A loss-of-function screen reveals SNX5 and SNX6 as potential components of the mammalian retromer

Wassmer, Thomas; Attar, Naomi; Bujny, Miriam V.; Oakley, Jacqueline; Traer, Colin J.; Cullen, Peter J.

doi:10.1242/jcs.03302

Cited by 230 publications

(316 citation statements)

References 38 publications

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“…Depletion of SNX6 did not have any effect on the membrane localization of SNX2, indicating that SNX2's association with membranes does not require SNX6 (Figure 3C, m-p). In contrast, consistent with previous reports [26], very weak if any SNX1 signals were detected in SNX6-depleted cells ( Figure 3C, e-h), suggesting that SNX1 might be stabilized via complexing with SNX6. Moreover, although depletion of PI(3)P caused dissociation of SNX1, SNX2 and SNX6 from the membrane, it did not weaken SNX1's interaction with SNX5 or SNX6 ( Figure 3D), indicating that the interactions between SNXs do not require their membrane association.…”

Section: Snx6's Recruitment To Endosomal Membranes Requires Pi(3)p Ansupporting

confidence: 93%

“…Consistent with previous reports [26], we observed colocalization of SNX6 with SNX1-labeled vesicles and tubulovesicular structures (Supplementary information, Movie S1). To confirm that SNX6 localizes to early endosomes, we also performed live imaging of HeLa cells co-expressing YFP-EEA1 and mCherry-SNX6.…”

Section: Snx6 Interacts With Snx1/2 and Associates With Retromer-posisupporting

confidence: 92%

“…Confocal immunofluorescence microscopy showed that CI-MPR distribution was unchanged upon SNX6 R68A mutant expression ( Figure 5A, d-f), possibly because the cytosolic mutant's affinity for SNX1 is much lower than that of WT SNX6 ( Figure 4B), which is abundant in cells. Since SNX6 binds to SNX1, and depletion of both proteins causes CI-MPR dispersal [26], we reasoned that suppressing protein levels of both SNX1 and SNX6 would decrease the intracellular concentrations of these retromer components to such a low level that they become limiting factors for the transport machinery required for CI-MPR retrieval (Supplementary information, Figure S5). To examine SNX6's role in endosome-to-TGN transport, we depleted SNX1/6 (Supplementary information, Figure S5) in HeLa cells by RNAi and attempted to rescue the disruption of CI-MPR retrieval by WT or mutant SNX6 proteins expressed at low levels.…”

Section: Ci-mpr Trafficking From Endosome To Tgn Requires the Full Acmentioning

confidence: 99%

“…SNX1 and SNX2 are mammalian orthologs of Vps5p [15,25], whereas there is no obvious sequence homolog of Vps17p in mammals. Most recently, an RNAi loss-of-function screen in HeLa cells identified SNX5 and SNX6 as putative components of the SNX subcomplex [26].…”

Section: Introductionmentioning

confidence: 99%

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The retromer component SNX6 interacts with dynactin p150Glued and mediates endosome-to-TGN transport

et al. 2009

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The retromer is a protein complex that mediates retrograde transport of transmembrane cargoes from endosomes to the trans-Golgi network (TGN). It is comprised of a cargo-selection subcomplex of Vps26, Vps29 and Vps35 and a membrane-binding coat subcomplex of sorting nexins (SNXs). Previous studies identified SNX1/2 as one of the components of the SNX subcomplex, and SNX5/6 as candidates for the second SNX. How the retromer-associated cargoes are recognized and transported by molecular motors are largely unknown. In this study, we found that one of SNX1/2's dimerization partners, SNX6, interacts with the p150 Glued subunit of the dynein/dynactin motor complex. We present evidence that SNX6 is a component of the retromer, and that recruitment of the motor complex to the membrane-associated retromer requires the SNX6-p150 Glued interaction. Disruption of the SNX6-p150 Glued interaction causes failure in formation and detachment of the tubulovesicular sorting structures from endosomes and results in block of CI-MPR retrieval from endosomes to the TGN. These observations indicate that in addition to SNX1/2, SNX6 in association with the dynein/dynactin complex drives the formation and movement of tubular retrograde intermediates.

