scholarly journals Visualization and Sequencing of Membrane Remodeling Leading to Influenza Virus Fusion

2016 ◽  
Vol 90 (15) ◽  
pp. 6948-6962 ◽  
Author(s):  
Long Gui ◽  
Jamie L. Ebner ◽  
Alexander Mileant ◽  
James A. Williams ◽  
Kelly K. Lee
Keyword(s):  
Influenza Virus ◽  
Membrane Fusion ◽  
Electron Tomography ◽  
Membrane Remodeling ◽  
Enveloped Virus ◽  
Virus Fusion ◽  
Cryo Electron Tomography ◽  
Membrane Reorganization ◽  
Target Membrane ◽  
Over Time

ABSTRACTProtein-mediated membrane fusion is an essential step in many fundamental biological events, including enveloped virus infection. The nature of protein and membrane intermediates and the sequence of membrane remodeling during these essential processes remain poorly understood. Here we used cryo-electron tomography (cryo-ET) to image the interplay between influenza virus and vesicles with a range of lipid compositions. By following the population kinetics of membrane fusion intermediates imaged by cryo-ET, we found that membrane remodeling commenced with the hemagglutinin fusion protein spikes grappling onto the target membrane, followed by localized target membrane dimpling as local clusters of hemagglutinin started to undergo conformational refolding. The local dimples then transitioned to extended, tightly apposed contact zones where the two proximal membrane leaflets were in most cases indistinguishable from each other, suggesting significant dehydration and possible intermingling of the lipid head groups. Increasing the content of fusion-enhancing cholesterol or bis-monoacylglycerophosphate in the target membrane led to an increase in extended contact zone formation. Interestingly, hemifused intermediates were found to be extremely rare in the influenza virus fusion system studied here, most likely reflecting the instability of this state and its rapid conversion to postfusion complexes, which increased in population over time. By tracking the populations of fusion complexes over time, the architecture and sequence of membrane reorganization leading to efficient enveloped virus fusion were thus resolved.IMPORTANCEEnveloped viruses employ specialized surface proteins to mediate fusion of cellular and viral membranes that results in the formation of pores through which the viral genetic material is delivered to the cell. For influenza virus, the trimeric hemagglutinin (HA) glycoprotein spike mediates host cell attachment and membrane fusion. While structures of a subset of conformations and parts of the fusion machinery have been characterized, the nature and sequence of membrane deformations during fusion have largely eluded characterization. Building upon studies that focused on early stages of HA-mediated membrane remodeling, here cryo-electron tomography (cryo-ET) was used to image the three-dimensional organization of intact influenza virions at different stages of fusion with liposomes, leading all the way to completion of the fusion reaction. By monitoring the evolution of fusion intermediate populations over the course of acid-induced fusion, we identified the progression of membrane reorganization that leads to efficient fusion by an enveloped virus.

scholarly journals Functions of the Stem Region of the Semliki Forest Virus Fusion Protein during Virus Fusion and Assembly

2006 ◽  
Vol 80 (22) ◽  
pp. 11362-11369 ◽  
Author(s):  
Maofu Liao ◽  
Margaret Kielian
Keyword(s):  
Fusion Protein ◽  
Membrane Fusion ◽  
Semliki Forest Virus ◽  
Virus Assembly ◽  
Protein A ◽  
Stem Length ◽  
Membrane Fusion Protein ◽  
Virus Fusion ◽  
Target Membrane ◽  
Domain Iii

ABSTRACT Membrane fusion of the alphaviruses is mediated by the E1 protein, a class II virus membrane fusion protein. During fusion, E1 dissociates from its heterodimer interaction with the E2 protein and forms a target membrane-inserted E1 homotrimer. The structure of the homotrimer is that of a trimeric hairpin in which E1 domain III and the stem region fold back toward the target membrane-inserted fusion peptide loop. The E1 stem region has a strictly conserved length and several highly conserved residues, suggesting the possibility of specific stem interactions along the trimer core and an important role in driving membrane fusion. Mutagenesis studies of the alphavirus Semliki Forest virus (SFV) here demonstrated that there was a strong requirement for the E1 stem in virus assembly and budding, probably reflecting its importance in lateral interactions of the envelope proteins. Surprisingly, however, neither the conserved length nor any specific residues of the stem were required for membrane fusion. Although the highest fusion activity was observed with wild-type E1, efficient fusion was mediated by stem mutants containing a variety of substitutions or deletions. A minimal stem length was required but could be conferred by a series of alanine residues. The lack of a specific stem sequence requirement during SFV fusion suggests that the interaction of domain III with the trimer core can provide sufficient driving force to mediate membrane merger.

scholarly journals Membrane Structures of the Hemifusion-Inducing Fusion Peptide Mutant G1S and theFusion-Blocking Mutant G1V of Influenza Virus HemagglutininSuggest a Mechanism for Pore Opening in MembraneFusion

2005 ◽  
Vol 79 (18) ◽  
pp. 12065-12076 ◽  
Author(s):  
Yinling Li ◽  
Xing Han ◽  
Alex L. Lai ◽  
John H. Bushweller ◽  
David S. Cafiso ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Membrane Fusion ◽  
Lipid Bilayer ◽  
Membrane Surface ◽  
Helical Structure ◽  
Fusion Peptide ◽  
Membrane Structures ◽  
Influenza Virus Hemagglutinin ◽  
Target Membrane ◽  
Pore Opening

