Vero cell-adapted SARS-CoV-2 strain shows increased viral growth through furin-mediated efficient spike cleavage
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) utilizes several host proteases to cleave the spike (S) protein to enter host cells. SARS-CoV-2 S protein is cleaved into S1 and S2 subunits by furin, which is closely involved in the pathogenicity of SARS-CoV-2. However, the effects of the modulated protease cleavage activity due to S protein mutations on viral replication and pathogenesis remain unclear. Herein, we serially passaged two SARS-CoV-2 strains in Vero cells and characterized the cell-adapted SARS-CoV-2 strains in vitro and in vivo. The adapted strains showed high viral growth, effective S1/S2 cleavage of the S protein, and low pathogenicity compared with the wild-type strain. Furthermore, the viral growth and S1/S2 cleavage were enhanced by the combination of the Δ68-76 and H655Y mutations using recombinant SARS-CoV-2 strains generated by the circular polymerase extension reaction. The recombinant SARS-CoV-2 strain, which contained the mutation of the adapted strain, showed increased susceptibility to the furin inhibitor, suggesting that the adapted SARS-CoV-2 strain utilized furin more effectively than the wild-type strain. Pathogenicity was attenuated by infection with effectively cleaved recombinant SARS-CoV-2 strains, suggesting that the excessive cleavage of the S proteins decreases virulence. Finally, the high-growth-adapted SARS-CoV-2 strain could be used as the seed for a low-cost inactivated vaccine; immunization with this vaccine can effectively protect the host from SARS-CoV-2 variants. Our findings provide novel insights into the growth and pathogenicity of SARS-CoV-2 in the evolution of cell-cell transmission.
IMPORTANCE: The efficacy of the S protein cleavage generally differs among the SARS-CoV-2 variants, resulting in distinct viral characteristics. The relationship between a mutation and the entry of SARS-CoV-2 into host cells remains unclear. In this study, we analyzed the sequence of high-growth Vero cell-adapted SARS-CoV-2 and factors determining the enhancement of the growth of the adapted virus and confirmed the characteristics of the adapted strain by analyzing the recombinant SARS-CoV-2 strain. We successfully identified mutations Δ68-76 and H655Y, which enhance viral growth and the S protein cleavage by furin. Using recombinant viruses enabled us to conduct a virus challenge experiment in vivo. The pathogenicity of SARS-CoV-2 introduced with the mutations Δ68-76, H655Y, P812L, and Q853L was attenuated in hamsters, indicating the possibility of the attenuation of excessive cleaved SARS-CoV-2. These findings provide novel insights into the infectivity and pathogenesis of SARS-CoV-2 strains, thereby significantly contributing to the field of virology.
| Medienart: |
E-Artikel |
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| Erscheinungsjahr: |
2024 |
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| Erschienen: |
2024 |
| Enthalten in: |
Zur Gesamtaufnahme - volume:12 |
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| Enthalten in: |
Microbiology spectrum - 12(2024), 4 vom: 02. Apr., Seite e0285923 |
| Sprache: |
Englisch |
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| Beteiligte Personen: |
Minami, Shohei [VerfasserIn] |
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| Links: |
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| Anmerkungen: |
Date Completed 08.04.2024 Date Revised 08.04.2024 published: Print-Electronic Citation Status MEDLINE |
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| doi: |
10.1128/spectrum.02859-23 |
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| Förderinstitution / Projekttitel: |
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| 245 | 1 | 0 | |a Vero cell-adapted SARS-CoV-2 strain shows increased viral growth through furin-mediated efficient spike cleavage |
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| 520 | |a Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) utilizes several host proteases to cleave the spike (S) protein to enter host cells. SARS-CoV-2 S protein is cleaved into S1 and S2 subunits by furin, which is closely involved in the pathogenicity of SARS-CoV-2. However, the effects of the modulated protease cleavage activity due to S protein mutations on viral replication and pathogenesis remain unclear. Herein, we serially passaged two SARS-CoV-2 strains in Vero cells and characterized the cell-adapted SARS-CoV-2 strains in vitro and in vivo. The adapted strains showed high viral growth, effective S1/S2 cleavage of the S protein, and low pathogenicity compared with the wild-type strain. Furthermore, the viral growth and S1/S2 cleavage were enhanced by the combination of the Δ68-76 and H655Y mutations using recombinant SARS-CoV-2 strains generated by the circular polymerase extension reaction. The recombinant SARS-CoV-2 strain, which contained the mutation of the adapted strain, showed increased susceptibility to the furin inhibitor, suggesting that the adapted SARS-CoV-2 strain utilized furin more effectively than the wild-type strain. Pathogenicity was attenuated by infection with effectively cleaved recombinant SARS-CoV-2 strains, suggesting that the excessive cleavage of the S proteins decreases virulence. Finally, the high-growth-adapted SARS-CoV-2 strain could be used as the seed for a low-cost inactivated vaccine; immunization with this vaccine can effectively protect the host from SARS-CoV-2 variants. Our findings provide novel insights into the growth and pathogenicity of SARS-CoV-2 in the evolution of cell-cell transmission | ||
| 520 | |a IMPORTANCE: The efficacy of the S protein cleavage generally differs among the SARS-CoV-2 variants, resulting in distinct viral characteristics. The relationship between a mutation and the entry of SARS-CoV-2 into host cells remains unclear. In this study, we analyzed the sequence of high-growth Vero cell-adapted SARS-CoV-2 and factors determining the enhancement of the growth of the adapted virus and confirmed the characteristics of the adapted strain by analyzing the recombinant SARS-CoV-2 strain. We successfully identified mutations Δ68-76 and H655Y, which enhance viral growth and the S protein cleavage by furin. Using recombinant viruses enabled us to conduct a virus challenge experiment in vivo. The pathogenicity of SARS-CoV-2 introduced with the mutations Δ68-76, H655Y, P812L, and Q853L was attenuated in hamsters, indicating the possibility of the attenuation of excessive cleaved SARS-CoV-2. These findings provide novel insights into the infectivity and pathogenesis of SARS-CoV-2 strains, thereby significantly contributing to the field of virology | ||
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| 700 | 1 | |a Sakai, Yusuke |e verfasserin |4 aut | |
| 700 | 1 | |a Okamura, Shinya |e verfasserin |4 aut | |
| 700 | 1 | |a Torii, Shiho |e verfasserin |4 aut | |
| 700 | 1 | |a Ono, Chikako |e verfasserin |4 aut | |
| 700 | 1 | |a Motooka, Daisuke |e verfasserin |4 aut | |
| 700 | 1 | |a Hamajima, Rina |e verfasserin |4 aut | |
| 700 | 1 | |a Nouda, Ryotaro |e verfasserin |4 aut | |
| 700 | 1 | |a Nurdin, Jeffery A |e verfasserin |4 aut | |
| 700 | 1 | |a Yamasaki, Moeko |e verfasserin |4 aut | |
| 700 | 1 | |a Kanai, Yuta |e verfasserin |4 aut | |
| 700 | 1 | |a Ebina, Hirotaka |e verfasserin |4 aut | |
| 700 | 1 | |a Maeda, Yusuke |e verfasserin |4 aut | |
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| 700 | 1 | |a Tachibana, Taro |e verfasserin |4 aut | |
| 700 | 1 | |a Matsuura, Yoshiharu |e verfasserin |4 aut | |
| 700 | 1 | |a Kobayashi, Takeshi |e verfasserin |4 aut | |
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