Biology subjects

Collins, B. M.

Publications and source records attributed to Collins, B. M..

At least 19 recordsLinked to original sources

bioRxiv2026

Global identification of neuronal and astrocytic integral membrane proteins that require Retromer for their endosomal recycling.

Efficient transport of membrane proteins through the endosomal network is essential for brain development and function, with perturbation implicated in disease. Deficiencies in Retromer, a key regulator of endosomal transport, have been linked to aging-related neurodegenerative disorders including Alzheimers and Parkinsons disease. To better define the neuroprotective role of Retromer, we have applied cell surface restricted proteomics to identify those integral membrane proteins whose recycling to the plasma membrane is mediated by Retromer and associated cargo adaptors, sorting nexin 3 (SNX3), its paralogue sorting nexin 12 (SNX12), and sorting nexin 27 (SNX27) (data available via ProteomeXchange: PXD078277). By comparing primary rat cortical neurons and astrocytes we have identified several cargoes that require either SNX3/SNX12- or SNX27-Retromer complexes for endosomal recycling, including proteins involved in synapse organisation, synaptic signalling and Alzheimers disease pathology. We highlight that perturbed Retromer function leads to endosomal enlargement, and we establish a key role of SNX27-Retromer in modulating transport of glutamate across both neuronal and astrocytic membranes via recycling of glutamate transporters EAAT3 (SLC1A1) and EAAT1 (SLC1A3) respectively. Our study provides further mechanistic insight into the consequences of Retromer deficiency for neuronal and astrocytic function, offering new avenues of research in the treatment of neurodegenerative disease. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=194 SRC="FIGDIR/small/724903v1_ufig1.gif" ALT="Figure 1"> View larger version (59K): org.highwire.dtl.DTLVardef@1225eedorg.highwire.dtl.DTLVardef@1863f1dorg.highwire.dtl.DTLVardef@99c21eorg.highwire.dtl.DTLVardef@976bfd_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical AbstractC_FLOATNO Suppression of Retromer and the sorting nexins (SNX27, SNX3/SNX12) leads to a significant change in the surface proteome of rat cortical neurons and astrocytes. Focusing on the glutamate transporters, SLC1A1 and SLC1A3, we have validated that SNX27-Retromer is required for their trafficking, with SNX27-Retromer suppression in astrocytes leading to a loss of glutamate uptake. C_FIG

molecular biology↗

bioRxiv2026

A remarkable case of conserved domain swapping in the COMMD family of proteins

The eukaryotic COMMD family of proteins are core subunits of the Commander protein complex, with a central role in endosomal membrane trafficking and signalling. Previous crystal and cryoEM structures show that COMMD and COMMD-like proteins form homo-oligomeric and hetero-decameric assemblies with a central ring structure formed by their small C-terminal COMM domains. Their -helical N-terminal (HN) domains decorate each side of these rings, and in the eukaryotic Commander complex engage the coiled-coil proteins CCDC22 and CCDC93. Here, we have determined new crystal structures of the isolated HN domains of human Commd4, Commd9 and Commd10, and find that all three proteins form domain swapped structures with a remarkably consistent topology. This occurs via a conformational change in a hinge-loop between helices 2 and 3, leading to exchange of helices 3 to 6 between protomers. The hinge-loops of Commd9 and Commd10 possess several serine and threonine residues that can be phosphorylated, and we find that specific phospho-mimicking mutations in Commd10 can promote or inhibit domain swapping. Whether the unique COMMD HN domains play any roles beyond assembly with CCDC proteins is unclear, but this work suggests a common conformational switch exists with a potential to regulate their function.

