scholarly journals A helical assembly of human ESCRT-I scaffolds reverse-topology membrane scission

2020 ◽  
Author(s):  
Thomas G. Flower ◽  
Yoshinori Takahashi ◽  
Arpa Hudait ◽  
Kevin Rose ◽  
Nicholas Tjahjono ◽  
...  
Keyword(s):  
Membered Ring ◽  
Coarse Grained ◽  
Virus Release ◽  
Cell Processes ◽  
Membrane Scission ◽  
Coarse Grained Simulations ◽  
Gag Assembly ◽  
Late Stages ◽  
Hiv 1

AbstractThe ESCRT complexes drive membrane scission in HIV-1 release, autophagosome closure, MVB biogenesis, cytokinesis, and other cell processes. ESCRT-I is the most upstream complex and bridges the system to HIV-1 Gag in virus release. The crystal structure of the headpiece of human ESCRT-I comprising TSG101:VPS28:VPS37B:MVB12A was determined, revealing an ESCRT-I helical assembly with a 12 molecule repeat. Electron microscopy confirmed that ESCRT-I subcomplexes form helical filaments in solution. Mutation of VPS28 helical interface residues blocks filament formation in vitro and autophagosome closure and HIV-1 release in human cells. Coarse grained simulations of ESCRT assembly at HIV-1 budding sites suggest that formation of a 12-membered ring of ESCRT-I molecules is a geometry-dependent checkpoint during late stages of Gag assembly and HIV-1 budding, and templates ESCRT-III assembly for membrane scission. These data show that ESCRT-I is not merely a bridging adaptor, but has an essential scaffolding and mechanical role in its own right.

scholarly journals The Conserved Tyr176/Leu177 Motif in the α-Helix 9 of the Feline Immunodeficiency Virus Capsid Protein Is Critical for Gag Particle Assembly

Viruses ◽  
2019 ◽  
Vol 11 (9) ◽  
pp. 816
Author(s):  
César A. Ovejero ◽  
Silvia A. González ◽  
José L. Affranchino
Keyword(s):  
Feline Immunodeficiency Virus ◽  
Particle Production ◽  
Equine Infectious Anemia Virus ◽  
Particle Assembly ◽  
Immunodeficiency Virus ◽  
Immature Particle ◽  
Gag Assembly ◽  
Hiv 1

The capsid domain (CA) of the lentiviral Gag polyproteins has two distinct roles during virion morphogenesis. As a domain of Gag, it mediates the Gag–Gag interactions that drive immature particle assembly, whereas as a mature protein, it self-assembles into the conical core of the mature virion. Lentiviral CA proteins are composed of an N-terminal region with seven α-helices and a C-terminal domain (CA-CTD) formed by four α-helices. Structural studies performed in HIV-1 indicate that the CA-CTD helix 9 establishes homodimeric interactions that contribute to the formation of the hexameric Gag lattice in immature virions. Interestingly, the mature CA core also shows inter-hexameric associations involving helix 9 residues W184 and M185. The CA proteins of feline immunodeficiency virus (FIV) and equine infectious anemia virus (EIAV) exhibit, at equivalent positions in helix 9, the motifs Y176/L177 and L169/F170, respectively. In this paper, we investigated the relevance of the Y176/L177 motif for FIV assembly by introducing a series of amino acid substitutions into this sequence and studying their effect on in vivo and in vitro Gag assembly, CA oligomerization, mature virion production, and viral infectivity. Our results demonstrate that the Y176/L177 motif in FIV CA helix 9 is essential for Gag assembly and CA oligomerization. Notably, mutations converting the FIV CA Y176/L177 motif into the HIV-1 WM and EIAV FL sequences allow substantial particle production and viral replication in feline cells.

scholarly journals Coarse-grained simulation reveals key features of HIV-1 capsid self-assembly

2016 ◽  
Author(s):  
John M A Grime ◽  
James F Dama ◽  
Barbie K Ganser-Pornillos ◽  
Cora L Woodward ◽  
Grant J Jensen ◽  
...  
Keyword(s):  
Self Assembly ◽  
Coarse Grained ◽  
Capsid Assembly ◽  
Molecular Crowding ◽  
Local Conditions ◽  
Conformational Variability ◽  
Multi Stage ◽  
Capsid Shell ◽  
Coarse Grained Simulations ◽  
Hiv 1

The maturation of HIV-1 viral particles is essential for viral infectivity. During maturation, many copies of the capsid protein (CA) self-assemble into a capsid shell to enclose the viral RNA. The mechanistic details of the initiation and early stages of capsid assembly remain to be delineated. We present coarse-grained simulations of capsid assembly under various conditions, considering not only capsid lattice self-assembly but also the potential disassembly of capsid upon delivery to the cytoplasm of a target cell. The effects of CA concentration, molecular crowding, and the conformational variability of CA are described, with results indicating that capsid nucleation and growth is a multi-stage process requiring well-defined metastable intermediates. Generation of the mature capsid lattice is sensitive to local conditions, with relatively subtle changes in CA concentration and molecular crowding influencing self-assembly and the ensemble of structural morphologies.

