Out of the ESCPE room: Emerging roles of endosomal SNX‐BARs in receptor transport and host–pathogen interaction

Abstract Several functions of the human cell, such as sensing nutrients, cell movement and interaction with the surrounding environment, depend on a myriad of transmembrane proteins and their associated proteins and lipids (collectively termed “cargoes”). To successfully perform their tasks, cargo must be sorted and delivered to the right place, at the right time, and in the right amount. To achieve this, eukaryotic cells have evolved a highly organized sorting platform, the endosomal network. Here, a variety of specialized multiprotein complexes sort cargo into itineraries leading to either their degradation or their recycling to various organelles for further rounds of reuse. A key sorting complex is the Endosomal SNX‐BAR Sorting Complex for Promoting Exit (ESCPE‐1) that promotes the recycling of an array of cargos to the plasma membrane and/or the trans‐Golgi network. ESCPE‐1 recognizes a hydrophobic‐based sorting motif in numerous cargoes and orchestrates their packaging into tubular carriers that pinch off from the endosome and travel to the target organelle. A wide range of pathogens mimic this sorting motif to hijack ESCPE‐1 transport to promote their invasion and survival within infected cells. In other instances, ESCPE‐1 exerts restrictive functions against pathogens by limiting their replication and infection. In this review, we discuss ESCPE‐1 assembly and functions, with a particular focus on recent advances in the understanding of its role in membrane trafficking, cellular homeostasis and host–pathogen interaction.


| The endolysosomal pathway
Endosomes are a series of intracellular membrane-bound compartments that receive transmembrane proteins and associated proteins and lipids (hereinafter referred as to cargoes) from the endocytic and the biosynthetic pathways. The discovery and characterization of endosomes arose from studies exploring Semliki Forest Virus (SFV) entry into hamster kidney cells, establishing their central importance in cellular infection. 1 The compartments first receiving incoming endocytic material are called "early" or "sorting" endosomes because they represent the primary platform for cargo sorting. Cargo essentially face one of two fates: either they are sorted for entry into lysosomes leading to their degradation, or cargo are retrieved from this fate and promoted for recycling to different cellular organelles 2 ( Figure 1A). Early endosomes undergo a process of maturation into late endosomes by progressively losing cargo via the process of tubular-based recycling and by accumulating changes in luminal pH and the protein and lipid content of their membrane bilayer. [2][3][4] When all the recycled material has been removed the late endosome fuses with the lysosome, leading to the formation of a hybrid organelle named endolysosome in which the remaining contents will eventually be degraded. 5 Cargoes fated for degradation (such as the activated epidermal growth factor receptor, EGFR) are ubiquitinated and sequestered into vesicular structures that invaginate from the endosomal membrane and pinch off in the lumen of the endosomal vacuole to form cargoenriched intraluminal vesicles (ILVs) 6,7 ( Figure 1A). The endosomal sorting complex required for transport (ESCRT), a series of multimeric protein complexes, serves to coordinate recognition of ubiquitinated cargo with the membrane remodelling required for ILV biogenesis. ILV biogenesis is an iterative process leading to late endosomes containing multiple ILVs, hence their alternative name of multivesicular bodies (MVBs). [8][9][10] Late endosomes become competent to fuse with the lysosome, forming a hybrid endolysosome, where the cargo present within the ILVs is degraded through exposure to a series of hydrolyses and lipases present in the lysosomal lumen. [8][9][10] Endolysosomes are subsequently resolved back to lysosomes to regenerate this degradative compartment. 5,11 For an extensive review of the mechanistic regulation of the ESCRT pathway we refer the reader to two excellent reviews. 