biochemistry↗

bioRxiv2025

Redox Regulation of Cell Migration via Nischarin S-glutathionylation

Reactive oxygen species (ROS) are central players in redox signaling, controlling all biological processes in human health. Many reports demonstrated that ROS play essential roles in regulating cell migration and invasion, while contributing to cancer progression and metastasis, potentially via inducing protein cysteine oxidations. Nevertheless, specific redox players involved in cell migration and invasion remain ill-defined. In this report, we found that Nischarin (NISCH), established as a tumor suppressor, is susceptible to S-glutathionylation, selectively at Cys185 located near its leucine-rich repeat (LRR) domains, which are implicated in protein-protein interactions with Rac1 and PAK1. We demonstrated that epithelial breast cancer cell lines, MCF7 and MDA-MB-231, expressing NISCH wild-type (WT), compared to its cysteine mutant (C185S), exhibit increased migration and invasion in response to oxidative stress, such as limited glucose. Mechanistically, NISCH S-glutathionylation reduced its binding to Rac1 and PAK1, without altering its binding to integrin 5. The dissociation of NISCH led to the activation of Rac1 and PAK1, resulting in the localization of Rac1 to the cell periphery, which facilitates lamellipodia formation. The activated PAK1 increased the phosphorylation of the LIMK1-cofilin axis, thereby further enhancing actin filament dynamics that promote cell migration. Based on the mechanistic analysis, we produced an engineered NISCH construct, composed of the N-terminal PX and LRR domains. We demonstrated that the engineered NISCH PX-LRR constructs, particularly one lacking the S-glutathionylation site (i.e., C185S), can suppress the migration, invasion, and colony formation of MDA-MB-231 cells, regardless of the presence of oxidative stress. Our data reports a new redox player in cell migration and invasion, while supporting the potential application of NISCH-derived protein-based therapeutics for breast cancer.

biochemistry↗

bioRxiv2025

The prokaryotic origins of the COMMD protein family involved in eukaryotic membrane trafficking

The ten eukaryotic COMMD proteins are core components of the Commander complex, with central roles in endosomal membrane trafficking and signalling. Each protein has an -helical N-terminal (HN) domain with a C-terminal copper metabolism gene MURR1 (COMM) domain. These ten family members assemble into a heterodecameric ring composed of five specific heterodimers. In this work we have combined structural homology searches with genome-wide predicted structures to identify ancestral COMMD-like proteins that exist as single genes in Bacteria and Archaea. Although there is limited sequence similarity to the eukaryotic proteins the bacterial and archaeal COMMD-like proteins are predicted to form homomeric ring-shaped assemblies like their eukaryotic counterparts. Our biophysical studies, crystal and cryo-EM structures confirm COMMD-like proteins readily form homooligomeric rings composed of eight or ten subunits assembled from core dimeric building blocks and inter-dimer interactions that are analogous to the heterodecameric core structure of the eukaryotic Commander complex. Phylogenetic analyses using amino acid sequences and FoldSeek structural alphabet (3Di) infer that the closest identified relatives to the eukaryotic COMMD proteins are found in Myxococcota bacteria. These findings indicate that COMMD genes emerged early in eukaryotic evolution through multiple rounds of duplication from a single ancestral gene likely acquired from bacteria.

biochemistry↗

bioRxiv2025

Identification of a VPS29 isoform with restricted association to Retriever and Retromer accessory proteins through auto-inhibition.

