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Induction of Cell-Cell Fusion by Ebola Virus Glycoprotein: Low pH Is Not a Trigger

Ebola virus (EBOV) is a highly pathogenic filovirus that causes hemorrhagic fever in humans and animals. Currently, how EBOV fuses its envelope membrane within an endosomal membrane to cause infection is poorly understood. We successfully measure cell-cell fusion mediated by the EBOV fusion protein,...

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Autores principales: Markosyan, Ruben M., Miao, Chunhui, Zheng, Yi-Min, Melikyan, Gregory B., Liu, Shan-Lu, Cohen, Fredric S.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2016
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4711667/
https://www.ncbi.nlm.nih.gov/pubmed/26730950
http://dx.doi.org/10.1371/journal.ppat.1005373
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author Markosyan, Ruben M.
Miao, Chunhui
Zheng, Yi-Min
Melikyan, Gregory B.
Liu, Shan-Lu
Cohen, Fredric S.
author_facet Markosyan, Ruben M.
Miao, Chunhui
Zheng, Yi-Min
Melikyan, Gregory B.
Liu, Shan-Lu
Cohen, Fredric S.
author_sort Markosyan, Ruben M.
collection PubMed
description Ebola virus (EBOV) is a highly pathogenic filovirus that causes hemorrhagic fever in humans and animals. Currently, how EBOV fuses its envelope membrane within an endosomal membrane to cause infection is poorly understood. We successfully measure cell-cell fusion mediated by the EBOV fusion protein, GP, assayed by the transfer of both cytoplasmic and membrane dyes. A small molecule fusion inhibitor, a neutralizing antibody, as well as mutations in EBOV GP known to reduce viral infection, all greatly reduce fusion. By monitoring redistribution of small aqueous dyes between cells and by electrical capacitance measurements, we discovered that EBOV GP-mediated fusion pores do not readily enlarge—a marked difference from the behavior of other viral fusion proteins. EBOV GP must be cleaved by late endosome-resident cathepsins B or L in order to become fusion-competent. Cleavage of cell surface-expressed GP appears to occur in endosomes, as evidenced by the fusion block imposed by cathepsin inhibitors, agents that raise endosomal pH, or an inhibitor of anterograde trafficking. Treating effector cells with a recombinant soluble cathepsin B or thermolysin, which cleaves GP into an active form, increases the extent of fusion, suggesting that a fraction of surface-expressed GP is not cleaved. Whereas the rate of fusion is increased by a brief exposure to acidic pH, fusion does occur at neutral pH. Importantly, the extent of fusion is independent of external pH in experiments in which cathepsin activity is blocked and EBOV GP is cleaved by thermolysin. These results imply that low pH promotes fusion through the well-known pH-dependent activity of cathepsins; fusion induced by cleaved EBOV GP is a process that is fundamentally independent of pH. The cell-cell fusion system has revealed some previously unappreciated features of EBOV entry, which could not be readily elucidated in the context of endosomal entry.
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spelling pubmed-47116672016-01-26 Induction of Cell-Cell Fusion by Ebola Virus Glycoprotein: Low pH Is Not a Trigger Markosyan, Ruben M. Miao, Chunhui Zheng, Yi-Min Melikyan, Gregory B. Liu, Shan-Lu Cohen, Fredric S. PLoS Pathog Research Article Ebola virus (EBOV) is a highly pathogenic filovirus that causes hemorrhagic fever in humans and animals. Currently, how EBOV fuses its envelope membrane within an endosomal membrane to cause infection is poorly understood. We successfully measure cell-cell fusion mediated by the EBOV fusion protein, GP, assayed by the transfer of both cytoplasmic and membrane dyes. A small molecule fusion inhibitor, a neutralizing antibody, as well as mutations in EBOV GP known to reduce viral infection, all greatly reduce fusion. By monitoring redistribution of small aqueous dyes between cells and by electrical capacitance measurements, we discovered that EBOV GP-mediated fusion pores do not readily enlarge—a marked difference from the behavior of other viral fusion proteins. EBOV GP must be cleaved by late endosome-resident cathepsins B or L in order to become fusion-competent. Cleavage of cell surface-expressed GP appears to occur in endosomes, as evidenced by the fusion block imposed by cathepsin inhibitors, agents that raise endosomal pH, or an inhibitor of anterograde trafficking. Treating effector cells with a recombinant soluble cathepsin B or thermolysin, which cleaves GP into an active form, increases the extent of fusion, suggesting that a fraction of surface-expressed GP is not cleaved. Whereas the rate of fusion is increased by a brief exposure to acidic pH, fusion does occur at neutral pH. Importantly, the extent of fusion is independent of external pH in experiments in which cathepsin activity is blocked and EBOV GP is cleaved by thermolysin. These results imply that low pH promotes fusion through the well-known pH-dependent activity of cathepsins; fusion induced by cleaved EBOV GP is a process that is fundamentally independent of pH. The cell-cell fusion system has revealed some previously unappreciated features of EBOV entry, which could not be readily elucidated in the context of endosomal entry. Public Library of Science 2016-01-05 /pmc/articles/PMC4711667/ /pubmed/26730950 http://dx.doi.org/10.1371/journal.ppat.1005373 Text en © 2016 Markosyan et al http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.
spellingShingle Research Article
Markosyan, Ruben M.
Miao, Chunhui
Zheng, Yi-Min
Melikyan, Gregory B.
Liu, Shan-Lu
Cohen, Fredric S.
Induction of Cell-Cell Fusion by Ebola Virus Glycoprotein: Low pH Is Not a Trigger
title Induction of Cell-Cell Fusion by Ebola Virus Glycoprotein: Low pH Is Not a Trigger
title_full Induction of Cell-Cell Fusion by Ebola Virus Glycoprotein: Low pH Is Not a Trigger
title_fullStr Induction of Cell-Cell Fusion by Ebola Virus Glycoprotein: Low pH Is Not a Trigger
title_full_unstemmed Induction of Cell-Cell Fusion by Ebola Virus Glycoprotein: Low pH Is Not a Trigger
title_short Induction of Cell-Cell Fusion by Ebola Virus Glycoprotein: Low pH Is Not a Trigger
title_sort induction of cell-cell fusion by ebola virus glycoprotein: low ph is not a trigger
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4711667/
https://www.ncbi.nlm.nih.gov/pubmed/26730950
http://dx.doi.org/10.1371/journal.ppat.1005373
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