Constitutively active ESCRT-II suppresses the MVB-sorting phenotype of ESCRT-0 and ESCRT-I mutants

Mageswaran, Shrawan Kumar; Johnson, Natalie; Odorizzi, Greg; Babst, Markus

doi:10.1091/mbc.e14-10-1469

Cited by 23 publications

(28 citation statements)

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“…Intriguingly, the Glue domain therefore seems to play an inhibitory role for the activation of ESCRT-II as upon removal of this region; the virus rescue is almost as efficient as that achieved by the FL protein (Figure 4). An autoinhibitory role within members of ESCRT is not uncommon (McCullough et al, 2008) and has been proposed also for ESCRT-II for its role in MVB sorting in yeast (Mageswaran et al, 2015). However, activation of ESCRT-II in MVB sorting is slightly different from that in HIV budding, as the removal of the Glue domain is not sufficient to rescue the cargo sorting defective phenotype; a mutation is additionally required at its C terminus.…”

Section: Discussionmentioning

confidence: 99%

“…In yeast, this domain contains two Npl4‐type zinc finger domains, NZF1 and NZF2; the former has been demonstrated to interact with VPS28 of ESCRT‐I (Teo et al, ). The interaction between ESCRT‐I and ‐II plays an important role in the endosomal recruitment of ESCRT‐II (Mageswaran, Johnson, Odorizzi, & Babst, ). However, in mammalian systems, both NZF motifs are absent.…”

Section: Introductionmentioning

confidence: 99%

“…The interaction between ESCRT-I and -II plays an important role in the endosomal recruitment of ESCRT-II (Mageswaran, Johnson, Odorizzi, & Babst, 2015). However, in mammalian systems, both NZF motifs are absent.…”

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confidence: 99%

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ESCRT‐II functions by linking to ESCRT‐I in human immunodeficiency virus‐1 budding

Meng

Abbink

et al. 2020

Cellular Microbiology

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Human immunodeficiency virus (HIV) uses the ESCRT (endosomal sorting complexes required for transport) protein pathway to bud from infected cells. Despite the roles of ESCRT‐I and ‐III in HIV budding being firmly established, participation of ESCRT‐II in this process has been controversial. EAP45 is a critical component of ESCRT‐II. Previously, we utilised a CRISPR‐Cas9 EAP45 knockout cell line to assess the involvement of ESCRT‐II in HIV replication. We demonstrated that the absence of ESCRT‐II impairs HIV budding. Here, we show that virus spread is also defective in physiologically relevant CRISPR/Cas9 EAP45 knockout T cells. We further show reappearance of efficient budding by re‐introduction of EAP45 expression into EAP45 knockout cells. Using expression of selected mutants of EAP45, we dissect the domain requirement responsible for this function. Our data show at the steady state that rescue of budding is only observed in the context of a Gag/Pol, but not a Gag expressor, indicating that the size of cargo determines the usage of ESCRT‐II. EAP45 acts through the YPXL‐ALIX pathway as partial rescue is achieved in a PTAP but not a YPXL mutant virus. Our study clarifies the role of ESCRT‐II in the late stages of HIV replication and reinforces the notion that ESCRT‐II plays an integral part during this process as it does in sorting ubiquitinated cargos and in cytokinesis.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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ESCRT‐II functions by linking to ESCRT‐I in human immunodeficiency virus‐1 budding

Meng

Abbink

et al. 2020

Cellular Microbiology

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“…In forming vesicles, 487 positively curved membrane is limited to the rim, in contrast to negative curvature, which 488 is found throughout the nascent bud. We, therefore, speculate that sensing positive 489 curvature allows ESCRT-II and Vps20 to target Snf7 nucleation to the rim of shallow 490 membrane invaginations pre-formed by crowded cargo and early ESCRT-proteins during 491 ILV formation (72)(73)(74)(75). Since Snf7 exclusively binds to low-curvature membrane and 492 polymerizes in spiral-shape, it seems conceivable that a nucleated filament grows around 493 the nascent bud.…”

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confidence: 98%

Vps4 triggers sequential subunit exchange in ESCRT-III polymers that drives membrane constriction and fission