The endosomal-lysosomal network is a hub of organelles that orchestrate the dynamic sorting of hundreds of integral membrane proteins to maintain cellular homeostasis. VPS29 is a central conductor of this network through its assembly into Retromer, Retriever and Commander endosomal sorting complexes, and its role in regulating RAB GTPase activity. Two VPS29 isoforms have been described, VPS29A and VPS29B, that differ solely in their amino-terminal sequences. Here we identify a third VPS29 isoform, which we term VPS29C, that harbours an extended amino-terminal sequence compared to VPS29A and VPS29B. Through a combination of AlphaFold predictive modelling, in vitro complex reconstitution, mass spectrometry and molecular cell biology, we find that the amino-terminal VPS29C extension constitutes an autoinhibitory sequence that limits access to a hydrophobic groove necessary for effector protein recruitment to Retromer, and association with Retriever and Commander. VPS29C is therefore unique in its ability to uncouple Retromer-dependent cargo sorting from the broader roles of VPS29A and VPS29B in regulating the endosomal-lysosomal network through accessory protein recruitment. Our identification and characterisation of VPS29C points to additional complexity in the differential subunit assembly of Retromer, an important consideration given the increasing interest in Retromer as a potential therapeutic target in neurodegenerative diseases. SIGNIFICANCE STATEMENTThe endosomal-lysosomal network is essential for normal cellular function with network defects being associated with numerous neurodegenerative diseases. Two heterotrimeric complexes, Retromer and Retriever, control transmembrane protein recycling through the network. Of these, reduced Retromer expression is observed in Alzheimers disease and Retromer mutations lead to familial Parkinsons disease. Here, we identify and characterise a new isoform of VPS29, a subunit shared between Retromer and Retriever. We reveal how this isoform, VPS29C, adopts an auto-inhibitory conformation to limit its association into Retriever and restrict the binding of VPS29C-containing Retromer to accessory proteins vital for regulating network function. By revealing added complexity in Retromer assembly and function, we provide new insight into Retromers potential as a therapeutic target in neurodegenerative diseases.

cell biology↗

bioRxiv2025

Identification of a RAB32-LRMDA-Commander membrane trafficking complex reveals the molecular mechanism of human oculocutaneous albinism type 7

Commander is an endosome associated sixteen protein assembly that associates with the sorting nexin 17 (SNX17) cargo adaptor to regulate cell surface recycling of internalised integral membrane proteins including integrins and lipoprotein receptors. Mutations in Commander are causative for Ritscher-Schinzel syndrome (RSS), a multiorgan developmental disease associated with a core triad of cerebellar-cardiac-craniofacial malformation. Here, using unbiased proteomics and computational modelling, we identify leucine rich melanocyte differentiation associated (LRMDA) as a novel Commander binding protein. Using recombinant protein reconstitution, we show that LRMDA simultaneously associates with Commander and active RAB32, and, by revealing that LRMDA and SNX17 share a common mechanism of Commander association, establish the mutually exclusive nature of RAB32-LRMDA-Commander and SNX17-Commander assemblies. From functional analysis in human melanocytes, we establish an essential role for RAB32-LRMDA-Commander in melanosome biogenesis and pigmentation and reveal a distinct functional role for SNX17-Commander in this organelle biogenesis pathway. We reveal how LRMDA mutations, causative for oculocutaneous albinism type 7 (OCA7), a hypopigmentation disorder accompanied by poor visual acuity, uncouple RAB32 and Commander binding thereby establishing the mechanistic basis of this disease. Our discovery and characterisation of this alternative Commander assembly establishes an unrecognised plasticity of Commander function within a highly complex organelle biogenesis pathway. This extends Commander function beyond the confines of SNX17-mediated cell surface recycling into RAB32-family mediated biogenesis of lysosome-related organelles and, potentially, other RAB32 regulated pathways including host-pathogen defence mechanisms. Our work also extends the breath of Commander pathway dysfunction for human disease.

cell biology↗

bioRxiv2024

Cavin1 binding nanobodies reveal structural flexibility and regulated interactions of the N-terminal coiled-coil domain

Caveolae are abundant plasma membrane structures that regulate signalling, membrane homeostasis, and mechanoprotection. Their formation is driven by caveolins and cavins and their coordinated interactions with lipids. We have developed nanobodies against the trimeric HR1 coiled-coil domain of Cavin1. We identify specific nanobodies that do not perturb Cavin1 membrane binding and localise to caveolae when expressed in cells. The crystal structure of a nanobody HR1 complex reveals a symmetric 3:3 architecture as validated by mutagenesis. In this structure, the C-terminal half of the HR1 domain is disordered suggesting the nanobody has stabilised an open conformation previously identified as important for membrane interactions. A phosphomimic mutation in a Thr-Ser pair proximal to this region reveals selective regulation of Cavin2/Cavin3 association. These studies provide new insights into Cavin domains required for assembly of multiprotein caveolar assemblies and describe new nanobody tools for structural and functional studies of caveolae. SUMMARY STATEMENTNanobodies are reported that can label the Cavin1 protein and caveola structures in cells.