12,13 Cargo proteins that require retrieval from the lysosomal degradative fate are sorted into branched tubular profiles from where they F I G U R E 1 Role of SNX-BAR proteins in tubular-based endosomal sorting of transmembrane proteins and associated proteins and lipids (cargoes). (A) Cargoes reach the endosomal system from either the biosynthetic pathway or from the plasma membrane through endocytosis. Within endosomes, cargoes can either be fated for degradation (such as the epidermal growth factor receptor [EGFR]) by the endosomal sorting complex required for transport (ESCRT). Alternatively, cargoes can be retrieved from this fate, and recycled through their sorting into endosomal tubular profiles, from where they are clustered in tubulovesicular transport carriers. These transport carriers can either deliver cargoes to the cell surface (e.g., the Retromer-dependent Glucose transporter 1 [GLUT1], and the Retromer-independent Transferrin receptor [TfR]) or to the trans-Golgi network (TGN) (e.g., cation-independent mannose 6-phosphate receptor [CI-MPR]). The family of endosome-associated SNX-BARs are among the most important orchestrators of tubular-based recycling from endosomes. (B) The endosomal SNX-BAR assemblies that participate in tubular-based recycling include: ESCPE-1 consisting of heterodimers of SNX1 or SNX2 with SNX5 or SNX6 (or the neuronal SNX32), heterodimers of SNX4 with SNX7 or SNX30, and SNX8 homodimers. Examples of transmembrane proteins that recycle through these different complexes are depicted in red. (C) Confocal microscopy micrograph of endosomes (in red) from where SNX1-positive tubular profiles (green) emanate to mediate cargo recycling to the trans-Golgi Network (TGN) or to the plasma membrane (PM). (D) Morphology of the cargo-enriched SNX1 intermediates examined by threedimensional electron tomography. Part figure D has been adapted with permission from the Traffic article. 165 are packed into tubulo-vesicular carriers that recycle them to the relevant compartment 2,14 ( Figure 1A). Three main recycling pathways have been described for prototypical cell surface receptors, these include: direct recycling from endosomes to the cell surface ("fast recycling"); transport from sorting endosomes to the perinuclear endocytic recycling compartment (ERC) and then to the cell surface, which can be followed by integrins ("slow recycling"); and transport from sorting endosomes to the trans-Golgi network (TGN) ("retrograde transport"), the route followed by cation-independent mannose 6-phosphate receptor (CI-MPR). 14 Other cargoes, such as the transferrin receptor (TfR), can use multiple routes to return back to the cell surface depending on several factors, including the kinetics of ligand dissociation, through either the fast route or a slower trafficking through the ERC first. 15,16 These different recycling pathways are orchestrated by several multiprotein complexes that associate on the endosomal membrane to coordinate the process of cargo selection with the biogenesis of tubule-vesicular carriers for cargo transport to the target compartment. These complexes included Retromer, Retriever, the CCDC22/ CCDC93 and COMMD (CCC) complex, the Arp2/3-activating WASH complex, and ESCPE-1. While most of the aforementioned sorting complexes have been widely reviewed, 14,17-20 the importance of ESCPE-1 and other related endosomal ESCPE complexes has only recently been reappraised and this will be the principal focus of this review.

| Overview of SNX-BAR proteins
Sorting nexins (SNXs) are a family of peripheral proteins that localize to endosomal membranes to regulate intracellular trafficking of cargo proteins through a combination of lipid-binding and protein-protein interactions. 21 SNXs are all defined by the presence of the phox (PX) domain that binds to phosphoinositide (PI) lipids found in organelle membranes. [21][22][23][24][25] A subset of SNXs contain a BAR (Bin/Amphiphysin/Rvs) domain in their carboxy-terminal region and are therefore named SNX-BARs. 21,24,26 SNX-BARs cycle between the cytosol and organelles of the endosomal network through a common mechanism of membrane association that involves simultaneous detection of several membrane properties via their PX and BAR domains, including curvature, lipid identity and cargo density. 24,[26][27][28] Importantly, a discrete series of hydrophobic and charged interactions in the BAR domain dimer interface ensures a limited scheme of homoand hetero-dimerization, allowing the formation of a restricted number of functional SNX-BAR dimers. [29][30][31] These include the plasma membrane (PM)-associated homodimers of SNX9, SNX18, and SNX33; and the endosome-localized homodimers of SNX8, heterodimers of SNX4:SNX7 and SNX4:SNX30, and heterodimers of SNX1/ SNX2 with SNX5/SNX6/SNX32 30 ( Figure 1B). SNX1/SNX2 and SNX5/SNX6/SNX32 associate in different dimeric combinations to comprise the endosome-associated ESCPE-1 complex, responsible for tubular-based endosome-to-TGN or endosome-to-PM recycling of a myriad of cargoes. ESCPE-1 is the best characterized, and hence prototypical, of the endosomal SNX-BAR complexes. SNX4 heterodimers and SNX8 homodimers are evolutionarily conserved complexes that localize on endosomes where they generate tubular profiles that are distinct from those of ESCPE-1 32-34 ( Figure 1B). In mammalian cells, SNX4 is involved in the recycling of TfR to the plasma membrane, and in autophagic membrane trafficking 32,35-37 while SNX8 has a largely uncharacterised function ( Figure 1B).
