Cargo- and compartment-selective endocytic scaffold proteins

Abstract

The endocytosis of membrane receptors is a complex and tightly controlled process that is essential for maintaining cellular homoeostasis. The removal of receptors from the cell surface can be constitutive or ligand-induced, and occurs in a clathrin-dependent or -independent manner. The recruitment of receptors into specialized membrane domains, the formation of vesicles and the trafficking of receptors together with their ligands within endocytic compartments are regulated by reversible protein modifications, and multiple protein-protein and protein-lipid interactions. Recent reports describe a variety of multidomain molecules that facilitate receptor endocytosis and function as platforms for the assembly of protein complexes. These scaffold proteins typically act in a cargo-specific manner, recognizing one or more receptor types, or function at the level of endocytic cellular microcompartments by controlling the movement of cargo molecules and linking endocytic machineries to signalling pathways. In the present review we summarize present knowledge on endocytic scaffold molecules and discuss their functions.

Figure 1. CIN85 is an endocytic scaffold

CIN85 is an example of a multimeric adaptor that is involved in cargo internalizaton and sorting. Through its multiple domains, CIN85 associates with a number of proteins, which are divided here into functional groups. CIN85 oligomerizes and simultaneously binds various molecules, changing its conformation and binding partners upon cell stimulation, thus co-ordinating the trafficking of several cargo proteins. Abbreviations: STAP, stem-cell-specific adaptor protein containing pleckstrin homology and SH2 domains; BLNK, B cell linker protein; SHIP-1, SH2-containing inositol phosphatase-1; GEF, guanine nucleotide exchange factor; ASAP1, Arf-GTPase-activating protein 1; ARAP3, Arf GAP and Rho GAP with ankyrin repeat and pleckstrin homology domains protein 3; PDGFR, platelet-derived growth factor receptor; HGFR, hepatocyte growth factor receptor; PI3K, phosphoinositide 3-kinase; PAK, p21-activated kinase; SFK, Src family kinase; CC, coiled-coil.

Figure 2. Examples of dynamic protein–lipid complexes that couple the cargo with the biogenesis and trafficking of vesicles in endocytic microcompartments

(A) Endophilin co-ordinates cargo entry and actin polymerization at clathrin-coated pits. Its BAR domain senses and induces membrane curvature. (B) The BAR domain of amphiphysin binds to curved negatively charged phospholipids, while its other domains associate with AP-2, clathrin and dynamin. (C) Epsin interacts with clathrin, AP-2, Eps15, intersectin and ubiquitin (purple oval). Its ENTH domain binds PtdIns(4,5)P₂ [PI(4,5)P₂] and introduces membrane curvature. (D) The Rab domain at the level of the early endosome is a functional protein multimer that is coupled to the membrane phospholipid PtdIns3P via binding of FYVE-domain-containing proteins. For details, see the text.

Figure 3. Ubiquitin traffic

Lysosomal degradation of ubiquitinated receptors is directed by the sequential action of the ubiquitin-binding protein complex composed of Hrs, Tsg101 and Alix with associated proteins. All three molecules form oligomers. Hrs and Tsg101 bind ubiquitin directly via UIM (ubiquitin-interacting motif) and UEV (ubiquitin-conjugating enzyme E2 variant) domains respectively. Hrs binds PtdIns3P [PI(3)P] and sequesters cargo in clathrin-coated regions of early endosomes, preventing it from recycling. The UEV domain of Tsg101 associates with PSAP motifs in Hrs and Alix (block arrows). Tsg101 and Alix co-ordinate the function of ESCRT protein complexes involved in the biogenesis of MVBs. Alix associates with LPBA, a lipid that is necessary for the formation of inner vesicles of MVBs. Upon ligand stimulation, several accessory proteins, including Hrs, undergo mono-ubiquitnation (indicated by a purple asterisk). This has been implicated in co-ordinating vesicular transport. The endocytic sorting machinery is recruited during the budding of RNA viruses, whose Gag proteins are ubiquitinated and have motifs that bind Tsg101 and Alix. Abbreviations: VHS, Vsp27p, Hrs, STAM; CC, coiled-coil; SNAP-25, 25 kDa synaptosome-associated protein; BRD, Bro1-rhophilin conserved domain; CHMP4, charged MVB protein; ALG, apoptosis-linked gene 2.