scholarly journals Actin crosslinker competition and sorting drive emergent GUV size-dependent actin network architecture

2021 ◽  
Vol 4 (1) ◽  
Author(s):  
Yashar Bashirzadeh ◽  
Steven A. Redford ◽  
Chatipat Lorpaiboon ◽  
Alessandro Groaz ◽  
Hossein Moghimianavval ◽  
...  
Keyword(s):  
Network Architecture ◽  
Molecular Size ◽  
Coarse Grained ◽  
General Mechanism ◽  
Complex Structures ◽  
Actin Bundles ◽  
Size Dependent ◽  
Apparent Stiffness ◽  
Coarse Grained Simulations

AbstractThe proteins that make up the actin cytoskeleton can self-assemble into a variety of structures. In vitro experiments and coarse-grained simulations have shown that the actin crosslinking proteins α-actinin and fascin segregate into distinct domains in single actin bundles with a molecular size-dependent competition-based mechanism. Here, by encapsulating actin, α-actinin, and fascin in giant unilamellar vesicles (GUVs), we show that physical confinement can cause these proteins to form much more complex structures, including rings and asters at GUV peripheries and centers; the prevalence of different structures depends on GUV size. Strikingly, we found that α-actinin and fascin self-sort into separate domains in the aster structures with actin bundles whose apparent stiffness depends on the ratio of the relative concentrations of α-actinin and fascin. The observed boundary-imposed effect on protein sorting may be a general mechanism for creating emergent structures in biopolymer networks with multiple crosslinkers.

scholarly journals Actin crosslinker competition and sorting drive emergent GUV size-dependent actin network architecture

2020 ◽  
Author(s):  
Yashar Bashirzadeh ◽  
Steven A. Redford ◽  
Chatipat Lorpaiboon ◽  
Alessandro Groaz ◽  
Thomas Litschel ◽  
...  
Keyword(s):  
Network Architecture ◽  
Coarse Grained ◽  
General Mechanism ◽  
Actin Networks ◽  
Cellular Processes ◽  
Size Dependent ◽  
Sensory Projections ◽  
Coarse Grained Simulations ◽  
Cytoskeletal Networks

AbstractRobust spatiotemporal organization of cytoskeletal networks is crucial, enabling cellular processes such as cell migration and division. α-Actinin and fascin are two actin crosslinking proteins localized to distinct regions of eukaryotes to form actin bundles with optimized spacing for cell contractile machinery and sensory projections, respectively. In vitro reconstitution assays and coarse-grained simulations have shown that these actin bundling proteins segregate into distinct domains with a bundler size-dependent competition-based mechanism, driven by the minimization of F-actin bending energy. However, it is not known how physical confinement imposed by the cell membrane contributes to sorting of actin bundling proteins and the concomitant reorganization of actin networks in intracellular environment. Here, by encapsulating actin, α-actinin, and fascin in giant unilamellar vesicles (GUVs), we show that the size of such a spherical boundary determines equilibrated structure of actin networks among three typical structures: single rings, astral structures, and star-like structures. We show that α-actinin bundling activity and its tendency for clustering actin is central to the formation of these structures. By analyzing physical features of crosslinked actin networks, we show that spontaneous sorting and domain formation of α-actinin and fascin are intimately linked to the resulting structures. We propose that the observed boundary-imposed effect on sorting and structure formation is a general mechanism by which cells can select between different structural dynamical steady states.

scholarly journals Coarse-Grained Simulations of the HIV-1 Matrix Protein Anchoring: Revisiting Its Assembly on Membrane Domains

2014 ◽  
Vol 106 (3) ◽  
pp. 577-585 ◽  
Author(s):  
Landry Charlier ◽  
Maxime Louet ◽  
Laurent Chaloin ◽  
Patrick Fuchs ◽  
Jean Martinez ◽  
...  
Keyword(s):  
Matrix Protein ◽  
Coarse Grained ◽  
Membrane Domains ◽  
Coarse Grained Simulations ◽  
Hiv 1

scholarly journals CryoET structures of immature HIV Gag reveal a complete six-helix bundle and stabilizing small molecules distinct from IP6

2020 ◽  
Author(s):  
Luiza Mendonça ◽  
Dapeng Sun ◽  
Jiying Ning ◽  
Jiwei Liu ◽  
Abhay Kotecha ◽  
...  
Keyword(s):  
Small Molecules ◽  
Structural Basis ◽  
Particle Assembly ◽  
Virus Maturation ◽  
Helix Bundle ◽  
Polyprotein Precursor ◽  
Subtomogram Averaging ◽  
Gag Assembly ◽  
Hiv 1

