scholarly journals The Large Extracellular Loop of CD63 Interacts with gp41 of HIV-1 and is Essential for Establishing the Virological Synapse

2020 ◽  
Author(s):  
Daniel Ivanusic ◽  
Kazimierz Madela ◽  
Norbert Bannert ◽  
Joachim Denner
Keyword(s):  
Molecular Mechanisms ◽  
Core Protein ◽  
Biological Significance ◽  
Virological Synapse ◽  
Extracellular Loop ◽  
Protein Gp41 ◽  
Large Extracellular Loop ◽  
Cell Spread ◽  
Hiv 1 ◽  
Cell Cell

Abstract Human immunodeficiency virus type 1 (HIV-1) persists lifelong in infected individuals and has evolved unique strategies in order to evade the immune system. One of these strategies is the direct cell-to-cell spread of HIV-1. The formation of a virological synapse (VS) between donor and target cell is important for this process. Tetraspanins are cellular proteins that are actively involved in the formation of a VS. However, the molecular mechanisms of recruiting host proteins for the cell-cell transfer of particles to the VS remains unclear. Our study has mapped the binding site for the transmembrane envelope protein gp41 of HIV-1 within the large extracellular loop (LEL) of CD63 and showed that this interaction occurs predominantly at the VS between T cells where viral particles are transferred. Mutations within the highly conserved CCG motif of the tetraspanin superfamily abrogated recruiting of expressed HIV-1 GFP fused Gag core protein and CD63 to the VS. This demonstrates the biological significance of CD63 for enhanced formation of a VS. Since cell-cell spread of HIV-1 is a major route of persistent infection, these results highlight the central role of CD63 as a member of the tetraspanin superfamily during HIV-1 infection and pathogenesis.

scholarly journals The large extracellular loop of CD63 interacts with gp41 of HIV-1 and is essential for establishing the virological synapse

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Daniel Ivanusic ◽  
Kazimierz Madela ◽  
Norbert Bannert ◽  
Joachim Denner
Keyword(s):  
Molecular Mechanisms ◽  
Core Protein ◽  
Biological Significance ◽  
Virological Synapse ◽  
Extracellular Loop ◽  
Protein Gp41 ◽  
Large Extracellular Loop ◽  
Cell Spread ◽  
Hiv 1 ◽  
Cell Cell

AbstractHuman immunodeficiency virus type 1 (HIV-1) persists lifelong in infected individuals and has evolved unique strategies in order to evade the immune system. One of these strategies is the direct cell-to-cell spread of HIV-1. The formation of a virological synapse (VS) between donor and target cell is important for this process. Tetraspanins are cellular proteins that are actively involved in the formation of a VS. However, the molecular mechanisms of recruiting host proteins for the cell–cell transfer of particles to the VS remains unclear. Our study has mapped the binding site for the transmembrane envelope protein gp41 of HIV-1 within the large extracellular loop (LEL) of CD63 and showed that this interaction occurs predominantly at the VS between T cells where viral particles are transferred. Mutations within the highly conserved CCG motif of the tetraspanin superfamily abrogated recruiting of expressed HIV-1 GFP fused Gag core protein and CD63 to the VS. This demonstrates the biological significance of CD63 for enhanced formation of a VS. Since cell–cell spread of HIV-1 is a major route of persistent infection, these results highlight the central role of CD63 as a member of the tetraspanin superfamily during HIV-1 infection and pathogenesis.

scholarly journals Contact-Induced Mitochondrial Polarization Supports HIV-1 Virological Synapse Formation

2014 ◽  
Vol 89 (1) ◽  
pp. 14-24 ◽  
Author(s):  
Elisabetta Groppelli ◽  
Shimona Starling ◽  
Clare Jolly
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Lymphocytes ◽  
Virological Synapse ◽  
Cell Contact ◽  
Target Cells ◽  
Cell Contacts ◽  
Cell Spread ◽  
Hiv 1 ◽  
Cell Cell