Pfitzner

Mercier

Roux

2019

Preprint

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1ESCRT-III is a ubiquitous complex which catalyzes membrane fission from 2 within membrane necks via an as yet unknown mechanism. Here, we reconstituted in 3 vitro the ESCRT-III complex onto membranes. We show that based on variable affinities 4 between ESCRT-III proteins and the ATPase Vps4, subunits are recruited to the 5 membrane in a Vps4-driven sequence that starts with Snf7 and ends with Did2 and Ist1 6 which, together, form a fission-active subcomplex. Sequential recruitment of ESCRT-III 7 subunits is coupled to membrane remodeling. Binding of Did2 promoted the formation of 8 membrane protrusions which later constricted and underwent fission mediated by the 9 recruitment of Ist1. Overall, our results provide a mechanism to explain how a sequence 10 of ESCRT-III subunits drives membrane deformation and fission. 14 catalyze membrane fission from within membrane necks in all investigated cellular 15 processes requiring this type of fission event (1-16). ESCRT-III polymers are nucleated 16 by several factors. ESCRT-II, which binds ESCRT-III core subunit Vps20, is the 17 canonical one. Besides Vps20, yeast ESCRT-III complex contains three other core 18 subunits: Snf7, which binds Vps20-ESCRT-II, as well as Vps2 and Vps24, which, in 19 tandem, bind Snf7 and recruit the AAA-ATPase Vps4 via their [17][18][19][20][21][22][23][24][25][26]. 20 While all ESCRT-III subunits have MIM-domains, their specific affinities for Vps4 differ 21 widely (22,23,25). Vps4 ATPase activity remodels ESCRT-III by inducing turnover of 22 ESCRT-III subunits, controlling the balance between polymer growth (27) and 23 disassembly (24-26). ESCRT-III core subunits assemble, alone or in various 24 stoichiometries, into single or multiple stranded filaments (24,(28)(29)(30)(31)(32)(33). These filaments 25 usually exhibit high spontaneous curvature, but a low rigidity, which leads to diverse 26 helical shapes like spirals (30,(32)(33)(34), conical spirals (28, 34) and tubular helices (24, 27 35). While the possibility that transitions occur between those shapes is under debate (36) 28 many findings support the notion that Vps4-dependent ESCRT-III remodeling promotes 29 membrane constriction and fission (7,27,37). The most direct evidence is the 30 asymmetric constriction of in vitro formed CHMP2-CHMP3 (mammalian homologs of 31 Vps2 and Vps24) tubular copolymers by Vps4 (38). While attempts to reconstitute fission 32 in vitro with ESCRT-III core subunits provided conflicting results regarding the role of 33 2 Vps4 (39, 40), the most constricted polymers observed with Snf7, Vps2 and Vps24 34 exceed a radius of 10 nm (24,27,30,32,33), far from the theoretical limit of 3 nm 35 required for spontaneous fission (41) and far from the radius of experimentally observed 36 dynamin pre-fission intermediates (42). This suggests that other subunits or mechanisms 37 are at play to reach sufficient constriction for membrane fission. 38 Aside from core ESCRT-III proteins, accessory subunits are thought to play a role 39 specific to subsets of ESCRT-III functio...

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“…This was first characterized in atomic detail for CHMP3/ hVps24 [26]. a1 and a2 form a 70 A helical hairpin and a3/a4 form short helices that pack against the hairpin (Fig. 3A) [26].…”

Section: Escrt-iii and Vacuolar Protein Sorting 4 (Vps4)mentioning

confidence: 99%

ESCRT‐III and Vps4: a dynamic multipurpose tool for membrane budding and scission

2016

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Complex molecular machineries bud, scission and repair cellular membranes. Components of the multi-subunit endosomal sorting complex required for transport (ESCRT) machinery are enlisted when multivesicular bodies are generated, extracellular vesicles are formed, the plasma membrane needs to be repaired, enveloped viruses bud out of host cells, defective nuclear pores have to be cleared, the nuclear envelope must be resealed after mitosis and for final midbody abscission during cytokinesis. While some ESCRT components are only required for specific processes, the assembly of ESCRT-III polymers on target membranes and the action of the AAAATPase Vps4 are mandatory for every process. In this review, we summarize the current knowledge of structural and functional features of ESCRT-III/Vps4 assemblies in the growing pantheon of ESCRT-dependent pathways. We describe specific recruitment processes for ESCRT-III to different membranes, which could be useful to selectively inhibit ESCRT function during specific processes, while not affecting other ESCRT-dependent processes. Finally, we speculate how ESCRT-III and Vps4 might function together and highlight how the characterization of their precise spatiotemporal organization will improve our understanding of ESCRT-mediated membrane budding and scission in vivo.

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Constitutively active ESCRT-II suppresses the MVB-sorting phenotype of ESCRT-0 and ESCRT-I mutants

Cited by 23 publications

References 50 publications

ESCRT‐II functions by linking to ESCRT‐I in human immunodeficiency virus‐1 budding

ESCRT‐II functions by linking to ESCRT‐I in human immunodeficiency virus‐1 budding

Vps4 triggers sequential subunit exchange in ESCRT-III polymers that drives membrane constriction and fission

ESCRT‐III and Vps4: a dynamic multipurpose tool for membrane budding and scission

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