AbstractGag is the major HIV-1 structural polyprotein precursor. The Gag SP1 domain with the last residues of CA have been hypothesized to form a six-helix bundle necessary for particle assembly, but this bundle has not been fully resolved. Here, we determined the structures of complete CA-SP1 six-helix bundle connecting to the NC domain, from both in vitro Gag assemblies and viral-like particles (VLPs) carrying a T8I mutation in SP1, to near-atomic resolutions using cryoET and subtomogram averaging. The structures revealed novel densities, however distinct from IP6, inside the six-helix bundle of Gag assemblies, stabilizing the immature lattice. Interestingly, the T8I mutation impaired proteolytic cleavage of Gag at both SP1 boundaries. Our findings signify the involvement of small molecules in immature Gag assembly and provide the structural basis for development of small molecule inhibitors that stabilize SP1 helix, thus interfere with PR-mediated virus maturation.

scholarly journals Apoptosis Induced by Infection of Primary Brain Cultures with Diverse Human Immunodeficiency Virus Type 1 Isolates: Evidence for a Role of the Envelope

1999 ◽  
Vol 73 (2) ◽  
pp. 897-906 ◽  
Author(s):  
Asa Ohagen ◽  
Sajal Ghosh ◽  
Jianglin He ◽  
Karen Huang ◽  
Youzhi Chen ◽  
...  
Keyword(s):  
Replication Capacity ◽  
Immunodeficiency Virus ◽  
The Central Nervous System ◽  
Late Stages ◽  
The Brain ◽  
V3 Region ◽  
Hiv 1

ABSTRACT Apoptosis of neurons and astrocytes is induced by human immunodeficiency type 1 (HIV-1) infection in vitro and has been demonstrated in brain tissue from patients with AIDS. We analyzed a panel of diverse HIV-1 primary isolates for the ability to replicate and induce neuronal and astrocyte apoptosis in primary human brain cultures. Apoptosis was induced three- to eightfold by infection with the blood-derived HIV-1 isolates 89.6, SG3, and ADA. In contrast, the brain-derived HIV-1 isolates YU2, JRFL, DS-br, RC-br, and KJ-br did not induce significant levels of apoptosis. The ability of HIV-1 isolates to induce apoptosis was independent of their replication capacity. Studies of recombinant chimeras between the SG3 and YU2 viruses showed that replacement of the YU2 Env with the SG3 Env was sufficient to confer the ability to induce apoptosis to the YU2 virus. Replacement of the Env V3 regions alone largely conferred the phenotypes of the parental clones. The SG3 Env used CXCR4 and CCR3 as coreceptors for virus entry, whereas YU2 used CCR5 and CCR3. The V3 regions of SG3 and YU2 conferred the ability to use CXCR4 and CCR5, respectively. In contrast, the 3′ region of Env, particularly the C3V4 region, was required in conjunction with the V3 region for efficient use of CCR3. These results provide evidence that Env is a major determinant of neurodegenerative mechanisms associated with HIV-1 infection in vitro and raise the possibility that blood-derived viruses which emerge during the late stages of disease may affect disease progression in the central nervous system.

scholarly journals Molecular details of protein condensates probed by microsecond-long atomistic simulations

2020 ◽  
Author(s):  
Wenwei Zheng ◽  
Gregory L. Dignon ◽  
Xichen Xu ◽  
Roshan M. Regy ◽  
Nicolas L. Fawzi ◽  
...  
Keyword(s):  
Phase Separation ◽  
Intrinsically Disordered Proteins ◽  
Low Complexity ◽  
Coarse Grained ◽  
Disordered Proteins ◽  
Intrinsically Disordered ◽  
Residue Interaction ◽  
Coarse Grained Simulations ◽  
Protein Density

AbstractThe formation of membraneless organelles in cells commonly occurs via liquid-liquid phase separation (LLPS), and is in many cases driven by multivalent interactions between intrinsically disordered proteins (IDPs). Molecular simulations can reveal the specific amino acid interactions driving LLPS, which is hard to obtain from experiment. Coarse-grained simulations have been used to directly observe the sequence determinants of phase separation but have limited spatial resolution, while all-atom simulations have yet to be applied to LLPS due to the challenges of large system sizes and long time scales relevant to phase separation. We present a novel multiscale computational framework by obtaining initial molecular configurations of a condensed protein-rich phase from equilibrium coarse-grained simulations, and back mapping to an all-atom representation. Using the specialized Anton 2 supercomputer, we resolve microscopic structural and dynamical details of protein condensates through microsecond-scale all-atom explicit-solvent simulations. We have studied two IDPs which phase separate in vitro: the low complexity domain of FUS and the N-terminal disordered domain of LAF-1. Using this approach, we explain the partitioning of ions between phases with low and high protein density, demonstrate that the proteins are remarkably dynamic within the condensed phase, identify the key residue-residue interaction modes stabilizing the dense phase, all while showing good agreement with experimental observations. Our approach is generally applicable to all-atom studies of other single and multi-component systems of proteins and nucleic acids involved in the formation of membraneless organelles.

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