Abstract
SARS-CoV spike (S) protein-mediated cell fusion is important for the viral entry mechanism and identification of SARS-CoV entry inhibitors. In order to avoid the high risks involved in handling SARS-CoV and to facilitate the study of viral fusion mechanism, we established the cell lines: SR-COS7 cells that stably express both SARS-CoV S protein and red fluorescence protein, R-COS7 cells that stably express red fluorescence protein, and AG-COS7 cells that stably express both ACE2 and green fluorescence protein, respectively. When SR-COS7 cells or R-COS7 cells were cocultured with AG-COS7 cells, syncytia with yellow fluorescence were conveniently observed after 12 h in SR-COS7 cells plus AG-COS7 cells, but not in R-COS7 cells plus AG-COS7 cells. The cell-to-cell fusion efficiency was simply determined for quantitative analysis based on the number of syncytium detected by flow cytometry. Such new cell-to-cell fusion model was further assessed by the potent HR2 peptide inhibitor, which led to the obvious decrease of the cell-to-cell fusion efficiency. The successful fusion and inhibition of cell-based binding assay shows that it can be well used for the study of SARS-CoV entry and inhibition.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Angiotensin-Converting Enzyme 2
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Animals
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Biological Assay*
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COS Cells
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Cell Fusion
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Chlorocebus aethiops
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Coculture Techniques
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Escherichia coli / genetics
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Fluorescent Dyes / metabolism
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Glutathione Transferase / metabolism
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Green Fluorescent Proteins / metabolism
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Luminescent Proteins / metabolism
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Membrane Glycoproteins / genetics
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Membrane Glycoproteins / metabolism*
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Peptides / chemistry
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Peptides / metabolism
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Peptidyl-Dipeptidase A / metabolism
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Recombinant Proteins / metabolism
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Red Fluorescent Protein
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Severe Acute Respiratory Syndrome / metabolism*
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Severe acute respiratory syndrome-related coronavirus / genetics
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Severe acute respiratory syndrome-related coronavirus / metabolism*
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Spike Glycoprotein, Coronavirus
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Time Factors
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Trypsin / pharmacology
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Viral Envelope Proteins / genetics
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Viral Envelope Proteins / metabolism*
Substances
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Fluorescent Dyes
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Luminescent Proteins
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Membrane Glycoproteins
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Peptides
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Recombinant Proteins
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Spike Glycoprotein, Coronavirus
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Viral Envelope Proteins
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spike glycoprotein, SARS-CoV
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spike protein, mouse hepatitis virus
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Green Fluorescent Proteins
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Glutathione Transferase
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Peptidyl-Dipeptidase A
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Angiotensin-Converting Enzyme 2
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Trypsin