ABSTRACTRapid HIV-1 spread between CD4 T lymphocytes occurs at retrovirus-induced immune cell contacts called virological synapses (VS). VS are associated with striking T cell polarization and localized virus budding at the site of contact that facilitates cell-cell spread. In addition to this, spatial clustering of organelles, including mitochondria, to the contact zone has been previously shown. However, whether cell-cell contact specifically induces dynamic T cell remodeling during VS formation and what regulates this process remain unclear. Here, we report that contact between an HIV-1-infected T cell and an uninfected target T cell specifically triggers polarization of mitochondria concomitant with recruitment of the major HIV-1 structural protein Gag to the site of cell-cell contact. Using fixed and live-cell imaging, we show that mitochondrial and Gag polarization in HIV-1-infected T cells occurs within minutes of contact with target T cells, requires the formation of stable cell-cell contacts, and is an active, calcium-dependent process. We also find that perturbation of mitochondrial polarization impairs cell-cell spread of HIV-1 at the VS. Taken together, these data suggest that HIV-1-infected T cells are able to sense and respond to contact with susceptible target cells and undergo dynamic cytoplasmic remodeling to create a synaptic environment that supports efficient HIV-1 VS formation between CD4 T lymphocytes.IMPORTANCEHIV-1 remains one of the major global health challenges of modern times. The capacity of HIV-1 to cause disease depends on the virus's ability to spread between immune cells, most notably CD4 T lymphocytes. Cell-cell transmission is the most efficient way of HIV-1 spread and occurs at the virological synapse (VS). The VS forms at the site of contact between an infected cell and an uninfected cell and is characterized by polarized assembly and budding of virions and clustering of cellular organelles, including mitochondria. Here, we show that cell-cell contact induces rapid recruitment of mitochondria to the contact site and that this supports efficient VS formation and consequently cell-cell spread. Additionally, we observed that cell-cell contact induces a mitochondrion-dependent increase in intracellular calcium, indicative of cellular signaling. Taken together, our data suggest that VS formation is a regulated process and thus a potential target to block HIV-1 cell-cell spread.

scholarly journals Requirement for an Intact T-Cell Actin and Tubulin Cytoskeleton for Efficient Assembly and Spread of Human Immunodeficiency Virus Type 1

2007 ◽  
Vol 81 (11) ◽  
pp. 5547-5560 ◽  
Author(s):  
Clare Jolly ◽  
Ivonne Mitar ◽  
Quentin J. Sattentau
Keyword(s):  
Human Immunodeficiency Virus ◽  
T Cells ◽  
Plasma Membrane ◽  
T Cell ◽  
Immunodeficiency Virus ◽  
Fixed Cell ◽  
Cell Spread ◽  
Hiv 1 ◽  
Cell Cell

ABSTRACT Human immunodeficiency virus type 1 (HIV-1) infection of CD4+ T cells leads to the production of new virions that assemble at the plasma membrane. Gag and Env accumulate in the context of lipid rafts at the inner and outer leaflets of the plasma membrane, respectively, forming polarized domains from which HIV-1 buds. HIV-1 budding can result in either release of cell-free virions or direct cell-cell spread via a virological synapse (VS). The recruitment of Gag and Env to these plasma membrane caps in T cells is poorly understood but may require elements of the T-cell secretory apparatus coordinated by the cytoskeleton. Using fixed-cell immunofluorescence labeling and confocal microscopy, we observed a high percentage of HIV-1-infected T cells with polarized Env and Gag in capped, lipid raft-like assembly domains. Treatment of infected T cells with inhibitors of actin or tubulin remodeling disrupted Gag and Env compartmentalization within the polarized raft-like domains. Depolymerization of the actin cytoskeleton reduced Gag release and viral infectivity, and actin and tubulin inhibitors reduced Env incorporation into virions. Live- and fixed-cell confocal imaging and assay of de novo DNA synthesis by real-time PCR allowed quantification of HIV-1 cell-cell transfer. Inhibition of actin and tubulin remodeling in infected cells interfered with cell-cell spread across a VS and reduced new viral DNA synthesis. Based on these data, we propose that HIV-1 requires both actin and tubulin components of the T-cell cytoskeleton to direct its assembly and budding and to elaborate a functional VS.