2 | MOLECULAR ASSEMBLY AND FUNCTION OF ESCPE-1 SNX1 and SNX2 are paralogous proteins with redundant functions, and the same is also true for SNX5 and SNX6 (and its neuronal variant SNX32). 38-42 SNX1/2 and SNX5/6 are homologs of the yeast Vps5 and Vps17, respectively, that form integral parts of the pentameric retromer complex in fungi. 20,39,40,43,44 In mammals, these SNX-BARs can exist as homodimeric combinations of SNX1 or SNX2, or heterodimeric combinations of SNX1 or SNX2 with either SNX5 or SNX6. 30,42,45 The SNX1/SNX2:SNX5/SNX6 heterodimers have been historically linked with the mammalian subunits of the Retromer complex (VPS26A [or VPS26B]/VPS35/VPS29), and are often referred as to Retromer-linked or Retromer-related SNX-BARs. 14,46 However, they have also been shown to coordinate the process of membrane remodelling and cargo sorting in a Retromer-independent fashion and the complex has been named the "Endosomal SNX-BAR Sorting Complex for Promoting Exit," or ESCPE-1 42 with cargo adaptors such as SNX27. 52 The direct association between SNX27 and the unstructured N-terminus of SNX1/SNX2 allows for Retromer cargoes (e.g., Glucose transporter 1, GLUT1 and β2-Adrenergic receptor, β2AR) to exit endosomes through ESCPE-1-positive tubular profiles ( Figure 1A). Accordingly, suppression of the ESCPE-1 subunits leads to the accumulation of Retromer cargoes such as GLUT1 in early endosomes, resulting in reduced levels on the plasma membrane. 52,53,56,57 Importantly, suppression of ESCPE-1 does not perturb the recycling of cargoes linked to other sorting complexes such as Retriever and the CCC complex, suggesting a specificity of the sorting of Retromer cargoes into ESCPE-1 tubules. 17 This process of tubular export from endosomes, which we will describe in detail in the following paragraphs, is achieved through the ability of the SNX-BAR dimer to (i) associate to the peripheral endosomal surface, (ii) directly interact with transmembrane proteins, and (iii) locally remodel these membranes into tubulovesicular cargoenriched transport carriers that are transported to the target compartment; together termed "co-incidence detection" 30,48 ( Figure 1C). The generation of cargo-enriched carriers and their trafficking by ESCPE-1 mirrors the general functional principles of other coat complexes: (1) cargo is recognized and concentrated on the donor compartment, (2) cargo-enriched membranes remodel to form vesicular or tubular profiles, (3) scission of these profiles lead to the formation of isolated transport carriers, (4) carriers couple to molecular motors for movement to the target compartment, (5) tethering molecules capture incoming carriers to facilitate SNARE-mediated fusion with the correct acceptor compartment. 58 We will explore in detail how the ESCPE-1 orchestrates each of these steps.

