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D614G Substitution of SARS-CoV-2 Spike Protein Increases Syncytium Formation and Virus Titer via Enhanced Furin-Mediated Spike Cleavage.
mBio. 2021 08 31; 12(4):e0058721.MBIO

Abstract

Since the D614G substitution in the spike (S) protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) emerged, the variant strain has undergone a rapid expansion to become the most abundant strain worldwide. Therefore, this substitution may provide an advantage for viral spreading. To explore the mechanism, we analyzed 18 viral isolates containing S proteins with either G614 or D614 (S-G614 and S-D614, respectively). The plaque assay showed a significantly higher virus titer in S-G614 than in S-D614 isolates. We further found increased cleavage of the S protein at the furin substrate site, a key event that promotes syncytium formation, in S-G614 isolates. The enhancement of the D614G substitution in the cleavage of the S protein and in syncytium formation has been validated in cells expressing S protein. The effect on the syncytium was abolished by furin inhibitor treatment and mutation of the furin cleavage site, suggesting its dependence on cleavage by furin. Our study pointed to the impact of the D614G substitution on syncytium formation through enhanced furin-mediated S cleavage, which might increase the transmissibility and infectivity of SARS-CoV-2 strains containing S-G614. IMPORTANCE Analysis of viral genomes and monitoring of the evolutionary trajectory of SARS-CoV-2 over time has identified the D614G substitution in spike (S) as the most prevalent expanding variant worldwide, which might confer a selective advantage in transmission. Several studies showed that the D614G variant replicates and transmits more efficiently than the wild-type virus, but the mechanism is unclear. By comparing 18 virus isolates containing S with either D614 or G614, we found significantly higher virus titers in association with higher furin protease-mediated cleavage of S, an event that promotes syncytium formation and virus infectivity, in the S-G614 viruses. The effect of the D614G substitution on furin-mediated S cleavage and the resulting enhancement of the syncytium phenotype has been validated in S-expressing cells. This study suggests a possible effect of the D614G substitution on S of SARS-CoV-2; the antiviral effect through targeting furin protease is worthy of being investigated in proper animal models.

Authors+Show Affiliations

Department of Microbiology, National Taiwan Universitygrid.19188.39 College of Medicine, Taipei, Taiwan.Department of Clinical Laboratory Sciences and Medical Biotechnology, National Taiwan Universitygrid.19188.39 College of Medicine, Taipei, Taiwan.Department of Microbiology, National Taiwan Universitygrid.19188.39 College of Medicine, Taipei, Taiwan.Hepatitis Research Center, National Taiwan Universitygrid.19188.39 Hospital, Taipei, Taiwan.Department of Clinical Laboratory Sciences and Medical Biotechnology, National Taiwan Universitygrid.19188.39 College of Medicine, Taipei, Taiwan.Department of Clinical Laboratory Sciences and Medical Biotechnology, National Taiwan Universitygrid.19188.39 College of Medicine, Taipei, Taiwan.Graduate Institute of Clinical Medicine, National Taiwan Universitygrid.19188.39 College of Medicine, Taipei, Taiwan.Department of Microbiology, National Taiwan Universitygrid.19188.39 College of Medicine, Taipei, Taiwan.Graduate Institute of Clinical Medicine, National Taiwan Universitygrid.19188.39 College of Medicine, Taipei, Taiwan.Graduate Institute of Clinical Medicine, National Taiwan Universitygrid.19188.39 College of Medicine, Taipei, Taiwan. Department of Internal Medicine, National Taiwan Universitygrid.19188.39 Hospital, Taipei, Taiwan. grid.19188.39National Taiwan University Center for Genomic Medicine, National Taiwan University grid.19188.39College of Medicine, Taipei, Taiwan.Department of Clinical Laboratory Sciences and Medical Biotechnology, National Taiwan Universitygrid.19188.39 College of Medicine, Taipei, Taiwan. Department of Laboratory Medicine, National Taiwan Universitygrid.19188.39 Hospital, Taipei, Taiwan.Department of Microbiology, National Taiwan Universitygrid.19188.39 College of Medicine, Taipei, Taiwan. Department of Clinical Laboratory Sciences and Medical Biotechnology, National Taiwan Universitygrid.19188.39 College of Medicine, Taipei, Taiwan. grid.19188.39National Taiwan University Center for Genomic Medicine, National Taiwan University grid.19188.39College of Medicine, Taipei, Taiwan.

