9.A.3 The Retromer Assembly Apparatus (RetromerAA) Family

The PDZ domain-containing sorting nexin 27 (SNX27) promotes recycling of internalized transmembrane proteins from endosomes to the plasma membrane by linking PDZ-dependent cargo recognition to retromer-mediated transport. Steinberg et al. (2013) employed quantitative proteomics of the SNX27 interactome and quantification of the surface proteome of SNX27- and retromer-suppressed cells to dissect the assembly of the SNX27 complex and provide an unbiased global view of SNX27-mediated sorting. Over 100 cell surface proteins, many of which interact with SNX27, including the glucose transporter GLUT1, the Menkes disease copper transporter ATP7A, various zinc and amino acid transporters, and numerous signalling receptors, require SNX27-retromer to prevent lysosomal degradation and maintain surface levels. Furthermore, direct interaction of the SNX27 PDZ domain with the retromer subunit VPS26 is necessary and sufficient to prevent lysosomal entry of SNX27 cargo. Thus, the SNX27-retromer is a major endosomal recycling hub required to maintain cellular nutrient homeostasis. The retromer complex and sorting nexins (SNXs) are involved in endosomal retrieval and recycling of various transmembrane proteins (Priya and Datta 2022).  Assembly and fission of tubular carriers mediate protein sorting in endosomes (Gopaldass et al. 2024).

Gallon and Cullen 2015 reviewed the molecular architecture and cellular roles of retromer and its various functional partners. The endosomal network trafficks proteins through the cellular endomembrane system. Transmembrane proteins enter endosomes by endocytosis from the plasma membrane or by trafficking from the trans-Golgi network. Endosomal cargo proteins face one of two fates: retention in the endosome, leading ultimately to lysosomal degradation, or export from the endosome for reuse ('recycling'). The balance of protein degradation and recycling is crucial to cellular homoeostasis; inappropriate sorting of proteins to either fate leads to cellular dysfunction.  The protein PTEN (403 aas; P60484) blocks SNX27 from interacting with VPS26, preventing recycling and thus reduces the amount of the glucose transporter, GLUT1 (TC# 2.A.1.1.28) in the plasma membrane (Shinde and Maddika 2017). See also TC family 9.A.63. Membrane trafficking and the retromer complex play roles in Parkinson's and Alzheimer's diseases (Abdul-Rahman et al. 2024).

Aberrations in membrane trafficking pathways have profound effects in cellular dynamics of cellular sorting processes. Sorting nexin 27 (SNX27) is a metazoan-specific sorting nexin protein from the PX-FERM domain family and is required for endosomal recycling of many important transmembrane receptors (Chandra et al. 2021). SNX27-mediated recycling requires association with retromer, one of the best-known regulators of endosomal trafficking. SNX27/retromer downregulation is strongly linked to Down's Syndrome (DS) via glutamate receptor dysfunction and to Alzheimer's Disease (AD) through increased intracellular production of amyloid peptides from amyloid precursor protein (APP) breakdown. SNX27 is further linked to addiction via its role in potassium channel trafficking, and its over-expression is linked to tumorigenesis, cancer progression, and metastasis. The role of SNX27 in regulating cargo recycling from endosomes to the cell surface is firmly established (Chandra et al. 2021).

Retromer and the associated actin polymerizing WASH-complex are essential for the endocytic recycling of a wide range of integral membrane proteins. A hereditary Parkinson's disease causing point mutation (D620N) in the retromer subunit VPS35 perturbs retromer's association with the WASH-complex and also with the uncharacterized protein Ankyrin Repeat Domain Containing Protein 50 (ANKRD50).  Kvainickas et al. 2016 established ANKRD50 as an essential component of the SNX27 -retromer-WASH supercomplex. Depletion of ANKRD50 phenocopied the loss of endosome to cell surface recycling of multiple transmembrane proteins seen upon suppression of SNX27, retromer or WASH-complex components. Mass spectrometric quantification of the cell surface proteome of ANKRD50 depleted cells identified amino acid transporters of the SLC1A family, among them SLC1A4, (TC# 2.A.3.3.1) as additional cargo molecules that depend on ANKRD50 and retromer for their endocytic recycling. Mechanistically, Kvainickas et al. 2016 showed that ANKRD50 simultaneously engages multiple parts of the SNX27 -retromer-WASH complex machinery in a direct and co-operative interaction network that is needed to efficiently recycle the nutrient transporters GLUT1 and many other surface proteins.

As noted above, endosomes are major protein sorting stations in cells. Endosomally localised multi-protein complexes sort integral proteins, including signaling receptors, nutrient transporters, adhesion molecules, and lysosomal hydrolase receptors, for lysosomal degradation or for retrieval and subsequent recycling to various membrane compartments (McNally and Cullen 2018). Correct endosomal sorting of these proteins is essential for maintaining cellular homeostasis, with defects in endosomal sorting implicated in various human pathologies including neurodegenerative disorders. Retromer, a multi-protein complex, is essential for the retrieval and recycling of hundreds of transmembrane proteins. While retromer is a major player in endosomal retrieval and recycling, several studies have identified retrieval mechanisms that are independent of retromer. McNally and Cullen 2018 reviewed endosomal retrieval complexes, particularly retromer-independent systems, and suggest that the yeast system described in TC family 9.A.63 is related to the systems described here. The small GTPase, Rab11A, involved in exocytosis, regulates dense granule transport and secretion during Toxoplasma gondii invasion of host cells and parasite replication (Venugopal et al. 2020).

The structure of the retromer complex (Vps26-Vps29-Vps35) assembled on membrane tubules with the bin/amphiphysin/rvs-domain-containing sorting nexin protein Vps5, has been solved using cryo-electron tomography and subtomogram averaging (Kovtun et al. 2018). It revealed a membrane-associated Vps5 array, from which arches of retromer extend away from the membrane surface. Vps35 forms the 'legs' of these arches, and Vps29 resides at the apex where it is free to interact with regulatory factors. The bases of the arches connect to each other and to Vps5 through Vps26, and the presence of the same arches on coated tubules within cells confirmed their functional importance. Vps5 binds to Vps26 at a position analogous to the previously described cargo- and Snx3-binding site, which suggesed the existence of distinct retromer-sorting nexin assemblies. The structure revealed the architecture of the coat and its mechanism of assembly, and suggested that retromer promotes tubule formation by directing the distribution of sorting nexin proteins on the membrane surface while providing a scaffold for regulatory-protein interactions (Kovtun et al. 2018).

The reaction catalyzed by the retomer assembly apparatus is:

Integral membrane protein in the plasma membrane → endosome → lysosome or plasma membrane