| Endosomal targeting by ESCPE-1
SNX1 and SNX2 are the key membrane-binding components of the ESCPE-1. They both display a steady state localization on early and late endosomes, and such localization does not require the partners SNX5/SNX6. 24,28,41,48 The isolated PX domain of SNX1/SNX2 bind PI(3)P, while full-length proteins have been reported to also bind PI(3,5)P 2 and PI(3,4)P 2 . 24,25,41,49 The BAR domain also facilitates membrane-association through a series of positively charged residues on the concave surface of the dimer that mediate electrostatic interactions with the lipid bilayer 24,59 (Figure 2A). Supporting the importance of the PX and BAR domains of SNX1 and SNX2 in the interaction with membrane, mutation of residues that compromise the PI binding site, as well as mutations that disrupt the BAR domain, result in cytosolic localization of the protein. 24,48 Early studies indicated that SNX5/SNX6/SNX32 displayed phosphoinositide binding activity, 60,61 but a recent screen indicated that the PX domains of these proteins do not display detectable lipid binding capability. 25 Consistent with this, crystal structures of the PX domains of SNX5 and SNX32 have established that they lack key residues required for lipid binding. Rather, it appears that the PX domain of these SNX-BARs is uniquely specialized to bind to the specific sequence motifs present in cytosolic tails of transmembrane cargo. 48,62,63 Full-length SNX5/SNX6/SNX32 rely on heterodimeric assembly with SNX1 or SNX2 to associate with endosomal membranes 30,42,48 (Figure 2A) Accordingly, mutations of the charged residues on the concave face of the SNX5-BAR domain, or mutation of the residues in the dimerization interface result in impaired membrane association. 64,65 F I G U R E 2 Molecular basis for the biogenesis of ESCPE-1 transport carriers. (A) Model of the mechanism by which ESCPE-1 heterodimers sense multiple features of endosomal membranes, including the presence of specific phosphoinositides, local membrane curvature and cytosolic tails of transmembrane proteins. By sensing these features, ESCPE-1 assembles into functional membrane-associated complexes that couple membrane remodelling with cargo recognition for the formation of cargo-enrich transport carriers. (B) Molecular details of the association between SNX5 and the cytosolic tail of cargoes such as CI-MPR and SEMA4C possessing a ФxΩxФ(x) n Ф consensus motif that folds into a beta harping structure (Ф = hydrophobic and Ω = aromatic sidechains). The first β-strand always consists of a highly hydrophobic sequence, 2349 VSYKYS 2354 in CI-MPR and 734 VGYYYS 739 in SEMA4C. The key side-chains of SNX5 that mediate binding include Tyr132, Leu133 and Phe136, with the central tyrosine of the βA forming a stacking interaction with SNX5 Phe136. The binding is further sustained by one hydrophobic side chain from βB that packs in the hydrophobic groove of SNX5. In SEMA4C this residue is Leu743, while in CI-MPR both Leu2370 or Met2371 can support the binding, suggesting that rather than a specific residue, any properly positioned hydrophobic residue in the βB could sustain the interaction. 48 (C) Model of the oligomeric assembly of SNX-BARs into tubular lattices that coordinate the biogenesis of cargo-ladened transport carriers.

| ESCPE-1 directly binds cargoes
Several SNXs act as cargo adaptors that directly bind transmembrane proteins to regulate, alone or in concert with multimeric complexes such as Retromer and Retriever, cargo trafficking through the endosomal network. 14 Among these, SNX27 and SNX17 mediate the endosome-to-plasma membrane recycling of a myriad of cargoes by binding PDZ binding motifs and NPxY/NxxY sorting motifs, respectively, 52,66-71 recently reviewed in Ref. [63]. All components of ESCPE-1 were initially characterized for their abilities to bind a variety of receptors including the epidermal growth factor receptor (EGFR), platelet-derived growth factor receptor (PDGFR), insulin receptor (INSR), leptin receptor and several members of the transforming growth factor (TGF)-β receptor family 22,23,[72][73][74] (Table 1). More recently, the proteomic interrogation of SNX-BAR immunoprecipitates and unbiased labelling of the plasma membrane and TGN have vastly expanded the cohort of receptors that were found to associate with SNX5/SNX6. 42,47,48,51,75 Furthermore, the use of recombinant binding assays such as isothermal titration calorimetry (ITC) has revealed the direct nature of some of these interactions. Among the cargoes found to directly interact with SNX5/SNX6 are the CI-MPR, the insulin-like growth factor-1 receptor (IGF1R), semaphorin 4C (SEMA4C) and neuropilin-1 (NRP1) 46,48,51 (Table 1).
