Helical Structures of ESCRT-III Are Disassembled by VPS4

Lata, Suman; Schoehn, Guy; Jain, Ankur; Pires, Ricardo H.; Piehler, Jacob; Göttlinger, Heinrich G.; Weissenhorn, Winfríed

doi:10.1126/science.1161070

Cited by 324 publications

(471 citation statements)

References 26 publications

(47 reference statements)

Supporting

Mentioning

443

Contrasting

Unclassified

Order By: Relevance

“…Less consistent with the yeast-based mechanism, CHMP6, CHMP3, and CHMP1A/B knockdowns have little or no HIV-1 release phenotype (26,32). These differences are hard to reconcile with live-cell imaging detecting CHMP1B at bud sites (33) and with the finding that CHMP2A and CHMP3 coassemble in vitro into tubules but do not do so on their own (34). The differences in the results with yeast ESCRT-III proteins and the knockdown, imaging, and tubule formation data for human proteins called out for further investigation.…”

mentioning

confidence: 68%

See 1 more Smart Citation

In vitro reconstitution of the ordered assembly of the endosomal sorting complex required for transport at membrane-bound HIV-1 Gag clusters

Carlson

Hurley

2012

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

Most membrane-enveloped viruses depend on host proteins of the endosomal sorting complex required for transport (ESCRT) machinery for their release. HIV-1 is the prototypic ESCRT-dependent virus. The direct interactions between HIV-1 and the early ESCRT factors TSG101 and ALIX have been mapped in detail. However, the full pathway of ESCRT recruitment to HIV-1 budding sites, which culminates with the assembly of the late-acting CHMP4, CHMP3, CHMP2, and CHMP1 subunits, is less completely understood. Here, we report the biochemical reconstitution of ESCRT recruitment to viral assembly sites, using purified proteins and giant unilamellar vesicles. The myristylated full-length Gag protein of HIV-1 was purified to monodispersity. Myr-Gag forms clusters on giant unilamellar vesicle membranes containing the plasma membrane lipid PIð4,5ÞP 2 . These Gag clusters package a fluorescent oligonucleotide, and recruit early ESCRT complexes ESCRT-I or ALIX with the appropriate dependence on the Gag PTAP and LYPðXÞ n L motifs. ALIX directly recruits the key ESCRT-III subunit CHMP4. ESCRT-I can only recruit CHMP4 when ESCRT-II and CHMP6 are present as intermediary factors. Downstream of CHMP4, CHMP3 and CHMP2 assemble synergistically, with the presence of both subunits required for efficient recruitment. The very late-acting factor CHMP1 is not recruited unless the pathway is completed through CHMP3 and CHMP2. These findings define the minimal sets of components needed to complete ESCRT assembly at HIV-1 budding sites, and provide a starting point for in vitro structural and biophysical dissection of the system. confocal microscopy | host-pathogen interaction | membrane traffic | virus budding M ost membrane-enveloped viruses utilize host proteins of the endosomal sorting complex required for transport (ESCRT) machinery for their release (1-3). This dependence was first described for HIV-1, where mutation of an ESCRTbinding motif (late domain) in the viral protein Gag was shown to stall virus bud release at a late stage in virus assembly (4, 5). Late domains have subsequently been found in other retroviruses and numerous other virus families including, e.g., flavi-, filo-and rhabdoviruses (6). Indeed, the only well-characterized case of ESCRT-independent release of a membrane-enveloped virus is influenza virus (7). In normal cell physiology, ESCRTs function in cytokinesis, formation of intralumenal vesicles in multivesicular endosomes, and vesicle release from the plasma membrane (8-10). These seemingly disparate processes all involve membrane budding away from the cytoplasm, and ESCRTs are the only described protein machinery to perform vesicle formation with this topology, which is analogous to virus release.The initial events in HIV-1 ESCRT recruitment are well characterized. The structural protein of HIV-1, Gag, binds earlyacting ESCRT factors through two late-domain motifs in its C-terminal p6 domain: a PTAP sequence that interacts with the TSG101 subunit of ESCRT-I (11-16) and a LYPðXÞ n L motif that interacts with the ESCRT...

show abstract

mentioning

confidence: 68%

“…CHMP2A and CHMP3 coassemble into tubes in vitro that may represent structures involved in membrane neck scission (34). The yeast cognates of CHMP2 and CHMP3 are Vps2 and Vps24, which seem to preferentially heterodimerize with one another (23).…”

Section: Discussionmentioning

confidence: 99%

In vitro reconstitution of the ordered assembly of the endosomal sorting complex required for transport at membrane-bound HIV-1 Gag clusters

Carlson

Hurley

2012

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

show abstract

“…CHMP4B also has been shown to form spiral-shaped filaments when overexpressed in cells (25). In addition, in vitro studies have shown that ESCRT-III components spontaneously polymerize into spiral filaments that expose their membrane interaction sites on the surface, allowing them to bind tightly to surrounding membranes (26). A plausible model (Fig.…”

