tetano
Editor, Senior Moderator
Microbiol Spectr
. 2024 Feb 28:e0285923.
doi: 10.1128/spectrum.02859-23. Online ahead of print. Vero cell-adapted SARS-CoV-2 strain shows increased viral growth through furin-mediated efficient spike cleavage
Shohei Minami[SUP] 1 [/SUP], Tomohiro Kotaki[SUP] 1 [/SUP], Yusuke Sakai[SUP] 2 [/SUP], Shinya Okamura[SUP] 3 4 [/SUP], Shiho Torii[SUP] 5 [/SUP], Chikako Ono[SUP] 5 6 [/SUP], Daisuke Motooka[SUP] 7 [/SUP], Rina Hamajima[SUP] 1 [/SUP], Ryotaro Nouda[SUP] 1 [/SUP], Jeffery A Nurdin[SUP] 1 [/SUP], Moeko Yamasaki[SUP] 1 [/SUP], Yuta Kanai[SUP] 1 [/SUP], Hirotaka Ebina[SUP] 3 4 6 8 [/SUP], Yusuke Maeda[SUP] 9 [/SUP], Toru Okamoto[SUP] 10 [/SUP], Taro Tachibana[SUP] 11 12 [/SUP], Yoshiharu Matsuura[SUP] 5 6 8 [/SUP], Takeshi Kobayashi[SUP] 1 6 8 [/SUP]
Affiliations
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.IMPORTANCEThe 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.
Keywords: infection; pathogenicity; severe acute respiratory syndrome virus-2 (SARS-CoV-2); spike (S) protein; viral replication.
. 2024 Feb 28:e0285923.
doi: 10.1128/spectrum.02859-23. Online ahead of print. Vero cell-adapted SARS-CoV-2 strain shows increased viral growth through furin-mediated efficient spike cleavage
Shohei Minami[SUP] 1 [/SUP], Tomohiro Kotaki[SUP] 1 [/SUP], Yusuke Sakai[SUP] 2 [/SUP], Shinya Okamura[SUP] 3 4 [/SUP], Shiho Torii[SUP] 5 [/SUP], Chikako Ono[SUP] 5 6 [/SUP], Daisuke Motooka[SUP] 7 [/SUP], Rina Hamajima[SUP] 1 [/SUP], Ryotaro Nouda[SUP] 1 [/SUP], Jeffery A Nurdin[SUP] 1 [/SUP], Moeko Yamasaki[SUP] 1 [/SUP], Yuta Kanai[SUP] 1 [/SUP], Hirotaka Ebina[SUP] 3 4 6 8 [/SUP], Yusuke Maeda[SUP] 9 [/SUP], Toru Okamoto[SUP] 10 [/SUP], Taro Tachibana[SUP] 11 12 [/SUP], Yoshiharu Matsuura[SUP] 5 6 8 [/SUP], Takeshi Kobayashi[SUP] 1 6 8 [/SUP]
Affiliations
- PMID: 38415690
- DOI: 10.1128/spectrum.02859-23
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.IMPORTANCEThe 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.
Keywords: infection; pathogenicity; severe acute respiratory syndrome virus-2 (SARS-CoV-2); spike (S) protein; viral replication.