scholarly journals The Chlamydia trachomatis Extrusion Exit Mechanism Is Regulated by Host Abscission Proteins

2019 ◽  
Vol 7 (5) ◽  
pp. 149 ◽  
Author(s):  
Meghan Zuck ◽  
Kevin Hybiske
Keyword(s):  
Host Cell ◽  
Intracellular Pathogens ◽  
Body Protein ◽  
Functional Roles ◽  
Endosomal Sorting ◽  
Exit Mechanism ◽  
Viral Budding ◽  
Membrane Fission ◽  
Exit Strategies ◽  
Vacuolar Protein

The cellular exit strategies of intracellular pathogens have a direct impact on microbial dissemination, transmission, and engagement of immune responses of the host. Chlamydia exit their host via a budding mechanism called extrusion, which offers protective benefits to Chlamydia as they navigate their extracellular environment. Many intracellular pathogens co-opt cellular abscission machinery to facilitate cell exit, which is utilized to perform scission of two newly formed daughter cells following mitosis. Similar to viral budding exit strategies, we hypothesize that an abscission-like mechanism is required to physically sever the chlamydial extrusion from the host cell, co-opting the membrane fission activities of the endosomal sorting complex required for transport (ESCRT) family of proteins that are necessary for cellular scission events, including abscission. To test this, C. trachomatis L2-infected HeLa cells were depleted of key abscission machinery proteins charged multivesicle body protein 4b (CHMP4B), ALIX, centrosome protein 55 (CEP55), or vacuolar protein sorting-associated protein 4A (VPS4A), using RNA interference (RNAi). Over 50% reduction in extrusion formation was achieved by depletion of CHMP4B, VPS4A, and ALIX, but no effect on extrusion was observed with CEP55 depletion. These results demonstrate a role for abscission machinery in C. trachomatis extrusion from the host cell, with ALIX, VPS4A and CHMP4B playing key functional roles in optimal extrusion release.

scholarly journals MITD1 is recruited to midbodies by ESCRT-III and participates in cytokinesis

2012 ◽  
Vol 23 (22) ◽  
pp. 4347-4361 ◽  
Author(s):  
Seongju Lee ◽  
Jaerak Chang ◽  
Benoît Renvoisé ◽  
Anita Tipirneni ◽  
Sarah Yang ◽  
...  
Keyword(s):  
Critical Role ◽  
Multivesicular Body ◽  
Strong Interactions ◽  
Body Protein ◽  
Endosomal Sorting ◽  
Cellular Processes ◽  
Viral Budding ◽  
Membrane Fission ◽  
Downstream Effects ◽  
Trafficking Molecules

Diverse cellular processes, including multivesicular body formation, cytokinesis, and viral budding, require the sequential functions of endosomal sorting complexes required for transport (ESCRTs) 0 to III. Of these multiprotein complexes, ESCRT-III in particular plays a key role in mediating membrane fission events by forming large, ring-like helical arrays. A number of proteins playing key effector roles, most notably the ATPase associated with diverse cellular activities protein VPS4, harbor present in microtubule-interacting and trafficking molecules (MIT) domains comprising asymmetric three-helical bundles, which interact with helical MIT-interacting motifs in ESCRT-III subunits. Here we assess comprehensively the ESCRT-III interactions of the MIT-domain family member MITD1 and identify strong interactions with charged multivesicular body protein 1B (CHMP1B), CHMP2A, and increased sodium tolerance-1 (IST1). We show that these ESCRT-III subunits are important for the recruitment of MITD1 to the midbody and that MITD1 participates in the abscission phase of cytokinesis. MITD1 also dimerizes through its C-terminal domain. Both types of interactions appear important for the role of MITD1 in negatively regulating the interaction of IST1 with VPS4. Because IST1 binding in turn regulates VPS4, MITD1 may function through downstream effects on the activity of VPS4, which plays a critical role in the processing and remodeling of ESCRT filaments in abscission.

Structure and function of ESCRT-III

2009 ◽  
Vol 37 (1) ◽  
pp. 156-160 ◽  
Author(s):  
Suman Lata ◽  
Guy Schoehn ◽  
Julianna Solomons ◽  
Ricardo Pires ◽  
Heinrich G. Göttlinger ◽  
...  
Keyword(s):  
Multivesicular Body ◽  
Multivesicular Bodies ◽  
Tubular Structures ◽  
Body Protein ◽  
Enveloped Viruses ◽  
Endosomal Sorting ◽  
Vacuolar Protein ◽  
And Function ◽  
Endosomal Vesicles

ESCRT-III (endosomal sorting complex required for transport III) is required for the formation and abscission of intraluminal endosomal vesicles, which gives rise to multivesicular bodies, budding of some enveloped viruses and cytokinesis. ESCRT-III is composed of 11 members in humans, which, except for one, correspond to the six ESCRT-III-like proteins in yeast. At least CHMP (charged multivesicular body protein) 2A and CHMP3 assemble into helical tubular structures that provide a platform for membrane interaction and VPS (vacuolar protein sorting) 4-catalysed effects leading to disassembly of ESCRT-III CHMP2A–CHMP3 polymers in vitro. Progress towards the understanding of the structures and function of ESCRT-III, its activation, its regulation by accessory factors and its role in abscission of membrane enveloped structures in concert with VPS4 are discussed.