cell biology↗

bioRxiv2024

Molecular basis for the assembly of the Vps5-Vps17 SNX-BAR proteins with Retromer

Retromer mediates endosomal retrieval of transmembrane proteins in all eukaryotes and was first discovered in yeast in complex with the Vps5 and Vps17 sorting nexins (SNXs). Cryoelectron tomography (cryoET) studies of Retromer-Vps5 revealed a pseudo-helical coat on membrane tubules where dimers of the Vps26 subunit bind Vps5 membrane-proximal domains. However, the Vps29 subunit is also required for Vps5-Vps17 association despite being far from the membrane. Here, we show that Vps5 binds both Vps29 and Vps35 subunits through its unstructured N-terminal domain. A Pro-Leu (PL) motif in Vps5 binds Vps29 and is required for association with Retromer on membrane tubules in vitro, and for the proper recycling of the Vps10 cargo in Saccharomyces cerevisiae. CryoET of Retromer tubules with Vps5-Vps17 heterodimers show a similar architecture to the coat with Vps5-Vps5 homodimers, however, the spatial relationship between Retromer units is highly restricted, likely due to more limited orientations for docking. These results provide new mechanistic insights into how Retromer and SNX-BAR association has evolved across species.

cell biology↗

bioRxiv2024

PPTC7 antagonizes mitophagy by promoting BNIP3 and NIX degradation via SCFFBXL4

Mitophagy must be carefully regulated to ensure that cells maintain appropriate numbers of functional mitochondria. The SCFFBXL4 ubiquitin ligase complex suppresses mitophagy by controlling the degradation of BNIP3 and NIX mitophagy receptors, and FBXL4 mutations result in mitochondrial disease as a consequence of elevated mitophagy. Here, we reveal that the mitochondrial phosphatase PPTC7 is an essential cofactor for SCFFBXL4-mediated destruction of BNIP3 and NIX, suppressing both basal and induced mitophagy. Disruption of the phosphatase activity of PPTC7 is not required for BNIP3 and NIX turnover. Rather, a pool of PPTC7 on the mitochondrial outer membrane acts as an adaptor linking BNIP3 and NIX to FBXL4, facilitating the turnover of these mitophagy receptors. PPTC7 accumulates on the outer mitochondrial membrane in response to mitophagy induction or the absence of FBXL4, suggesting a homeostatic feedback mechanism that attenuates high levels of mitophagy. We mapped critical residues required for PPTC7-NIX/BNIP3 and PPTC7-FBXL4 interactions and their disruption interferes with both NIX/BNIP3 degradation and mitophagy suppression. Collectively, these findings delineate a complex regulatory mechanism that restricts NIX/BNIP3-induced mitophagy.

cell biology↗

bioRxiv2024

Mechanism and regulation of cargo entry into the Commander recycling pathway

Commander is a multiprotein complex that orchestrates endosomal recycling of diverse integral cargo proteins and in humans is required for normal skeletal, brain, kidney, and cardiovascular development. While the structure of this complex has recently been described, the central question of how cargo proteins are selected for entry into the Commander recycling pathway remains unclear. Here using recombinant protein reconstitution and in silico predictions we identify the evolutionary conserved mechanism through which the unstructured carboxy-terminal tail of the integral protein adaptor sorting nexin-17 (SNX17) directly binds to the Retriever sub-complex of Commander. SNX17 adopts an autoinhibited conformation where its carboxy-terminal tail occupies the cargo binding groove. Competitive cargo binding overcomes this autoinhibition, promoting SNX17 endosomal residency and the release of the carboxy tail for Retriever association. Using molecular cell biology and high-resolution microscopy, we establish the central importance of SNX17-Retriever association in the handover of integrin and lipoprotein receptor cargoes into pre-existing endosomal retrieval sub-domains for entry into the recycling pathway. In describing the principal mechanism of cargo entry into the Commander recycling pathway we provide key insight into the function and regulation of this evolutionary conserved sorting complex.