Importantly, structural analysis of the minimal CI-MPR and SEMA4C peptides bound to the PX domain of SNX5 revealed the molecular basis for these interactions 48 ( Figure 2B). CI-MPR and SEMA4C bind an evolutionarily conserved extension that is unique to the PX domain of SNX5/SNX6 (this extension is absent in fungal Vps17). 48 Cargo interaction with the SNX5 PX domain occurs via a bipartite structure, consisting of two antiparallel β-strands (βA and βB) connected by a variable region ( Figure 2B). 48 The βA of the aforementioned cargoes conforms to a ФxΩxФ consensus T A B L E 1 Summary of known ESCPE-1 cargoes and pathogens reported to hijack them as host factors for infection of human cells. Upon localisation to membranes, it is thought that the increased concentration of SNX-BARs serves to promote their oligomerisation into loosely ordered spiral arrays, which result in vesicle-to-tubule membrane remodeling 30,76,77 ( Figure 2D). In vitro liposome-based membrane remodeling assays revealed that both purified SNX1 and SNX2 are able to induce the formation of tubules in a concentrationdependent fashion, while SNX5 and SNX6 fail to do so. 24,30,77 Also, overexpression of SNX1 and SNX2 in HeLa cells induces the formation of extended tubules emanating from endosomes. 24,41,42 The transition between the local membrane deformation induced by SNX-BAR dimers to the formation of tubular profiles is a cooperative process that involves three mechanisms: (1) adherence of the curved BAR-domain dimer onto the membrane via co-incidence detection of membrane features; (2) the insertion of shallow amphipathic helices (AHs) that disrupts local lipid organization imposing further membrane curvature and (3) tip-to-tip and side-to-side interactions between dimers that organize the formation of an oligomeric tubular lattice that translates the local membrane-sculpting properties of the single dimer into a global tubular remodeling of the membrane. 30,77 Interestingly, molecular simulation has estimated that a 30%-40% coverage of BAR proteins on the membrane is sufficient to form a continuous coat that shapes and stabilizes these tubular profiles 78,79 ( Figure 2D).
Cryoelectron tomography has provided structural validation of the tenets of SNX-BAR domain binding. In the structure of homodimeric membrane-bound SNX1, electrostatic associations and insertion of an AH facilitate membrane binding and curvature. 77,80 Moreover, the PX domains of several SNXs contain a membrane insertion loop that further promotes membrane deformation. 81 Tip-to-tip interactions promote the association between adjacent SNX-BAR dimers. 30 Figure 2B,C).

| Elongation and fission of ESCPE-1 tubular carriers
Following cargo enrichment, carrier scission is mediated by a complex symphony of biochemical and biophysical forces imposing tension on the tubule. The WASH complex has a primary role in facilitating the extension of the ESCPE-1 tubular profiles and in regulating their fission through the activation of a localized Arp2/3-mediated actin nucleation. 19,82,83 The WASH complex is recruited to ESCPE-1 tubular domains via the association with the SNX1 interactor RME-8 thereby coordinating its activity with the membrane remodeling ability of the SNX-BARs. 84 The WASH-dependent formation of branched filamentous actin may also contribute to the fission of the tubular profiles by providing a pushing force to induce membrane tension 19,85 ( Figure 3A). Accordingly, depletion of RME-8 or the WASH complex gives rise to long membrane tubules extending from endosomes and prevents the correct retrograde trafficking of the ESCPE-1 cargo CI-MPR. [82][83][84]86 More recently, it has been shown that tubular profiles with a BAR-protein coat can undergo spontaneous friction-driven scission. 87 A biophysical model suggests that the elongation force of molecular motors pulling on a coated tubular membrane causes tension between the lipids, leading to pore nucleation and tube scission. 87 Conceptually  ( Figure 3B).