ABSTRACT Influenza virus hemagglutinin (HA)-mediated membrane fusion is initiated by a conformational change that releases a V-shaped hydrophobic fusion domain, the fusion peptide, into the lipid bilayer of the target membrane. The most N-terminal residue of this domain, a glycine, is highly conserved and is particularly critical for HA function; G1S and G1V mutant HAs cause hemifusion and abolish fusion, respectively. We have determined the atomic resolution structures of the G1S and G1V mutant fusion domains in membrane environments. G1S forms a V with a disrupted “glycine edge” on its N-terminal arm and G1V adopts a slightly tilted linear helical structure in membranes. Abolishment of the kink in G1V results in reduced hydrophobic penetration of the lipid bilayer and an increased propensity to formβ -structures at the membrane surface. These results underline the functional importance of the kink in the fusion peptide and suggest a structural role for the N-terminal glycine ridge in viral membrane fusion.

scholarly journals 9Å structure of the COPI coat reveals that the Arf1 GTPase occupies two contrasting molecular environments

eLife ◽  
2017 ◽  
Vol 6 ◽  
Author(s):  
Svetlana O Dodonova ◽  
Patrick Aderhold ◽  
Juergen Kopp ◽  
Iva Ganeva ◽  
Simone Röhling ◽  
...  
Keyword(s):  
Electron Tomography ◽  
Molecular Model ◽  
Adaptor Proteins ◽  
Small Gtpase ◽  
Coated Vesicle ◽  
Cryo Electron Tomography ◽  
Target Membrane ◽  
Clathrin Adaptor ◽  
Subtomogram Averaging

COPI coated vesicles mediate trafficking within the Golgi apparatus and between the Golgi and the endoplasmic reticulum. Assembly of a COPI coated vesicle is initiated by the small GTPase Arf1 that recruits the coatomer complex to the membrane, triggering polymerization and budding. The vesicle uncoats before fusion with a target membrane. Coat components are structurally conserved between COPI and clathrin/adaptor proteins. Using cryo-electron tomography and subtomogram averaging, we determined the structure of the COPI coat assembled on membranes in vitro at 9 Å resolution. We also obtained a 2.57 Å resolution crystal structure of βδ-COP. By combining these structures we built a molecular model of the coat. We additionally determined the coat structure in the presence of ArfGAP proteins that regulate coat dissociation. We found that Arf1 occupies contrasting molecular environments within the coat, leading us to hypothesize that some Arf1 molecules may regulate vesicle assembly while others regulate coat disassembly.

scholarly journals Cryo-electron tomography reveals structural insights into the membrane binding and remodeling activity of dynamin-like EHDs

2021 ◽  
Author(s):  
Arthur A. Melo ◽  
Thiemo Sprink ◽  
Jeffrey K. Noel ◽  
Elena Vázquez Sarandeses ◽  
Chris van Hoorn ◽  
...  
Keyword(s):  
Structural Information ◽  
Electron Tomography ◽  
Spontaneous Curvature ◽  
Membrane Remodeling ◽  
Closed Conformation ◽  
Membrane Surfaces ◽  
Cryo Electron Tomography ◽  
Membrane Bound ◽  
Subtomogram Averaging ◽  
Structural Insights

AbstractDynamin-related Eps15-homology domain containing proteins (EHDs) oligomerize on membrane surfaces into filaments leading to membrane remodeling. EHD crystal structures in an open and a closed conformation were previously reported, but structural information on the membrane-bound EHD oligomeric structure has remained enigmatic. Consequently, mechanistic insight into EHD-mediated membrane remodeling is lacking. Here, by using cryo-electron tomography and subtomogram averaging, we determined the structure of an EHD4 filament on a tubular membrane template at an average resolution of 7.6 Å. Assembly of EHD4 is mediated via interfaces in the G-domain and the helical domain. The oligomerized EHD4 structure resembles the closed conformation, where the tips of the helical domains protrude into the membrane. The variation in filament geometry and tube radius suggests the AMPPNP-bound filament has a spontaneous curvature of approximately 1/70 nm-1. Combining the available structural and functional data, we propose a model of EHD-mediated membrane remodeling.

Characterization of influenza virus PR8 strain cultured in embryonated eggs by cryo-electron tomography

2019 ◽  
Vol 516 (1) ◽  
pp. 57-62 ◽  
Author(s):  
Qiang Chen ◽  
Xinrui Huang ◽  
Risheng Wei ◽  
Lei Zhang ◽  
Changcheng Yin
Keyword(s):  
Influenza Virus ◽  
Electron Tomography ◽  
Cryo Electron Tomography

scholarly journals HIV fusion: Catch me if you can

2020 ◽  
Vol 295 (45) ◽  
pp. 15196-15197
Author(s):  
Solène Denolly ◽  
François-Loïc Cosset
Keyword(s):  
Cell Membrane ◽  
Membrane Fusion ◽  
Lipid Membrane ◽  
Electron Tomography ◽  
Fusion Process ◽  
Target Cells ◽  
Tirf Microscopy ◽  
Enveloped Viruses ◽  
Cryo Electron Tomography

The penetration of enveloped viruses into target cells requires the fusion of the lipid envelope of their virions with the host lipid membrane though a stepwise and highly sophisticated process. However, the intermediate steps in this process have seldom been visualized due to their rarity and rapidity. Here, using cryo-electron tomography, TIRF microscopy, and cell membrane–derived vesicles called blebs, Ward et al. visualize intermediates of the HIV-cell membrane fusion process and demonstrate how Serinc proteins prevent full fusion by interfering with this process.

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