Section: Discussionmentioning

confidence: 99%

Dynamics of endosomal sorting complex required for transport (ESCRT) machinery during cytokinesis and its role in abscission

Elia

Sougrat

Spurlin

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

354

550

View full text Add to dashboard Cite

The final stage of cytokinesis is abscission, the cutting of the narrow membrane bridge connecting two daughter cells. The endosomal sorting complex required for transport (ESCRT) machinery is required for cytokinesis, and ESCRT-III has membrane scission activity in vitro, but the role of ESCRTs in abscission has been undefined. Here, we use structured illumination microscopy and timelapse imaging to dissect the behavior of ESCRTs during abscission. Our data reveal that the ESCRT-I subunit tumor-susceptibility gene 101 (TSG101) and the ESCRT-III subunit charged multivesicular body protein 4b (CHMP4B) are sequentially recruited to the center of the intercellular bridge, forming a series of cortical rings. Late in cytokinesis, however, CHMP4B is acutely recruited to the narrow constriction site where abscission occurs. The ESCRT disassembly factor vacuolar protein sorting 4 (VPS4) follows CHMP4B to this site, and cell separation occurs immediately. That arrival of ESCRT-III and VPS4 correlates both spatially and temporally with the abscission event suggests a direct role for these proteins in cytokinetic membrane abscission.

show abstract

“…A spiral polymer of ESCRT-III subunits forms at the bud neck to allow ILVs to pinch off into the lumen. This same membrane constriction activity of ESCRT-III that is harnessed for ILV formation on endosomes is also used for budding of retroviruses from the plasma membrane and membrane scission during cytokinesis in animal cells and some Archea (Hanson et al 2008;Lata et al 2008;Lindas et al 2008;Samson et al 2008;Wollert et al 2009;Henne et al 2012Henne et al , 2013. Before ILV scission, Ub can be removed from cargo to help replenish the cytosolic pool of Ub.…”

Section: Ubiquitin-dependent Sorting In Endocytosismentioning

confidence: 99%

Ubiquitin-Dependent Sorting in Endocytosis

Piper

Đikić

Lukacs

2014

Cold Spring Harbor Perspectives in Biology

191

176

View full text Add to dashboard Cite

When ubiquitin (Ub) is attached to membrane proteins on the plasma membrane, it directs them through a series of sorting steps that culminate in their delivery to the lumen of the lysosome where they undergo complete proteolysis. Ubiquitin is recognized by a series of complexes that operate at a number of vesicle transport steps. Ubiquitin serves as a sorting signal for internalization at the plasma membrane and is the major signal for incorporation into intraluminal vesicles of multivesicular late endosomes. The sorting machineries that catalyze these steps can bind Ub via a variety of Ub-binding domains. At the same time, many of these complexes are themselves ubiquitinated, thus providing a plethora of potential mechanisms to regulate their activity. Here we provide an overview of how membrane proteins are selected for ubiquitination and deubiquitination within the endocytic pathway and how that ubiquitin signal is interpreted by endocytic sorting machineries. HOW PROTEINS ARE SELECTED FOR UBIQUITINATIONU biquitin (Ub) is covalently attached to substrate proteins by the concerted action of E2-conjugating enzymes and E3 ligases (Varshavsky 2012). Most of the specificity and regulation of ubiquitination is at the level of the E3 ligases, which vastly outnumber the E2-conjugating enzymes. Ubiquitination results from the transfer of Ub, held either by the E2 or in some cases by the E3 as a high-energy thioester bond, onto a substrate protein, typically on the terminal amide of a substrate acceptor lysine side chain. Owing to the intense interest in the ubiquitination system, many of the simple rules and distinctions concerning different types of ligases, their specificity for forming particular polyUb chains, and even the kinds of residues in substrate proteins that can be ubiquitin modified, have been blurred with the discovery of mixed poly-Ub chains, new enzymatic mechanisms for Ub transfer, and ubiquitination of nonlysine residues (Kulathu and Komander 2012;Wenzel and Klevit 2012;McDowell and Philpott 2013). Nonetheless, in basic terms, Ub ligases function as platforms that coordinate substrate recognition with Ub transfer as well as configuring poly-Ub chain topology by the E2-conjugating enzyme. Substrates can be bound directly by the E3 ligase or by adaptor proteins that in turn bind to ligases, and selecEditors:

show abstract

Helical Structures of ESCRT-III Are Disassembled by VPS4

Cited by 324 publications

References 26 publications

In vitro reconstitution of the ordered assembly of the endosomal sorting complex required for transport at membrane-bound HIV-1 Gag clusters

In vitro reconstitution of the ordered assembly of the endosomal sorting complex required for transport at membrane-bound HIV-1 Gag clusters

Dynamics of endosomal sorting complex required for transport (ESCRT) machinery during cytokinesis and its role in abscission

Ubiquitin-Dependent Sorting in Endocytosis

Contact Info

Product

Resources

About