The CORVET complex promotes tethering and fusion of Rab5/Vps21-positive membranes

Department of Biology/Chemistry, University of Osnabrück, 49076 Osnabrück, Germany.
Proceedings of the National Academy of Sciences (Impact Factor: 9.67). 03/2013; 110(10):3823-8. DOI: 10.1073/pnas.1221785110
Source: PubMed


Membrane fusion along the endocytic pathway occurs in a sequence of tethering, docking, and fusion. At endosomes and vacuoles, the CORVET (class C core vacuole/endosome tethering) and HOPS (homotypic fusion and vacuole protein sorting) tethering complexes require their organelle-specific Rabs for localization and function. Until now, despite the absence of experimental evidence, it has been assumed that CORVET is a membrane-tethering factor. To test this theory and understand the mechanistic analogies with the HOPS complex, we set up an in vitro system, and establish CORVET as a bona-fide tether for Vps21-positive endosome/vacuole membranes. Purified CORVET binds to SNAREs and Rab5/Vps21-GTP. We then demonstrate that purified CORVET can specifically tether Vps21-positive membranes. Tethering via CORVET is dose-dependent, stimulated by the GEF Vps9, and inhibited by Msb3, the Vps21-GAP. Moreover, CORVET supports fusion of isolated membranes containing Vps21. In agreement with its role as a tether, overexpressed CORVET drives Vps21, but not the HOPS-specific Ypt7 into contact sites between vacuoles, which likely represent vacuole-associated endosomes. We therefore conclude that CORVET is a tethering complex that promotes fusion of Rab5-positive membranes and thus facilitates receptor down-regulation and recycling at the late endosome.

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    • "Rab5 also recruits the Class C core vacuole/endosome tethering (CORVET) complex to the membrane of EEs (Balderhaar et al., 2013). The yeast CORVET complex consists of four conserved core subunits (Vps11, Vps16, Vps18, and Vps33) and two additional subunits (Vps3 and Vps8) that directly interact with activated Rab5/Vps21 (Balderhaar et al., 2013; Peplowska et al., 2007). The homotypic fusion and protein sorting (HOPS) complex shares the same four core subunits with the CORVET complex, but has Vps39 and Vps41 as effectors for Rab7/Ypt7 (Price et al., 2000). "
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    ABSTRACT: Endosomes undergo extensive spatiotemporal rearrangements as proteins and lipids flux through them in a series of fusion and fission events. These controlled changes enable the concentration of cargo for eventual degradation while ensuring the proper recycling of other components. A growing body of studies has now defined multiple recycling pathways from endosomes to the trans-Golgi network (TGN) which differ in their molecular machineries. The recycling process requires specific sets of lipids, coats, adaptors, and accessory proteins that coordinate cargo selection with membrane deformation and its association with the cytoskeleton. Specific tethering factors and SNARE (SNAP (Soluble NSF Attachment Protein) Receptor) complexes are then required for the docking and fusion with the acceptor membrane. Herein, we summarize some of the current knowledge of the machineries that govern the retrograde transport from endosomes to the TGN. Copyright © 2015 Elsevier Inc. All rights reserved. Portions of this chapter is prepared by US government employees. Published by Elsevier Inc. All rights reserved.
    International review of cell and molecular biology 08/2015; 318:159-202. DOI:10.1016/bs.ircmb.2015.05.005 · 3.42 Impact Factor
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    • "In analogy, it is likely that the Vps41-homolog Vps8 and the Vps39-like Vps3 subunit are positioned similarly in CORVET. Indeed, both complexes can tether Vps21 and Ypt7-decorated membranes in vitro.17-19 "
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    ABSTRACT: Endosomal and vacuole fusion depends on the two homologous tethering complexes CORVET and HOPS. HOPS binds the activated Rab GTPase Ypt7 via two distinct subunits, Vps39 and Vps41. To understand the participation and possible polarity of Vps41 and Vps39 during tethering, we used an in vivo approach. For this, we established the ligand-induced relocalization to the plasma membrane, using the Mon1-Ccz1 GEF complex that activates Ypt7 on endosomes. We then employed slight overexpression to compare the mobility of the HOPS-specific Vps41 and Vps39 subunits during this process. Our data indicate an asymmetry in the Rab-specific interaction of the two HOPS subunits: Vps39 is more tightly bound to the vacuole, and relocalizes the entire vacuole to the plasma membrane, whereas Vps41 behaved like the more mobile subunit. This is due to their specific Rab binding, as the mobility of both subunits was similar in ypt7∆ cells. In contrast, both HOPS subunits were far less mobile if tagged endogenously, suggesting that the entire HOPS complex is tightly bound to the vacuole in vivo. Similar results were obtained for the endosomal association of CORVET, when we followed its Rab-specific subunit Vps8. Our data provide in vivo evidence for distinct Rab specificity within HOPS, which may explain its function during tethering, and indicate that these tethering complexes are less mobile within the cell than previously anticipated.
    04/2014; 4(2):e29191. DOI:10.4161/cl.29191
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    • "Endosomes contain at least two characterized Rab effectors that function in fusion, the dimeric early endosomal antigen (EEA1) and similar coiled-coil tethers [9]–[12] and the heterohexameric CORVET complex [13], [14]. Whereas EEA1 seems to have a function restricted to heterotypic fusion of endocytic vesicles with early endosomes [15], [16], CORVET has been implicated in endosome-endosome fusion in yeasts [14], [17], [18]. CORVET, like its sibling, the late endosomal/vacuolar HOPS complex, is a heterohexamer [7], [13], [14], [18], [19]. "
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    ABSTRACT: Endosomal biogenesis depends on multiple fusion and fission events. For fusion, the heterohexameric CORVET complex as an effector of the endosomal Rab5/Vps21 GTPase has a central function in the initial tethering event. Here, we show that the CORVET-specific Vps3 and Vps8 subunits, which interact with Rab5/Vps21, require their N-terminal domains for localization and function. Surprisingly, CORVET may lack either one of the two N-terminal domains, but not both, to promote protein sorting via the endosome. The dually truncated complex mislocalizes to the cytosol and is impaired in endocytic protein sorting, but not in assembly. Furthermore, the endosomal localization can be rescued by overexpression of Vps21 or one of the truncated CORVET subunits, even though CORVET assembly is not impaired by loss of the N-terminal domains or in strains lacking all endosomal Rab5s and Ypt7. We thus conclude that CORVET requires only its C-terminal domains for assembly and has beyond its putative β-propeller domains additional binding sites for endosomes, which could be important to bind Vps21 and other endosome-specific factors for efficient endosome tethering.
    PLoS ONE 06/2013; 8(6):e67307. DOI:10.1371/journal.pone.0067307 · 3.23 Impact Factor
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