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J Virol. 2018 Sep 12;92(19). pii: e00683-18. doi: 10.1128/JVI.00683-18. Print 2018 Oct 1.

Middle East Respiratory Syndrome Coronavirus Spike Protein Is Not Activated Directly by Cellular Furin during Viral Entry into Target Cells.

Author information

1
Department of Virology III, National Institute of Infectious Diseases, Tokyo, Japan matuyama@nih.go.jp.
2
Department of Virology III, National Institute of Infectious Diseases, Tokyo, Japan.
3
Laboratory of Clinical Research on Infectious Diseases, Osaka University, Osaka, Japan.
4
Department of Virology I, National Institute of Infectious Diseases, Tokyo, Japan.

Abstract

Middle East respiratory syndrome coronavirus (MERS-CoV) utilizes host cellular proteases to enter cells. A previous report shows that furin, which is distributed mainly in the Golgi apparatus and cycled to the cell surface and endosomes, proteolytically activates the MERS-CoV spike (S) protein following receptor binding to mediate fusion between the viral and cellular membranes. In this study, we reexamined furin usage by MERS-CoV using a real-time PCR-based virus cell entry assay after inhibition of cellular proteases. We found that the furin inhibitor dec-RVKR-CMK blocked entry of MERS-CoV harboring an S protein lacking furin cleavage sites; it even blocked entry into furin-deficient LoVo cells. In addition, dec-RVKR-CMK inhibited not only the enzymatic activity of furin but also those of cathepsin L, cathepsin B, trypsin, papain, and TMPRSS2. Furthermore, a virus cell entry assay and a cell-cell fusion assay provided no evidence that the S protein was activated by exogenous furin. Therefore, we conclude that furin does not play a role in entry of MERS-CoV into cells and that the inhibitory effect of dec-RVKR-CMK is specific for TMPRSS2 and cathepsin L rather than furin.IMPORTANCE Previous studies using the furin inhibitor dec-RVKR-CMK suggest that MERS-CoV utilizes a cellular protease, furin, to activate viral glycoproteins during cell entry. However, we found that dec-RVKR-CMK inhibits not only furin but also other proteases. Furthermore, we found no evidence that MERS-CoV uses furin. These findings suggest that previous studies in the virology field based on dec-RVKR-CMK should be reexamined carefully. Here we describe appropriate experiments that can be used to assess the effect of protease inhibitors on virus cell entry.

KEYWORDS:

TMPRSS2; cathepsin L; coronavirus; dec-RVKR-CMK; furin

PMID:
30021905
PMCID:
PMC6146822
DOI:
10.1128/JVI.00683-18
[Indexed for MEDLINE]
Free PMC Article

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