cell biology↗

bioRxiv2023

Ggnbp2 regulates synaptic development and autophagy in motor neurons

AbstractGenome-wide association studies (GWAS) have identified numerous candidate ALS risk variants, but their cellular functions are often unknown. Recent studies have identified a variant of GGNBP2 that results in increased expression. To better understand how this gene might contribute to disease, we investigated the function of Drosophila Ggnbp2 (dGgnbp2) in motor neurons. Loss of function studies showed that dGgnbp2 is required for motor neuron synaptic development. A human transgene completely rescued these phenotypes indicating that the gene is functionally conserved between humans and flies. Overexpression of dGgnbp2 caused severe locomotor defects in adult flies, consistent with ALS pathology. At the cellular level, dGgnbp2 regulated autophagy, a process commonly defective in ALS. Both overexpression and removal of dGgnbp2 reduced levels of the phosphorylated lipid, PI(3)P, an essential component of autophagosomes. Our study provides strong evidence that Ggnbp2 functions in motor neurons to regulate a cellular process commonly defective in ALS. TeaserThis study investigated the function of the ALS risk variant GGNBP2, in flies, and showed that it regulates autophagy in motor neurons.

neuroscience↗

bioRxiv2023

Polarity-dependent expression and localization of secretory glucoamylase mRNA in filamentous fungal cells

In multinuclear and multicellular filamentous fungi little is known about how mRNAs encoding secreted enzymes are transcribed and localized spatiotemporally. To better understand this process we visualized mRNA encoding GlaA, a glucoamylase secreted in large amounts by the industrial filamentous fungus Aspergillus oryzae, in living cells by the MS2 system. We found that glaA mRNA was significantly transcribed and localized near the hyphal tip and septum, which are the sites of protein secretion. We also revealed that glaA mRNA exhibits long-range dynamics in the vicinity of the endoplasmic reticulum (ER) in a manner that is dependent on the microtubule motor proteins kinesin-1 and kinesin-3, but independent of early endosomes. Moreover, we elucidated that the regulatory mechanisms of glaA mRNA by stress granules and processing bodies were different under high temperature and ER stress. Collectively, this study uncovers a dynamic regulatory mechanism of mRNA encoding a secretory enzyme in filamentous fungi.

cell biology↗

bioRxiv2023

Structural basis for coupling of the WASH subunit FAM21 with the endosomal SNX27-Retromer complex

Endosomal membrane trafficking is mediated by specific protein coats and formation of actin-rich membrane domains. The Retromer complex coordinates with sorting nexin (SNX) cargo adaptors including SNX27, and the SNX27-Retromer assembly interacts with the WASH complex which nucleates actin filaments establishing the endosomal reycling domain. Crystal structures, modelling, biochemical and cellular validation reveal how the FAM21 subunit of WASH interacts with both Retromer and SNX27. FAM21 binds the FERM domain of SNX27 using acidic-Asp-Leu-Phe (aDLF) motifs similar to those found in the SNX1 and SNX2 subunits of the ESCPE-1 complex. Overlapping FAM21 repeats and a specific Pro-Leu containing motif bind three distinct sites on Retromer involving both the VPS35 and VPS29 subunits. Mutation of the major VPS35-binding site does not prevent cargo recycling, however it partially reduces endosomal WASH association indicating that a network of redundant interactions promote endosomal activity of the WASH complex. These studies establish the molecular basis for how SNX27-Retromer is coupled to the WASH complex via overlapping and multiplexed motif-based interactions required for the dynamic assembly of endosomal membrane recycling domains. Significance StatementCell surface transmembrane proteins are regulated by a constant cycle of internalization and recycling from intracellular endosomal compartments. The Retromer protein complex and the sorting nexin adaptor protein play a critical role in the retrieval of hundreds of proteins responsible for ion transport, glucose metabolism, neurotransmission, and other cell functions. We have defined the mechanism by which both Retromer and SNX27 engage the actin-nucleating complex called WASH through multiple repeated sequences in the subunit FAM21. Dysfunction in WASH, Retromer and SNX27 are implicated in various disorders including Parkinsons disease, Alzheimers disease, hereditary spastic paraplegia, and this work provides important insights into the assembly of these essential endosomal sorting machineries.