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Section: Snx6's Recruitment To Endosomal Membranes Requires Pi(3)p Ansupporting

confidence: 93%

Section: Snx6 Interacts With Snx1/2 and Associates With Retromer-posisupporting

confidence: 92%

Section: Ci-mpr Trafficking From Endosome To Tgn Requires the Full Acmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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The retromer component SNX6 interacts with dynactin p150Glued and mediates endosome-to-TGN transport

et al. 2009

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“…Since then, retromer has been implicated in a broad range of protein-sorting pathways and has most recently been shown to be required for the maintenance of proper Wnt secretion in the development of both Caenorhabditis elegans and Drosophila melanogaster (Belenkaya et al, 2008;Franch-Marro et al, 2008;Pan et al, 2008;Port et al, 2008;Yang et al, 2008), underscoring its importance in retrograde transport. In yeast retromer is composed of five proteins that assemble into two subcomplexes: Vps5 and Vps17 (Snx1/ Snx2 and Snx5/Snx6 in mammals) and Vps26, Vps29, and Vps35 (and the homologous proteins in metazoans; Seaman et al, 1998;Haft et al, 2000;Wassmer et al, 2007). Vps5 and Vps17 are members of the sorting nexin family of proteins and are responsible for localizing the retromer complex to tubular endosomal membranes via the concerted action of their BAR (Bin/Amphiphysin/RVS) and PX (phox homology) domains (Carlton et al, 2004;Seet and Hong, 2006).…”

Section: Introductionmentioning

confidence: 99%

Opposing Activities of the Snx3-Retromer Complex and ESCRT Proteins Mediate Regulated Cargo Sorting at a Common Endosome

Strochlic

Schmiedekamp

Lee

et al. 2008

MBoC

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Endocytosed proteins are either delivered to the lysosome to be degraded or are exported from the endosomal system and delivered to other organelles. Sorting of the Saccharomyces cerevisiae reductive iron transporter, composed of the Fet3 and Ftr1 proteins, in the endosomal system is regulated by available iron; in iron-starved cells, Fet3-Ftr1 is sorted by Snx3/Grd19 and retromer into a recycling pathway that delivers it back to the plasma membrane, but when starved cells are exposed to iron, Fet3-Ftr1 is targeted to the lysosome-like vacuole and is degraded. We report that iron-induced endocytosis of Fet3-Ftr1 is independent of Fet3-Ftr1 ubiquitylation, and after endocytosis, degradation of Fet3-Ftr1 is mediated by the multivesicular body (MVB) sorting pathway. In mutant cells lacking any component of the ESCRT protein-dependent MVB sorting machinery, the Rsp5 ubiquitin ligase, or in wild-type cells expressing Fet3-Ftr1 lacking cytosolic lysyl ubiquitin acceptor sites, Fet3-Ftr1 is constitutively sorted into the recycling pathway independent of iron status. In the presence and absence of iron, Fet3-Ftr1 transits an endosomal compartment where a subunit of the MVB sorting receptor (Vps27), Snx3/Grd19, and retromer proteins colocalize. We propose that this endosome is where Rsp5 ubiquitylates Fet3-Ftr1 and where the recycling and degradative pathways diverge. INTRODUCTIONThe endocytic pathway is initiated with the internalization of protein and lipid cargo from the plasma membrane. After endocytosis, internalized molecules transit through the endosomal system, a network of interconnected organelles that process and sort cargo molecules back to the plasma membrane, to other organelles, or to lysosomes for degradation. In mammalian cells, early endosomes are pleiomorphic organelles composed of vacuolar domains that contain lumenal vesicles and tubular domains that lead into and emanate from the vacuolar portion. This distinct architecture has functional significance. The tubular elements of the organelle are regions where the majority of internalized plasma membrane proteins and lipids are exported and recycled from the endosomal system via bulk flow and cargo-specific processes (Mayor et al., 1993;Maxfield and McGraw, 2004;Bonifacino and Rojas, 2006). Cargo proteins that are not exported are delivered to lysosomes and many of these are degraded. This occurs via an active, signal-mediated event in which these proteins are targeted to the vacuolar region of the endosome where they are incorporated into vesicles that bud from the limiting membrane into the lumen of the organelle (Piper and Katzmann, 2007). During endosome maturation lumenal vesicles accrue until fusion of the late endosome with the lysosome results in degradation of these vesicles and their contents. A central sorting function of the endosomal system is thus to segregate cargo to be exported from the endosome from cargo to be degraded via lysosomal proteolysis.The core components of the recycling (retrograde) and degradative sorting pathways have be...