scholarly journals The envelope cytoplasmic tail regulates HIV-1 assembly and spread

2021 ◽  
Author(s):  
Xenia Snetkov ◽  
Tafhima Haider ◽  
Dejan Mesner ◽  
Nicholas Groves ◽  
Schuyler van Engelenburg ◽  
...  
Keyword(s):  
T Cells ◽  
Structural Integrity ◽  
Alpha Helix ◽  
Cytoplasmic Tail ◽  
Virological Synapse ◽  
Cell Contact ◽  
Viral Fusion ◽  
And Function ◽  
Hiv 1 ◽  
Cell Cell

AbstractThe HIV-1 envelope (Env) is an essential determinant of viral infectivity, tropism and spread between T cells. Lentiviral Env contain an unusually long 150 amino acid cytoplasmic tail (EnvCT) but the function of the EnvCT and conserved domains within it remain largely uncharacterised. Here we identified a highly conserved tryptophan motif at position 757 (W757) in the LLP-2 alpha helix of the EnvCT as a key determinant for HIV-1 replication and spread between T cells. Strikingly we find that mutating W757 had wide-ranging consequences including altering Env mobility in the plasma membrane, preventing Env and Gag recruitment to sites of cell-cell contact for virological synapse (VS) formation and cell-cell spread, and impeding viral fusion. Notably, W757 was also required for efficient virus budding, revealing a previously unappreciated role for the EnvCT in regulating HIV-1 assembly and egress. We conclude that W757 is a key residue that stabilises the structural integrity and function of Env, consistent with the recent model that this region of the EnvCT acts as a critical supporting baseplate for Env.

scholarly journals A Conserved Tryptophan in the Envelope Cytoplasmic Tail Regulates HIV-1 Assembly and Spread

Viruses ◽  
2022 ◽  
Vol 14 (1) ◽  
pp. 129
Author(s):  
Xenia Snetkov ◽  
Tafhima Haider ◽  
Dejan Mesner ◽  
Nicholas Groves ◽  
Schuyler B. van Engelenburg ◽  
...  
Keyword(s):  
T Cells ◽  
Single Molecule ◽  
Virus Assembly ◽  
Alpha Helix ◽  
Cytoplasmic Tail ◽  
Virological Synapse ◽  
Cell Contact ◽  
Viral Spread ◽  
Hiv 1 ◽  
Cell Cell

The HIV-1 envelope (Env) is an essential determinant of viral infectivity, tropism and spread between T cells. Lentiviral Env contain an unusually long 150 amino acid cytoplasmic tail (EnvCT), but the function of the EnvCT and many conserved domains within it remain largely uncharacterised. Here, we identified a highly conserved tryptophan motif at position 757 (W757) in the LLP-2 alpha helix of the EnvCT as a key determinant for HIV-1 replication and spread between T cells. Alanine substitution at this position potently inhibited HIV-1 cell–cell spread (the dominant mode of HIV-1 dissemination) by preventing recruitment of Env and Gag to sites of cell–cell contact, inhibiting virological synapse (VS) formation and spreading infection. Single-molecule tracking and super-resolution imaging showed that mutation of W757 dysregulates Env diffusion in the plasma membrane and increases Env mobility. Further analysis of Env function revealed that W757 is also required for Env fusion and infectivity, which together with reduced VS formation, result in a potent defect in viral spread. Notably, W757 lies within a region of the EnvCT recently shown to act as a supporting baseplate for Env. Our data support a model in which W757 plays a key role in regulating Env biology, modulating its temporal and spatial recruitment to virus assembly sites and regulating the inherent fusogenicity of the Env ectodomain, thereby supporting efficient HIV-1 replication and spread.

