Cholesterol suppresses membrane leakage by decreasing water penetrability

Soft Matter ◽  
2018 ◽  
Vol 14 (25) ◽  
pp. 5277-5282 ◽  
Author(s):  
Bing Bu ◽  
Michael Crowe ◽  
Jiajie Diao ◽  
Baohua Ji ◽  
Dechang Li
Keyword(s):  
Membrane Fusion ◽  
In Silico ◽  
Pore Formation ◽  
Membrane Leakage

In silico simulations and biochemical experiments show that cholesterol decreases water penetrability to inhibit leakage pore formation during membrane fusion.

Mechanics of membrane fusion/pore formation

2015 ◽  
Vol 185 ◽  
pp. 109-128 ◽  
Author(s):  
Marc Fuhrmans ◽  
Giovanni Marelli ◽  
Yuliya G. Smirnova ◽  
Marcus Müller
Keyword(s):  
Membrane Fusion ◽  
Pore Formation ◽  
Fusion Pore

scholarly journals Restricted movement of lipid and aqueous dyes through pores formed by influenza hemagglutinin during cell fusion.

1994 ◽  
Vol 127 (6) ◽  
pp. 1885-1894 ◽  
Author(s):  
J Zimmerberg ◽  
R Blumenthal ◽  
D P Sarkar ◽  
M Curran ◽  
S J Morris
Keyword(s):  
Cell Membrane ◽  
Membrane Fusion ◽  
Cell Fusion ◽  
Pore Formation ◽  
Fusion Pore ◽  
Restricted Movement ◽  
Influenza Hemagglutinin ◽  
Simultaneous Measurements ◽  
Cell Membrane Capacitance ◽  
Total Fusion

The fusion of cells by influenza hemagglutinin (HA) is the best characterized example of protein-mediated membrane fusion. In simultaneous measurements of pairs of assays for fusion, we determined the order of detectable events during fusion. Fusion pore formation in HA-triggered cell-cell fusion was first detected by changes in cell membrane capacitance, next by a flux of fluorescent lipid, and finally by flux of aqueous fluorescent dye. Fusion pore conductance increased by small steps. A retardation of lipid and aqueous dyes occurred during fusion pore fluctuations. The flux of aqueous dye depended on the size of the molecule. The lack of movement of aqueous dyes while total fusion pore conductance increased suggests that initial HA-triggered fusion events are characterized by the opening of multiple small pores: the formation of a "sieve".

scholarly journals Exportins can inhibit major mitotic assembly events in vitro: membrane fusion, nuclear pore formation, and spindle assembly

Nucleus ◽  
2020 ◽  
Vol 11 (1) ◽  
pp. 178-193
Author(s):  
Matthew S. Nord ◽  
Cyril Bernis ◽  
Sarah Carmona ◽  
Dennis C. Garland ◽  
Anna Travesa ◽  
...  
Keyword(s):  
Membrane Fusion ◽  
Pore Formation ◽  
Spindle Assembly ◽  
Nuclear Pore

scholarly journals Multifaceted Sequence-Dependent and -Independent Roles for Reovirus FAST Protein Cytoplasmic Tails in Fusion Pore Formation and Syncytiogenesis

2009 ◽  
Vol 83 (23) ◽  
pp. 12185-12195 ◽  
Author(s):  
Christopher Barry ◽  
Roy Duncan
Keyword(s):  
Membrane Fusion ◽  
Fusion Proteins ◽  
Pore Formation ◽  
Syncytium Formation ◽  
Fusion Pore ◽  
Protein Fusion ◽  
Sequence Dependent ◽  
Cytoplasmic Tails ◽  
Fast Proteins ◽  
Pore Expansion

ABSTRACT Fusogenic reoviruses utilize the FAST proteins, a novel family of nonstructural viral membrane fusion proteins, to induce cell-cell fusion and syncytium formation. Unlike the paradigmatic enveloped virus fusion proteins, the FAST proteins position the majority of their mass within and internal to the membrane in which they reside, resulting in extended C-terminal cytoplasmic tails (CTs). Using tail truncations, we demonstrate that the last 8 residues of the 36-residue CT of the avian reovirus p10 FAST protein and the last 20 residues of the 68-residue CT of the reptilian reovirus p14 FAST protein enhance, but are not required for, pore expansion and syncytium formation. Further truncations indicate that the membrane-distal 12 residues of the p10 and 47 residues of the p14 CTs are essential for pore formation and that a residual tail of 21 to 24 residues that includes a conserved, membrane-proximal polybasic region present in all FAST proteins is insufficient to maintain FAST protein fusion activity. Unexpectedly, a reextension of the tail-truncated, nonfusogenic p10 and p14 constructs with scrambled versions of the deleted sequences restored pore formation and syncytiogenesis, while reextensions with heterologous sequences partially restored pore formation but failed to rescue syncytiogenesis. The membrane-distal regions of the FAST protein CTs therefore exert multiple effects on the membrane fusion reaction, serving in both sequence-dependent and sequence-independent manners as positive effectors of pore formation, pore expansion, and syncytiogenesis.