Following engagement of incoming membrane carriers, the precise mechanism that brings them into direct proximity of the TGN membrane is unknown but may be mediated by the Golgin "collaps- ing" at hinge regions where coiled-coils are disrupted, or the "hopping" of vesicles between Rab-binding sites on the Golgin coils to bring them into closer proximity to the membranes. 98 108 and the TGN-enriched PI(4)P that facilitate the release of the ESCPE-1 coat 61 ( Figure 3C).
As ESCPE-1 also mediates endosome-to-plasma membrane recycling, coupling to a plus-end directed microtubule motor complex(es) is likely to be required for carrier transport toward the cell periphery.
However, at the present time the motor protein responsible for the plasma membrane-directed transport has not been identified. It has been shown that kinesin-1 is required for the motility of SNX1-positive endosomes and the organization of SNX1 subdomains, 89 whether the same motor complex also contributes to the peripheral transport of the ESCPE-1-decorated cargo-enriched carriers remains to be investigated.

| ESCPE-1 IN HOST-PATHOGEN INTERACTION
The catabolic endosomal and autophagic clearance pathways represent an innate cellular defense against invading pathogens. Accordingly, many intracellular pathogens subvert host endosomal sorting pathways to circumvent lysosomal degradation and promote survival and proliferation within cells. 109 For example, various bacterial pathogens exploit endosomal-associated machineries to promote the generation of permissive replication niches. 110 Moreover, many viruses are internalized into the cells through endocytic pathways after binding of host receptors, then rely on endosomal maturation and acidification to trigger their uncoating and genome release, thereby preventing exposure of the genome to nuclease enzymes present in the mature lysosome. 111 Endosome-associated machineries such as Retromer, Retriever and the CCC and WASH complexes are key regulators of endosomal biology and hence it does not surprise that they are often subverted by infecting agents for their advantage. 112-114 ESCPE-1 is no exception and recent work has reappraised the importance of this coat complex in the tug-of-war between host and pathogens.  Figure  4A).

S. Typhimurium invades cells, and resides within the SCV, which
matures in a distinct pathway to that of the endo-lysosomal network. 116 During the early stages of infection, S. Typhimurium releases its effector SigD/SopB into the cytosol, an inositol polyphosphatase that hydrolyses a variety of PIs and promotes the recruitment of Rab5 and the PI3-kinase VPS34, overall contributing to the increase of PI (3) P on the SCV 116,117 ( Figure 4A). The manipulation of PI levels drives the enrichment of SNX-BAR proteins on to the SCV membrane and triggers the formation of vacuole-associated tubules that allow SCV maturation. 115,116 The exaggerated ESCPE-1 and SNX-BAR-driven tubulation leads to a faster condensation of the pathogen-containing membrane that shrinks toward the bacteria until it eventually adheres to the pathogen, forming the SCV 115,116 ( Figure 4A).