Virological Synapse for Cell-Cell Spread of Viruses

2006 ◽  
pp. 288-297
Author(s):  
Eduardo Garcia ◽  
Vincent Piguet
Keyword(s):  
Virological Synapse ◽  
Cell Spread ◽  
Cell Cell

Nef does not inhibit F-actin remodelling and HIV-1 cell–cell transmission at the T lymphocyte virological synapse

2011 ◽  
Vol 90 (11) ◽  
pp. 913-921 ◽  
Author(s):  
Claudia Haller ◽  
Nadine Tibroni ◽  
Jochen M. Rudolph ◽  
Robert Grosse ◽  
Oliver T. Fackler
Keyword(s):  
T Lymphocyte ◽  
Virological Synapse ◽  
Cell Transmission ◽  
Actin Remodelling ◽  
Hiv 1 ◽  
Cell Cell

scholarly journals EWI-2 Inhibits Cell–Cell Fusion at the HIV-1 Virological Presynapse

Viruses ◽  
2019 ◽  
Vol 11 (12) ◽  
pp. 1082
Author(s):  
Emily E. Whitaker ◽  
Nicholas J. Matheson ◽  
Sarah Perlee ◽  
Phillip B. Munson ◽  
Menelaos Symeonides ◽  
...  
Keyword(s):  
Cell Fusion ◽  
Virological Synapse ◽  
Virus Particles ◽  
Fusion Inhibitors ◽  
Producer Cell ◽  
Cellular Components ◽  
Cell To Cell Transfer ◽  
Hiv 1 ◽  
Cell Cell

Cell-to-cell transfer of virus particles at the Env-dependent virological synapse (VS) is a highly efficient mode of HIV-1 transmission. While cell–cell fusion could be triggered at the VS, leading to the formation of syncytia and preventing exponential growth of the infected cell population, this is strongly inhibited by both viral (Gag) and host (ezrin and tetraspanins) proteins. Here, we identify EWI-2, a protein that was previously shown to associate with ezrin and tetraspanins, as a host factor that contributes to the inhibition of Env-mediated cell–cell fusion. Using quantitative fluorescence microscopy, shRNA knockdowns, and cell–cell fusion assays, we show that EWI-2 accumulates at the presynaptic terminal (i.e., the producer cell side of the VS), where it contributes to the fusion-preventing activities of the other viral and cellular components. We also find that EWI-2, like tetraspanins, is downregulated upon HIV-1 infection, most likely by Vpu. Despite the strong inhibition of fusion at the VS, T cell-based syncytia do form in vivo and in physiologically relevant culture systems, but they remain small. In regard to that, we demonstrate that EWI-2 and CD81 levels are restored on the surface of syncytia, where they (presumably) continue to act as fusion inhibitors. This study documents a new role for EWI-2 as an inhibitor of HIV-1-induced cell–cell fusion and provides novel insight into how syncytia are prevented from fusing indefinitely.

scholarly journals Macrophage Cell-Cell Interactions Promoting HIV-1 Infection

Viruses ◽  
2020 ◽  
Vol 12 (5) ◽  
pp. 492 ◽  
Author(s):  
Maeva Dupont ◽  
Quentin James Sattentau
Keyword(s):  
T Cells ◽  
Viral Entry ◽  
Macrophage Infection ◽  
Macrophage Cell ◽  
Current State ◽  
Direct Cell ◽  
Cell Spread ◽  
Hiv 1 ◽  
Cell Cell

Many pathogens infect macrophages as part of their intracellular life cycle. This is particularly true for viruses, of which HIV-1 is one of the best studied. HIV-1 infection of macrophages has important consequences for viral persistence and pathogenesis, but the mechanisms of macrophage infection remain to be fully elucidated. Despite expressing viral entry receptors, macrophages are inefficiently infected by cell-free HIV-1 virions, whereas direct cell-cell spread is more efficient. Different modes of cell-cell spread have been described, including the uptake by macrophages of infected T cells and the fusion of infected T cells with macrophages, both leading to macrophage infection. Cell-cell spread can also transmit HIV-1 between macrophages and from macrophages to T cells. Here, we describe the current state of the field concerning the cell-cell spread of HIV-1 to and from macrophages, discuss mechanisms, and highlight potential in vivo relevance.

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