scholarly journals The penta-EF-hand protein ALG-2 interacts directly with the ESCRT-I component TSG101, and Ca2+-dependently co-localizes to aberrant endosomes with dominant-negative AAA ATPase SKD1/Vps4B

2005 ◽  
Vol 391 (3) ◽  
pp. 677-685 ◽  
Author(s):  
Keiichi Katoh ◽  
Hidenori Suzuki ◽  
Yoshinori Terasawa ◽  
Takako Mizuno ◽  
Jiro Yasuda ◽  
...  
Keyword(s):  
Hela Cells ◽  
Fluorescent Protein ◽  
Dominant Negative ◽  
Positive Interaction ◽  
Interacting Protein ◽  
Body Protein ◽  
Endosomal Sorting ◽  
Aaa Atpase ◽  
Ef Hand ◽  
Vacuolar Protein

ALG-2 (apoptosis-linked gene 2) is a Ca2+-binding protein that belongs to the PEF (penta-EF-hand) protein family. Alix (ALG-2-interacting protein X)/AIP1 (ALG-2-interacting protein 1), one of its binding partners, interacts with TSG101 and CHMP4 (charged multivesicular body protein 4), which are components of ESCRT-I (endosomal sorting complex required for transport I) and ESCRT-III respectively. In the present study, we investigated the association between ALG-2 and ESCRT-I. By a GST (glutathione S-transferase) pull-down assay using HEK-293T (human embryonic kidney 293T) cell lysates, endogenous TSG101 and two other exogenously expressed ESCRT-I components [hVps28 (human vacuolar protein sorting 28) and hVps37A] were shown to associate with GST–ALG-2 in the presence of Ca2+. By the yeast two-hybrid assay, however, a positive interaction was observed with only TSG101 among the three ESCRT-I components, suggesting that ALG-2 associates with hVps28 and hVps37A indirectly through TSG101. Using various deletion mutants of TSG101, the central PRR (proline-rich region) was found to be sufficient for interaction with ALG-2 by the GST-pull-down assay. Direct binding of ALG-2 to the TSG101 PRR was demonstrated by an overlay assay using biotin-labelled ALG-2 as a probe. In immunofluorescence microscopic analysis of HeLa cells that overexpressed a GFP (green fluorescent protein)-fused ATPase-defective dominant-negative form of SKD1/Vps4B (GFP–SKD1E235Q), ALG-2 exhibited a punctate distribution at the perinuclear area and co-localized with GFP–SKD1E235Q to aberrant endosomes. This punctate distribution of ALG-2 was markedly diminished by treatment of HeLa cells with a membrane-permeant Ca2+ chelator. Moreover, a Ca2+-binding-defective mutant of ALG-2 did not co-localize with GFP–SKD1E235Q. Our findings suggest that ALG-2 may function as a Ca2+-dependent accessory protein of the endosomal sorting machinery by interacting directly with TSG101 as well as with Alix.

scholarly journals A Duplicated ESCRT-III Factor Blocks Enveloped Virus Budding Without Inhibiting Cellular ESCRT Functions

2020 ◽  
Author(s):  
Lara Rheinemann ◽  
Diane Miller Downhour ◽  
Kate Bredbenner ◽  
Gaelle Mercenne ◽  
Kristen A. Davenport ◽  
...  
Keyword(s):  
Cellular Membrane ◽  
New World Monkeys ◽  
Virus Budding ◽  
Enveloped Viruses ◽  
Endosomal Sorting ◽  
Adaptive Mutations ◽  
Enveloped Virus ◽  
Viral Budding ◽  
Membrane Fission ◽  
Infected Cells

SummaryMany enveloped viruses require the endosomal sorting complexes required for transport (ESCRT) pathway to exit infected cells. This highly conserved pathway mediates essential cellular membrane fission events and therefore has limited potential to acquire adaptive mutations to counteract this co-option by viruses. Here, we describe duplicated and truncated copies of the ESCRT-III factor CHMP3 that arose independently in New World monkeys and mice and that block ESCRT-dependent virus budding. When expressed in human cells, these retroCHMP3 proteins potently inhibit the release of retroviruses, paramyxoviruses and filoviruses. RetroCHMP3 proteins have evolved to reduce interactions with other ESCRT-III factors, and to have little effect on cellular ESCRT processes, revealing routes for decoupling cellular ESCRT functions from exploitation by viruses. The repurposing of duplicated ESCRT-III proteins thus provides a mechanism to generate broad-spectrum viral budding inhibitors without disrupting highly conserved essential cellular ESCRT functions.

scholarly journals Dual roles of an Arabidopsis ESCRT component FREE1 in regulating vacuolar protein transport and autophagic degradation