The Q fever-causing bacterium Coxiella burnetii requires both Retromer and the ESCPE-1 to progress in its replication. 118 Furthermore, Listeria monocytogenes, the causative agent of listeriosis, secretes the virulence factor Lmo1656 that binds SNX6 to subvert its function during early stages of oral listeriosis. 119 The ESCPE-1 coat complex is also used by toxins en route to different subcellular compartments, including Shiga toxin B-subunit which requires ESCPE-1 for efficient endosome-to-TGN transport. 120-122

| ESCPE-1 and antiviral xenophagy
On the other hand, SNX5 also plays a role in cellular immunity against some viruses, thereby protecting cells and organisms from viral infections. 65,138 Using a genome-wide short interfering RNA screen, SNX5 and SNX32 were identified as essential factors promoting virusinduced, but not basal or stress-induced, autophagy (specifically called xenophagy). 65  (CVB3) and Influenza A virus (IAV). 65 Consistently, it was observed that Snx5 knockout mice displayed higher lethality after infection with several of these human viruses (SINV, WNV, HSV-1 ΔBBD and CHIKV). 65 The mechanism by which SNX5 facilitates xenophagy depends on its ability to interact with Beclin-1 and ATG14-containing class III phosphatidylinotol-3-kinase (PI3KC3) complex 1 (PI3KC3-C1). 65 This interaction promotes the generation of PI(3)P on virus-containing endosomes and drives the recruitment of the PI(3)P-binding protein WIPI2 that initiate the autophagosome assembly 65 ( Figure 5A). An outstanding question relates to how luminal viruses stimulate the ESCPE-1-PI3KC3 axis on the cytoplasmic face of endosomes? Presently, we can only speculate on likely mechanisms. SNX5 associates with integral receptor tyrosine kinases that serve as viral entry factors, 139 such as AXL (e.g., ZIKV), IGF1 receptor (e.g., HRSV), EPHA2 (e.g., Epstein-Barr virus, EBV), MET (e.g., adeno-associated viruses) and EGFR (e.g., IAV) 42,48,140,141 (Table 1). Viral-induced enrichment of these membrane proteins may provide a compartment "signature" for recruiting the SNX5-PI3KC3 axis to virus-containing endosomes ( Figure 5A).
Additional mechanisms, such as viral induction of local membrane curvature (sensed by the SNX5 BAR domain) and/or virally encoded membrane penetrating peptides that present ESCPE-1 binding motifs to the cytoplasm (as in the case of the human papillomavirus, HPV) may serve to add complexity to the recruitment signature of viral containing endosomes. 142 It also remains to be addressed whether the SNX1 and SNX2 subunits of ESCPE-1 contribute to this these processes.

| ESCPE-1 and SARS-CoV-2
In     have also been shown to use NRP1 for entry and infection. 158,159 It is, therefore, tempting to speculate that the role of ESCPE-1 in SARS-CoV-2 infection could be extended to other NRP1-dependent viruses.
Given that ESCPE-1 also regulates the sorting of a wider array of viral receptors, including AXL, IGF1R, EPHA2, MET, EGFR, this effect may also apply to multiple distinct microbial pathogens that rely on these receptors for infection (Table 1). Future studies aiming to modulate or inhibit this pathogenic subversion of ESCPE-1 cargo sorting, or SNX5-driven initiation of xenophagy, may provide insights into how to manipulate this pathway to block intracellular pathogen survival.

| Other ESCPE complexes
Interestingly, recent work seems to suggest that endosomal-SNX-BARs can assemble into dimeric sorting complexes outside of the conventional dimerization pairing. This is the case for the Recycler complex, composed of SNX4:SNX5 heterodimers and SNX17, that plays an essential role in autophagosomal components recycling. 160 With limited evidence for a functional role of SNX4:SNX7 and SNX4: SNX30 heterodimers, and SNX8 homodimers in direct sorting of endosomal cargoes, future work will be required to dissect the molecular basis for how these heterodimeric SNX-BAR assemblies participate to the process of cargo recycling, and how this contributes to cellular pathways for host-pathogen interaction. SNX4 and SNX8 have been linked to the retrograde trafficking of the ricin toxin and Shiga toxin subunit B (STxB) toxins respectively, although mechanistic details of endogenous cargo recycling through this route remain unclear. 122,161 Interestingly, SNX8 has been reported to be part of a host defense mechanism against Listeria monocytogenes and a regulator of innate cellular responses to viruses. [162][163][164] Clearly, there is still a lot to learn about the cellular functions of endosomal SNX-BAR proteins, and how their role is subverted by pathogens for infection.