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

34311586

Citation

Cheng, Ya-Wen, et al. "D614G Substitution of SARS-CoV-2 Spike Protein Increases Syncytium Formation and Virus Titer Via Enhanced Furin-Mediated Spike Cleavage." MBio, vol. 12, no. 4, 2021, pp. e0058721.
Cheng YW, Chao TL, Li CL, et al. D614G Substitution of SARS-CoV-2 Spike Protein Increases Syncytium Formation and Virus Titer via Enhanced Furin-Mediated Spike Cleavage. mBio. 2021;12(4):e0058721.
Cheng, Y. W., Chao, T. L., Li, C. L., Wang, S. H., Kao, H. C., Tsai, Y. M., Wang, H. Y., Hsieh, C. L., Lin, Y. Y., Chen, P. J., Chang, S. Y., & Yeh, S. H. (2021). D614G Substitution of SARS-CoV-2 Spike Protein Increases Syncytium Formation and Virus Titer via Enhanced Furin-Mediated Spike Cleavage. MBio, 12(4), e0058721. https://doi.org/10.1128/mBio.00587-21
Cheng YW, et al. D614G Substitution of SARS-CoV-2 Spike Protein Increases Syncytium Formation and Virus Titer Via Enhanced Furin-Mediated Spike Cleavage. mBio. 2021 08 31;12(4):e0058721. PubMed PMID: 34311586.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - D614G Substitution of SARS-CoV-2 Spike Protein Increases Syncytium Formation and Virus Titer via Enhanced Furin-Mediated Spike Cleavage. AU - Cheng,Ya-Wen, AU - Chao,Tai-Ling, AU - Li,Chiao-Ling, AU - Wang,Sheng-Han, AU - Kao,Han-Chieh, AU - Tsai,Ya-Min, AU - Wang,Hurng-Yi, AU - Hsieh,Chi-Ling, AU - Lin,You-Yu, AU - Chen,Pei-Jer, AU - Chang,Sui-Yuan, AU - Yeh,Shiou-Hwei, Y1 - 2021/07/27/ PY - 2021/7/28/pubmed PY - 2021/9/15/medline PY - 2021/7/27/entrez KW - SARS-CoV-2 KW - furin KW - spike KW - syncytium SP - e0058721 EP - e0058721 JF - mBio JO - mBio VL - 12 IS - 4 N2 - Since the D614G substitution in the spike (S) protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) emerged, the variant strain has undergone a rapid expansion to become the most abundant strain worldwide. Therefore, this substitution may provide an advantage for viral spreading. To explore the mechanism, we analyzed 18 viral isolates containing S proteins with either G614 or D614 (S-G614 and S-D614, respectively). The plaque assay showed a significantly higher virus titer in S-G614 than in S-D614 isolates. We further found increased cleavage of the S protein at the furin substrate site, a key event that promotes syncytium formation, in S-G614 isolates. The enhancement of the D614G substitution in the cleavage of the S protein and in syncytium formation has been validated in cells expressing S protein. The effect on the syncytium was abolished by furin inhibitor treatment and mutation of the furin cleavage site, suggesting its dependence on cleavage by furin. Our study pointed to the impact of the D614G substitution on syncytium formation through enhanced furin-mediated S cleavage, which might increase the transmissibility and infectivity of SARS-CoV-2 strains containing S-G614. IMPORTANCE Analysis of viral genomes and monitoring of the evolutionary trajectory of SARS-CoV-2 over time has identified the D614G substitution in spike (S) as the most prevalent expanding variant worldwide, which might confer a selective advantage in transmission. Several studies showed that the D614G variant replicates and transmits more efficiently than the wild-type virus, but the mechanism is unclear. By comparing 18 virus isolates containing S with either D614 or G614, we found significantly higher virus titers in association with higher furin protease-mediated cleavage of S, an event that promotes syncytium formation and virus infectivity, in the S-G614 viruses. The effect of the D614G substitution on furin-mediated S cleavage and the resulting enhancement of the syncytium phenotype has been validated in S-expressing cells. This study suggests a possible effect of the D614G substitution on S of SARS-CoV-2; the antiviral effect through targeting furin protease is worthy of being investigated in proper animal models. SN - 2150-7511 UR - https://www.unboundmedicine.com/medline/citation/34311586/D614G_Substitution_of_SARS_CoV_2_Spike_Protein_Increases_Syncytium_Formation_and_Virus_Titer_via_Enhanced_Furin_Mediated_Spike_Cleavage_ L2 - https://doi.org/10.1128/mBio.00587-21 DB - PRIME DP - Unbound Medicine ER -