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SARS-CoV-2 Spike Alterations Enhance Pseudoparticle Titers and Replication-Competent VSV-SARS-CoV-2 Virus.
Viruses. 2020 12 18; 12(12)V

Abstract

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of the most recent global pandemic that has caused more than a million deaths around the world. The spike glycoprotein (S) drives the entry and fusion of this virus and is the main determinant of cell tropism. To explore S requirements for entry under BSL2 conditions, S has been pseudotyped onto vesicular stomatitis virus (VSV) or retroviral particles with varied success. Several alterations to S were demonstrated to improve pseudoparticle titers, but they have not been systematically compared. In this study, we produced pseudotyped VSV particles with multiple modifications to S, including truncation, mutation, and tagging strategies. The main objective of this study was to determine which modifications of the S protein optimize cell surface expression, incorporation into pseudotyped particles, and pseudoparticle entry. Removal of the last 19 residues of the cytoplasmic tail produced a hyper-fusogenic S, while removal of 21 residues increased S surface production and VSV incorporation. Additionally, we engineered a replication-competent VSV (rVSV) virus to produce the S-D614G variant with a truncated cytoplasmic tail. While the particles can be used to assess S entry requirements, the rVSV∆G/SMet1D614G∆21 virus has a poor specific infectivity (particle to infectious titer ratio).

Authors+Show Affiliations

Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, GA 30602, USA.Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, GA 30602, USA.Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, GA 30602, USA.Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, GA 30602, USA.Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, GA 30602, USA.Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, GA 30602, USA.Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, GA 30602, USA. Department of Population Health, College of Veterinary Medicine, University of Georgia, Athens, GA 30602, USA.

Pub Type(s)

Journal Article
Research Support, N.I.H., Extramural

Language

eng

PubMed ID

33353101

Citation

Havranek, Katherine Elizabeth, et al. "SARS-CoV-2 Spike Alterations Enhance Pseudoparticle Titers and Replication-Competent VSV-SARS-CoV-2 Virus." Viruses, vol. 12, no. 12, 2020.
Havranek KE, Jimenez AR, Acciani MD, et al. SARS-CoV-2 Spike Alterations Enhance Pseudoparticle Titers and Replication-Competent VSV-SARS-CoV-2 Virus. Viruses. 2020;12(12).
Havranek, K. E., Jimenez, A. R., Acciani, M. D., Lay Mendoza, M. F., Reyes Ballista, J. M., Diaz, D. A., & Brindley, M. A. (2020). SARS-CoV-2 Spike Alterations Enhance Pseudoparticle Titers and Replication-Competent VSV-SARS-CoV-2 Virus. Viruses, 12(12). https://doi.org/10.3390/v12121465
Havranek KE, et al. SARS-CoV-2 Spike Alterations Enhance Pseudoparticle Titers and Replication-Competent VSV-SARS-CoV-2 Virus. Viruses. 2020 12 18;12(12) PubMed PMID: 33353101.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - SARS-CoV-2 Spike Alterations Enhance Pseudoparticle Titers and Replication-Competent VSV-SARS-CoV-2 Virus. AU - Havranek,Katherine Elizabeth, AU - Jimenez,Ariana R, AU - Acciani,Marissa Danielle, AU - Lay Mendoza,Maria Fernanda, AU - Reyes Ballista,Judith Mary, AU - Diaz,Darren Austin, AU - Brindley,Melinda Ann, Y1 - 2020/12/18/ PY - 2020/12/02/received PY - 2020/12/14/revised PY - 2020/12/16/accepted PY - 2020/12/23/entrez PY - 2020/12/24/pubmed PY - 2021/1/5/medline KW - SARS-CoV-2 KW - fusion KW - recombinant VSV KW - spike JF - Viruses JO - Viruses VL - 12 IS - 12 N2 - Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of the most recent global pandemic that has caused more than a million deaths around the world. The spike glycoprotein (S) drives the entry and fusion of this virus and is the main determinant of cell tropism. To explore S requirements for entry under BSL2 conditions, S has been pseudotyped onto vesicular stomatitis virus (VSV) or retroviral particles with varied success. Several alterations to S were demonstrated to improve pseudoparticle titers, but they have not been systematically compared. In this study, we produced pseudotyped VSV particles with multiple modifications to S, including truncation, mutation, and tagging strategies. The main objective of this study was to determine which modifications of the S protein optimize cell surface expression, incorporation into pseudotyped particles, and pseudoparticle entry. Removal of the last 19 residues of the cytoplasmic tail produced a hyper-fusogenic S, while removal of 21 residues increased S surface production and VSV incorporation. Additionally, we engineered a replication-competent VSV (rVSV) virus to produce the S-D614G variant with a truncated cytoplasmic tail. While the particles can be used to assess S entry requirements, the rVSV∆G/SMet1D614G∆21 virus has a poor specific infectivity (particle to infectious titer ratio). SN - 1999-4915 UR - https://www.unboundmedicine.com/medline/citation/33353101/SARS_CoV_2_Spike_Alterations_Enhance_Pseudoparticle_Titers_and_Replication_Competent_VSV_SARS_CoV_2_Virus_ L2 - https://www.mdpi.com/resolver?pii=v12121465 DB - PRIME DP - Unbound Medicine ER -