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The functional roles of retromer in Parkinson's disease

2017

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The endosomal system is critical for the maintenance of intracellular homeostasis, and defects in this system are often linked to neurological disorders. The retromer complex is a critical coordinator of endosomal dynamics and has functional roles in multiple cellular processes through sorting cargoes from endosomes to the trans-Golgi network (TGN) or to the plasma membrane. Mammalian retromer comprises a core Vps26-Vps35-Vps29 trimer and associates with a range of proteins to generate endosomal tubular-vesicular carriers. Alterations in retromer function or its molecular organization have been a rising risk factor for Parkinson's disease (PD). Although genetic evidence has shown several variants within retromer in late-onset PD cases, the exact molecular machineries by which retromer variants induce the development of PD are still not completely elucidated. In this Review, we will focus on the functional roles of retromer in neuronal health, summarize advances in defining the cellular pathological phenotype caused by retromer deficiency or PD-linked retromer variants and discuss the potential clues of how retromer deregulation may contribute to PD pathogenesis.Keywords: autophagy; endosomal trafficking; lysosome; mitochondria; Parkinson's disease (PD); retromer The endosomal network is a highly dynamic and orchestrated system, which serves a vital function in coordinating the communication and exchange of associated proteins and lipids between intracellular membrane-bounded compartments. Once within the network, cargo molecules originating from the plasma membrane and biosynthetic pathways are targeted to a common sorting station, the early endosome, from where cargoes are sorted towards specialized intracellular locations following multiple trafficking routes. They can be either sorted into late endosomes/ lysosomes for degradation or retrieved to the plasma membrane or the trans-Golgi network (TGN). The efficient and accurate delivery of cargo contents within intracellular compartments is critical for the maintenance of intracellular homeostasis, and defects on it are often associated with multiple human diseases. The evolutionary conserved protein complex, termed Abbreviations AMPAR, a-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor; APEX, ascorbate peroxidase; APP, b-amyloid precursor protein; ATG9A, autophagy-associated protein; Ab, elevated b-amyloid peptide; BAR, Bin-Amphiphysin-Rvs; BDNF, brain-derived neurotrophic factor; CI-M6PR, cation-independent mannose 6-phosphate receptor; CLN5, ceroid lipouscinosis neuronal protein-5; CMA, chaperon-mediated autophagy; Crb, crumb; DA, dopaminergic; DMTI-II, divalent metal transporter II; DRD1, dopamine receptor D1; Drp1, dynamin-related protein 1; EHD1, Eps15 homology domain-containing protein-1; FERM, 4.1/ezrin/radixin/moesin; GLUT1, glucose transporter 1; Kir3, G protein-gated inward rectifying potassium channels; Lamp2a, lysosome-associated membrane glycoprotein 2a; LBs, Lewy bodies; MAPL, mitochondria-anchored protein ligase; MDVs, mitochondria-de...

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A loss-of-function screen reveals SNX5 and SNX6 as potential components of the mammalian retromer

Cited by 230 publications

References 38 publications

The retromer component SNX6 interacts with dynactin p150Glued and mediates endosome-to-TGN transport

The retromer component SNX6 interacts with dynactin p150Glued and mediates endosome-to-TGN transport

Opposing Activities of the Snx3-Retromer Complex and ESCRT Proteins Mediate Regulated Cargo Sorting at a Common Endosome

The functional roles of retromer in Parkinson's disease

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