scholarly journals Identification and Characterization of Influenza Virus Entry Inhibitors through Dual Myxovirus High-Throughput Screening

2016 ◽  
Vol 90 (16) ◽  
pp. 7368-7387 ◽  
Author(s):  
Marco Weisshaar ◽  
Robert Cox ◽  
Zachary Morehouse ◽  
Shiva Kumar Kyasa ◽  
Dan Yan ◽  
...  
Keyword(s):  
Membrane Fusion ◽  
Small Molecule ◽  
High Throughput ◽  
In Silico ◽  
High Throughput Screening ◽  
Target Identification ◽  
Viral Escape ◽  
Reporter Strain ◽  
Secondary Resistance ◽  
Developmental Potential

ABSTRACTInfluenza A virus (IAV) infections cause major morbidity and mortality, generating an urgent need for novel antiviral therapeutics. We recently established a dual myxovirus high-throughput screening protocol that combines a fully replication-competent IAV-WSN strain and a respiratory syncytial virus reporter strain for the simultaneous identification of IAV-specific, paramyxovirus-specific, and broad-spectrum inhibitors. In the present study, this protocol was applied to a screening campaign to assess a diverse chemical library with over 142,000 entries. Focusing on IAV-specific hits, we obtained a hit rate of 0.03% after cytotoxicity testing and counterscreening. Three chemically distinct hit classes with nanomolar potency and favorable cytotoxicity profiles were selected. Time-of-addition, minigenome, and viral entry studies demonstrated that these classes block hemagglutinin (HA)-mediated membrane fusion. Antiviral activity extends to an isolate from the 2009 pandemic and, in one case, another group 1 subtype. Target identification through biolayer interferometry confirmed binding of all hit compounds to HA. Resistance profiling revealed two distinct escape mechanisms: primary resistance, associated with reduced compound binding, and secondary resistance, associated with unaltered binding. Secondary resistance was mediated, unusually, through two different pairs of cooperative mutations, each combining a mutation eliminating the membrane-proximal stalk N-glycan with a membrane-distal change in HA1 or HA2. Chemical synthesis of an analog library combined within silicodocking extracted a docking pose for the hit classes. Chemical interrogation spotlights IAV HA as a major druggable target for small-molecule inhibition. Our study identifies novel chemical scaffolds with high developmental potential, outlines diverse routes of IAV escape from entry inhibition, and establishes a path toward structure-aided lead development.IMPORTANCEThis study is one of the first to apply a fully replication-competent third-generation IAV reporter strain to a large-scale high-throughput screen (HTS) drug discovery campaign, allowing multicycle infection and screening in physiologically relevant human respiratory cells. A large number of potential druggable targets was thus chemically interrogated, but mechanistic characterization, positive target identification, and resistance profiling demonstrated that three chemically promising and structurally distinct hit classes selected for further analysis all block HA-mediated membrane fusion. Viral escape from inhibition could be achieved through primary and secondary resistance mechanisms.In silicodocking predicted compound binding to a microdomain located at the membrane-distal site of the prefusion HA stalk that was also previously suggested as a target site for chemically unrelated HA inhibitors. This study identifies an unexpected chemodominance of the HA stalk microdomain for small-molecule inhibitors in IAV inhibitor screening campaigns and highlights a novel mechanism of cooperative resistance to IAV entry blockers.

Molecular mechanisms of membrane fusion: Steps during phospholipid and exocytotic membrane fusion

1987 ◽  
Vol 7 (4) ◽  
pp. 251-268 ◽  
Author(s):  
Joshua Zimmerberg
Keyword(s):  
Membrane Fusion ◽  
Pore Formation ◽  
Molecular Mechanisms ◽  
Fusion Pore ◽  
Pore Widening

Exocytosis is considered as four separate steps: adhesion, fusion/pore formation, pore widening, and content discharge. Experiments on both synthetic and natural membranes are presented to show each of these steps. Major differences are seen in the two fusing systems. These differences are discussed in terms of molecular mechanisms of fusion.

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