Human ESCRT-II Complex and Its Role in Human Immunodeficiency Virus Type 1 Release

Langelier, Charles; Schwedler, Uta K. von; Fisher, Robert D.; Domenico, Ivana De; White, Paul L.; Hill, Christopher P.; Kaplan, Jerry; Ward, Diane McVey; Sundquist, Wesley I.

doi:10.1128/jvi.01049-06

Cited by 155 publications

(230 citation statements)

References 89 publications

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“…The main discrepancy between our reconstitution and the published knockdown data is that in the latter case, essentially no budding defect is observed when ESCRT-II subunits or the most upstream ESCRT-III subunit CHMP6 are knocked down (26). In the knockdown study, protein bands for ESCRT-II EAP20/VPS25 and CHMP6 were still visible on Western blots following knockdown, and it cannot be ruled out that the residual levels of these proteins are above the threshhold needed for function.…”

Section: Discussioncontrasting

confidence: 48%

“…Consistent with the analogy to yeast ESCRTs, simultaneous knockdown of CHMP4A/B/C or CHMP2A/B reduces HIV-1 budding to the late-domain independent baseline (32). 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).…”

mentioning

confidence: 68%

“…In the yeast ESCRT system, ESCRT-I recruits the ESCRT-II complex, which in turn binds to the upstream ESCRT-III subunit Vps20 (CHMP6), initiating ESCRT-III assembly (23)(24)(25). However, RNA knockdown experiments have been interpreted as indicating that ESCRT-II and CHMP6 might be dispensable for HIV-I release (26). It has been suggested that ALIX could serve as an alternative to ESCRT-II in bridging ESCRT-I and ESCRT-III, given that ALIX directly binds to both ESCRT-I and ESCRT-III.…”

mentioning

confidence: 99%

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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.

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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...

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Section: Discussioncontrasting

confidence: 48%

mentioning

confidence: 68%

mentioning

confidence: 99%

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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.

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“…In addition, studies have shown a crucial role for cholesterol-rich microdomains of the plasma membrane for shedding vesicle formation 32 as well as for biogenesis of exosomes 14 . In our in vivo experiments, we have observed a decrease in Hh signalling by knocking down Vps22, an ESCRT-II complex component described as dispensable for vesicles derived directly from the plasma membrane 33,34 , but required for MVB formation 35 (reviewed in ref. 36) supporting an MVB fusion origin.…”

Section: Discussionmentioning

confidence: 99%

Exosomes as Hedgehog carriers in cytoneme-mediated transport and secretion

et al. 2014

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The Hedgehog signalling pathway is crucial for development, adult stem cell maintenance, cell migration and axon guidance in a wide range of organisms. During development, the Hh morphogen directs tissue patterning according to a concentration gradient. Lipid modifications on Hh are needed to achieve graded distribution, leading to debate about how Hh is transported to target cells despite being membrane-tethered. Cytonemes in the region of Hh signalling have been shown to be essential for gradient formation, but the carrier of the morphogen is yet to be defined. Here we show that Hh and its co-receptor Ihog are in exovesicles transported via cytonemes. These exovesicles present protein markers and other features of exosomes. Moreover, the cell machinery for exosome formation is necessary for normal Hh secretion and graded signalling. We propose Hh transport via exosomes along cytonemes as a significant mechanism for the restricted distribution of a lipid-modified morphogen.

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“…It is also clear that not all components of the ESCRT machinery are required for virus assembly in the MVB and subsequent exocytosis. HIV-1 budding only requires TSG101 and/or ALIX, and a subset of ESCRT-III proteins (CHMP4, CHMP2 and SKD4) (Langelier, et al 2006, Morita, et al 2011. Herpes simplex virus type I production also requires a functional ESCRT-III complex, but not ALIX or TSG101, whereas Ebola virus budding requires host Vps4/SKD1 activity, but can also bypass the requirement for the TSG101 ESCRT-I component (Pawliczek andCrump 2009, Silvestri, et al 2007).…”

Section: Endocytic Trafficking and Diseasementioning

confidence: 99%

Endocytic regulation of alkali metal transport proteins in mammals, yeast and plants

et al. 2013

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Mulet Salort, JM.; Llopis Torregrosa, V.; Primo Planta, C.; Marques Romero, MC.; Yenush, L. (2013) The relative concentrations of ions and solutes inside cells are actively maintained by several classes of transport proteins, in many cases against their concentration gradient. These transport processes, which consume a large portion of cellular energy, must be constantly regulated. Many structurally distinct families of channels, carriers, and pumps have been characterized in considerable detail during the past decades and defects in the function of some of these proteins have been linked to a growing list of human diseases. The dynamic regulation of the transport proteins present at the cell surface is vital for both normal cellular function and for the successful adaptation to changing environments. The composition of proteins present at the cell surface is controlled on both the transcriptional and post-translational level. Post-translational regulation involves highly conserved mechanisms of phosphorylation-and ubiquitylation-dependent signal transduction routes used to modify the cohort of receptors and transport proteins present under any given circumstances. In this review, we will summarize what is currently known about one facet of this regulatory process: the endocytic regulation of alkali metal transport proteins. The physiological relevance, major contributors, parallels and missing pieces of the puzzle in mammals, yeast and plants will be discussed.

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Human ESCRT-II Complex and Its Role in Human Immunodeficiency Virus Type 1 Release

Cited by 155 publications

References 89 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

Exosomes as Hedgehog carriers in cytoneme-mediated transport and secretion

Endocytic regulation of alkali metal transport proteins in mammals, yeast and plants

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