2015 ◽  
Vol 112 (6) ◽  
pp. 1886-1891 ◽  
Author(s):  
Caiji Gao ◽  
Xiaohong Zhuang ◽  
Yong Cui ◽  
Xi Fu ◽  
Yilin He ◽  
...  
Keyword(s):  
Protein Trafficking ◽  
Protein Transport ◽  
Multivesicular Body ◽  
Organelle Biogenesis ◽  
Functional Roles ◽  
Endosomal Sorting ◽  
Escrt Machinery ◽  
Autophagic Degradation ◽  
Vacuolar Protein ◽  
Vacuole Biogenesis

Protein turnover can be achieved via the lysosome/vacuole and the autophagic degradation pathways. Evidence has accumulated revealing that efficient autophagic degradation requires functional endosomal sorting complex required for transport (ESCRT) machinery. However, the interplay between the ESCRT machinery and the autophagy regulator remains unclear. Here, we show that FYVE domain protein required for endosomal sorting 1 (FREE1), a recently identified plant-specific ESCRT component essential for multivesicular body (MVB) biogenesis and plant growth, plays roles both in vacuolar protein transport and autophagic degradation. FREE1 also regulates vacuole biogenesis in both seeds and vegetative cells of Arabidopsis. Additionally, FREE1 interacts directly with a unique plant autophagy regulator SH3 DOMAIN-CONTAINING PROTEIN2 and associates with the PI3K complex, to regulate the autophagic degradation in plants. Thus, FREE1 plays multiple functional roles in vacuolar protein trafficking and organelle biogenesis as well as in autophagic degradation via a previously unidentified regulatory mechanism of cross-talk between the ESCRT machinery and autophagy process.

scholarly journals A cryo–electron tomography workflow reveals protrusion-mediated shedding on injured plasma membrane

2021 ◽  
Vol 7 (13) ◽  
pp. eabc6345
Author(s):  
Shrawan Kumar Mageswaran ◽  
Wei Yuan Yang ◽  
Yogaditya Chakrabarty ◽  
Catherine M. Oikonomou ◽  
Grant J. Jensen
Keyword(s):  
Plasma Membrane ◽  
Molecular Mechanisms ◽  
Electron Tomography ◽  
Membrane Damage ◽  
Light And Electron Microscopy ◽  
Actin Dynamics ◽  
Endosomal Sorting ◽  
Plasma Membrane Damage ◽  
Cryo Electron Tomography ◽  
Vacuolar Protein

Cryo–electron tomography (cryo-ET) provides structural context to molecular mechanisms underlying biological processes. Although straightforward to implement for studying stable macromolecular complexes, using it to locate short-lived structures and events can be impractical. A combination of live-cell microscopy, correlative light and electron microscopy, and cryo-ET will alleviate this issue. We developed a workflow combining the three to study the ubiquitous and dynamic process of shedding in response to plasma membrane damage in HeLa cells. We found filopodia-like protrusions enriched at damage sites and acting as scaffolds for shedding, which involves F-actin dynamics, myosin-1a, and vacuolar protein sorting 4B (a component of the ‘endosomal sorting complex required for transport’ machinery). Overall, shedding is more complex than current models of vesiculation from flat membranes. Its similarities to constitutive shedding in enterocytes argue for a conserved mechanism. Our workflow can also be adapted to study other damage response pathways and dynamic cellular events.

scholarly journals The role of VPS4 in ESCRT-III polymer remodeling

2019 ◽  
Vol 47 (1) ◽  
pp. 441-448 ◽  
Author(s):  
Christophe Caillat ◽  
Sourav Maity ◽  
Nolwenn Miguet ◽  
Wouter H. Roos ◽  
Winfried Weissenhorn
Keyword(s):  
Active Role ◽  
Mechanistic Models ◽  
Membrane Remodeling ◽  
Filament Formation ◽  
Enveloped Viruses ◽  
Endosomal Sorting ◽  
Membrane Fission ◽  
Dynamic Assembly ◽  
Inside Out

Abstract The endosomal sorting complex required for transport-III (ESCRT-III) and VPS4 catalyze a variety of membrane-remodeling processes in eukaryotes and archaea. Common to these processes is the dynamic recruitment of ESCRT-III proteins from the cytosol to the inner face of a membrane neck structure, their activation and filament formation inside or at the membrane neck and the subsequent or concomitant recruitment of the AAA-type ATPase VPS4. The dynamic assembly of ESCRT-III filaments and VPS4 on cellular membranes induces constriction of membrane necks with large diameters such as the cytokinetic midbody and necks with small diameters such as those of intraluminal vesicles or enveloped viruses. The two processes seem to use different sets of ESCRT-III filaments. Constriction is then thought to set the stage for membrane fission. Here, we review recent progress in understanding the structural transitions of ESCRT-III proteins required for filament formation, the functional role of VPS4 in dynamic ESCRT-III assembly and its active role in filament constriction. The recent data will be discussed in the context of different mechanistic models for inside-out membrane fission.

scholarly journals Pathways of host cell exit by intracellular pathogens

2018 ◽  
Vol 5 (12) ◽  
pp. 525-544 ◽  
Author(s):  
Antje Flieger ◽  
Freddy Frischknecht ◽  
Georg Haecker ◽  
Mathias W. Hornef ◽  
Gabriele Pradel
Keyword(s):  
Host Cell ◽  
Intracellular Pathogens
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