cell biology↗

bioRxiv2023

Copine-6 is a Ca2+ sensor for activity-induced AMPA receptor exocytosis

The recruitment of synaptic AMPA (-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid) receptors underlies the strengthening of neuronal connectivity during learning and memory. This process is triggered by NMDA (N-methyl-D-aspartate) receptor-dependent postsynaptic Ca2+ influx. Synaptotagmin (Syt)-1 and -7 have been proposed as Ca2+-sensors for AMPA receptor exocytosis, but are functionally redundant. Here we identify a cytosolic C2 domain-containing Ca2+-binding protein Copine-6 that forms a complex with AMPA receptors. Loss of Copine-6 expression impairs activity-induced exocytosis of AMPA receptors in primary hippocampal neurons, which is rescued by wild-type Copine-6, but not Ca2+-binding mutants. In contrast, Copine-6 loss-of-function has no effects on steady-state expression or tetrodotoxin-induced synaptic upscaling of surface AMPA receptors. Loss of Syt-1/-7 significantly reduces Copine-6 protein expression. Interestingly, overexpression of wild-type Copine-6, but not the Ca2+-binding mutant, restores activity-dependent exocytosis of AMPA receptors in Syt-1/-7 double-knockdown neurons. We conclude that Copine-6 is a postsynaptic Ca2+-sensor that mediates AMPA receptor exocytosis during synaptic potentiation.

neuroscience↗

bioRxiv2023

Caveola mechanotransduction reinforces the cortical cytoskeleton to promote epithelial resistance

As physical barriers, epithelia must preserve their integrity when challenged by mechanical stresses. Cell-cell junctions linked to the cortical cytoskeleton play key roles in this process, often with mechanotransduction mechanisms that reinforce tissues. Caveolae are mechanosensitive organelles that buffer tension via disassembly. Loss of caveolae, through caveolin-1 or cavin1 depletion, causes activation of PtdIns(4, 5)P2 signalling, recruitment of FMNL2 formin, and enhanced cortical actin assembly. How this equates to physiological responses in epithelial cells containing endogenous caveolae is unknown. Here we examined the effect of mechanically-inducing acute disassembly of caveolae in epithelia. We show that perturbation of caveolae, through direct mechanical stress, reinforces the actin cortex at adherens junctions. Increasing interactions with membrane lipids by introducing multiple phosphatidylserine-binding undecad cavin1 (UC1) repeat domains into cavin1 rendered caveolae more stable to mechanical stimuli. This molecular stabilization blocked cortical reinforcement in response to mechanical stress. Cortical reinforcement elicited by the mechanically-induced disassembly of caveolae increased epithelial resilience against tensile stresses. These findings identify the actin cortex as a target of caveola mechanotransduction that contributes to epithelial integrity.

cell biology↗

bioRxiv2022

FBXL4 suppresses mitophagy by restricting the accumulation of NIX and BNIP3 mitophagy receptors

