative pathways. The disruption of this balance is associated with several diseases, such as Liddle's and long QT syndromes. Because of the vital role of these proteins in human health and disease, knowledge of ICh turnover is essential. Clathrin-dependent and -independent mechanisms have been the primary mechanisms identified with ICh endocytosis and degradation. Several molecular determinants recognized by the cellular internalization machinery have been discovered. Moreover, specific conditions can trigger the endocytosis of many IChs, such as the activation of certain receptors, hypokalemia, and some drugs. Ligand-dependent receptor activation primarily results in the posttranslational modification of IChs and the recruitment of important mediators, such as β-arrestins and ubiquitin ligases. However, endocytosis is not a final fate. Once internalized into endosomes, IChs are either sorted to lysosomes for degradation or recycled back to the plasma membrane. Rab proteins are crucial participants during these turnover steps. In this review, we describe the major ICh endocytic pathways, the signaling inputs triggering ICh internalization, and the key mediators of this essential cellular process.

Keywords: endocytosis; ion channels; turnover; ubiquitination.

Figure 1

Schematic representation of the clathrin-coated pit (CCP) structure. Clathrin, the major protein in CCPs, forms a triskelion that polymerizes to form a hexagonal coat that covers the membrane. The AP-2 complex links clathrin to the membrane and coordinates the assembly of the coat with cargo proteins and lipids. In addition, epsins couple ubiquitinated membrane proteins into CCPs and contribute to the membrane curvature during the formation of clathrin-coated buds. Finally, dynamin catalyzes the constriction of the “neck” of the membrane invagination and the scission of the clathrin-coated vesicles (CCVs) from the plasma membrane. See the text for further details. Color code: red, clathrin; orange, AP-2 complex; blue, epsin; black, dynamin; light red circle, ubiquitin; and pink, ICh.

Figure 2

The endocytic ICh network. The balance between the secretory pathway and the endosomal network regulates ICh abundance at the plasma membrane. The endocytic network starts with the internalization of protein cargo by distinct mechanisms (1). ICh exhibits constitutive turnover but undergoes massive internalization upon specific stimulation. Thus, receptor activation (e.g., GPCR activation) provides a platform by which β-arrestins associate with receptors and ICh to mediate channel endocytosis (2). Receptor stimulation also triggers posttranslational modifications of IChs, such as phosphorylation (P) and ubiquitination (UBQ), which is realized through specific kinases and ubiquitin ligases (E3), respectively, which induce ICh endocytosis (3). Different internalization routes for IChs can be classified by their dependence on clathrin in (i) clathrin-mediated endocytosis (CME) (4) or (ii) clathrin-independent endocytosis (CIE) (5). CME is the most common internalization pathway for IChs. CIE mechanisms can be classified as caveolin-, RhoA- or ARF6-mediated. Internalized vesicles fuse to early endosomes (EEs) and are partially sorted (6). Target channels can be recycled (either back to the plasma membrane or to the secretory pathway) (7) or degraded in lysosomes (8). Recycling to the plasma membrane is achieved through tubule-vesicular transport carriers, known as the “fast recycling” process (9), or by the endocytic recycling compartment (ERC), known as the “slow recycling” process (10). Moreover, vesicles can bypass the trans-Golgi network and enter the secretory pathway (7). Therefore, there is an interplay between E3 ligases and deubiquitinating (DUB) enzymes. Ubiquitin is released from ubiquitinated substrates by DUBs. IChs sorted for degradation continue to move through the endocytic pathway from the EEs to the late endosomes (LEs) and are ultimately degraded by lysosomes. Rab proteins guide these turnover stages. Rab5 and Rab4 collaborate to regulate channel entry into and exit from EEs, respectively. Thus, Rab5 regulates the fusion of endocytic vesicles with EEs, whereas Rab4 is involved in the “fast recycling” pathway. On the other hand, Rab7 mediates the vacuolar fusion of EEs with LEs, and Rab11 participates in the “slow recycling” pathway. See the text for further details.