SNX-BAR proteins in phosphoinositide-mediated, tubular-based endosomal sorting

Abstract

The endocytic network is morphologically characterized by a wide variety of membrane bound compartments that are able to undergo dynamic re-modeling through tubular and vesicular structures. The precise molecular mechanisms governing such re-modeling, and the events that co-ordinated this with the major role of endosomes, cargo sorting, remain unclear. That said, recent work on a protein family of sorting nexins (SNX) - especially a subfamily of SNX that contain a BAR domain (SNX-BARs) - has begun to shed some much needed light on these issues and in particular the process of tubular-based endosomal sorting. SNX-BARs are evolutionary conserved in endosomal protein complexes such as retromer, where they co-ordinate membrane deformation with cargo selection. Furthermore a central theme emerges of SNX-BARs linking the forming membrane carrier to cytoskeletal elements for transport through motor proteins such as dynein. By studying these SNX-BARs, we are gaining an increasingly detailed appreciation of the mechanistic basis of endosomal sorting and how this highly dynamic process functions in health and disease.

Figure 1. Evolutionary conservation and homology of the different SNX–BAR proteins

(A) Phylogenic tree of all SNX–BAR proteins annotated in the human genome. The following phylogenic trees show the relation between human SNX–BAR with known homologues in Mus musculus, Danio rerio, Drosophila melanogaster, Caenorhabditis elegans and Saccharomyces cerevisiae of (B) SNX–BARs involved in endosomal recycling SNX4, and possibly SNX7 and SNX30, (C) SNX–BARs involved in endosome-to-TGN traffic (SNX1, SNX2, SNX5, SNX6 and SNX8, and possibly SNX32) and (D) SNX–BARs with a SH3 domain, possibly involved in clathrin-mediated endocytosis and endosomal sorting (SNX9, SNX18 and SNX33). Bars indicate substitutions per site.

Figure 2. Membrane association/deformation by SNX–BARs

(A) Crystal structure of the SNX9 BAR-PX dimer; one SNX9 molecule is shown in red/orange/yellow, the other in green/cyan/blue. Red/blue indicate the two PX domains, yellow/green the BAR dimerization domain and orange/cyan the so-called yoke domain, which links the PX domain to the BAR domain [16]. Space-fill models showing the electrostatic distribution on the surface of the SNX9 dimer, red indicates acidic, negative residues while blue indicates positive, basic residues. Note the dominant blue patches on the concave surface, binding the lipid bilayer [16]. (B) A highly speculative view for the oligomerization of SNX–BARs based on the tip-to-tip and lateral contact model for F-BARs [5]. With the presence of the PX domains on either side of the BAR dimer, lateral interactions may occur through two possible routes: via association of a PX domain with its neighbouring lateral BAR domain (top), or through PX domain lateral contact with a neighbouring PX domain (bottom).

Figure 3. Hypotheses on how SNX–BARs could deform membranes

Cartoon of the different models how SNX–BARs could deform membranes, the SNX–BAR domains are shown in similar colours as figure 2. (A). Detection of phosphoinositides in the flat membrane allow the PX domains of the SNX–BAR dimer to bind and amphipathic helixes to insert into the lipid bilayer, which produces local curvature. This local curvature is stabilized and extended to form a deformation by oligomerization of the SNX–BAR domains. (B). Co-incidence detection of both local curvature and phosphoinositides by the PX domain and the BAR domain of the SNX–BAR dimer. This local curvature is extended to global deformation by oligomerization of the SNX–BAR domains. (C). Co-incidence detection of phosphoinositides by the PX domain and curvature by the amphipathic helix (shown as cyan/orange circles) [27]. On the curved membrane BAR domains dimerize to stabilize and extend the local curvature.

Figure 4. SNX–BAR regulated sorting in the maturing endosome

Cartoon illustrating the maturation of endosomes from endocytosis through to fusion with the lysosome, and how different SNX–BARs may regulate distinct tubular-based sorting events. The maturation of the endosome coincides with changes in phosphoinositide composition, the presence of different Rab proteins and the increase in number of intraluminal vesicles. In brief, membrane is internalized in PI(4,5)P₂-enriched pits. SNX9 plays an important role in this internalization (see text Section 3.4). The endocytic vesicle fuses with the early endosome. In this compartment, proteins are sorted for different destinations while the early endosomal vacuole matures into a late endosome. Proteins targeted for degradation are collected in intraluminal vesicles, which the endosome accumulates during maturation. However, several proteins are retrieved away from this pathway. The transferrin receptor is recycled, at relatively early stages of endosomal maturation, to the plasma membrane either through fast recycling or more slowly via the endosomal recycling compartment (ERC). SNX4 has been established to play a role in regulating tubular-based sorting into the ERC [22], while the roles of SNX7 and SNX30 in this and other recycling pathways are unclear. It also remains to be determined whether SNX4, SNX7 and SNX30 form a restricted series of homo- and/or heterodimeric interactions. The retromer SNX–BARs SNX1, SNX2, SNX5 and SNX6 regulate the retrieval of cation-independent mannose 6-phosphate receptors to the trans-Golgi network (TGN) in a later stage of maturation [33]. SNX8 is also involved in endosome-to-TGN transport, but little is known about its cargo or timing in the maturing endosome [62]. The role(s) of SNX18 and SNX33 remain to be defined ([24,79]. See text (Sections 3.1-3.4) for more detailed discussion of the function of these SNX–BARs.

Figure 5. SNX–BAR recruit motor proteins to assist in fission and long-range transport

Cartoon of different SNX–BAR coats recruiting motor proteins to link the forming membrane carrier to cytoskeletal elements. (A). SNX5 and SNX6 associate with p150^glued, which is part of the dynactin complex that activates the minus-end directed transport on microtubules by the dynein motor thereby generating longitudinal force. (B). Illustrates the association of the SNX4 coat with the dynein motor [22,72], again generating longitudinal force. (C). SNX9 and SNX33 coats associate with N-WASP, which activates the Arp2/3 complex to polymerize actin filaments. By stimulating this actin polymerization, longitudinal force is generated that may assist the efficiency of membrane fission.