Cells selectively remove damaged or excessive mitochondria through mitophagy, a specialized form of autophagy, to maintain mitochondrial quality and quantity. Mitophagy is induced in response to diverse conditions, including hypoxia, cellular differentiation, and mitochondrial damage. However, the mechanisms by which cells remove specific dysfunctional mitochondria under steady-state conditions to fine-tune mitochondrial content are not well understood. Here, we report that SCFFBXL4, an SKP1/CUL1/F-box protein ubiquitin ligase complex, localizes to the mitochondrial outer membrane in unstressed cells and mediates the constitutive ubiquitylation and degradation of the mitophagy receptors NIX and BNIP3 to suppress basal levels of mitophagy. We demonstrate that, unlike wild-type FBXL4, pathogenic variants of FBXL4 that cause encephalopathic mtDNA depletion syndrome (MTDPS13), do not efficiently interact with the core SCF ubiquitin ligase machinery or mediate the degradation of NIX and BNIP3. Thus, we reveal a molecular mechanism that actively suppresses mitophagy via preventing NIX and BNIP3 accumulation and propose that excessive basal mitophagy in the FBXL4-associated mtDNA depletion syndrome is caused by dysregulation of NIX and BNIP3 turnover. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=195 SRC="FIGDIR/small/511867v1_ufig1.gif" ALT="Figure 1"> View larger version (60K): org.highwire.dtl.DTLVardef@19c97f6org.highwire.dtl.DTLVardef@1bca5a3org.highwire.dtl.DTLVardef@1e9419org.highwire.dtl.DTLVardef@18d28a9_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

bioRxiv2022

ESCPE-1 Mediates Retrograde Endosomal Sorting of the SARS-CoV-2 Host Factor Neuropilin-1

Endosomal sorting maintains cellular homeostasis by recycling transmembrane proteins and associated proteins and lipids (termed cargoes) from the endosomal network to multiple subcellular destinations, including retrograde traffic to the trans-Golgi network (TGN). Viral and bacterial pathogens subvert retrograde trafficking machinery to facilitate infectivity. Here, we develop a proteomic screen to identify novel retrograde cargo proteins of the Endosomal SNX-BAR Sorting Complex Promoting Exit-1 (ESCPE-1). Using this methodology, we identify Neuropilin-1 (NRP1), a recently characterised host factor for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, as a cargo directly bound and trafficked by ESCPE-1. ESCPE-1 mediates retrograde trafficking of engineered nanoparticles functionalised with the NRP1-interacting peptide of the SARS-CoV-2 Spike protein. ESCPE-1 sorting of NRP1 may therefore play a role in the intracellular membrane trafficking of NRP1-interacting viruses such as SARS-CoV-2.

cell biology↗

bioRxiv2021

Mechanistic basis for SNX27-Retromer coupling to ESCPE-1 in promoting endosomal cargo recycling.

Sorting nexin-27 (SNX27)-Retromer is an endosomal sorting complex that orchestrates endosome-to-plasma membrane recycling of hundreds of internalized receptors, channels and transporters, enzymes and adhesion molecules. While SNX27-Retromer is essential for development, subtle functional defects are observed in human disease, most notably neurodegenerative and neurological disorders. Achieving a thorough mechanistic dissection of SNX27-Retromer is central to understanding endosomal sorting in health and disease. Here we combine biochemical, structural and cellular analyses to establish the mechanistic basis through which SNX27-Retromer couples to the membrane tubulating ESCPE-1 complex (Endosomal SNX-BAR sorting complex for promoting exit 1). We show that a conserved surface in the FERM (4.1/ezrin/radixin/moesin) domain of SNX27 directly binds acidic-Asp-Leu-Phe (aDLF) motifs in the disordered amino-termini of the SNX1 and SNX2 subunits of ESCPE-1. This interaction hands-over SNX27-Retromer captured integral membrane proteins into ESCPE-1 tubular profiles to promote their cell surface recycling. Through phylogenetic analysis, we reveal that SNX27:Retromer:ESCPE-1 assembly evolved in a stepwise manner during the early evolution of metazoans, which reflects the increasing complexity of endosomal sorting from the ancestral opisthokont to modern animals.

cell biology↗