Deletion of the SARS-CoV-2 Spike Cytoplasmic Tail Increases Infectivity in Pseudovirus Neutralization Assays

Jingyou Yu; Zhenfeng Li; Xuan He; Makda S Gebre; Esther A Bondzie; Huahua Wan; Catherine Jacob-Dolan; David R Martinez; Joseph P Nkolola; Ralph S Baric; Dan H Barouch

doi:10.1128/JVI.00044-21

Deletion of the SARS-CoV-2 Spike Cytoplasmic Tail Increases Infectivity in Pseudovirus Neutralization Assays

J Virol. 2021 May 10;95(11):e00044-21. doi: 10.1128/JVI.00044-21. Epub 2021 Mar 16.

Authors

Jingyou Yu¹, Zhenfeng Li¹, Xuan He¹, Makda S Gebre^{1

2}, Esther A Bondzie¹, Huahua Wan¹, Catherine Jacob-Dolan^{1

2

3}, David R Martinez⁴, Joseph P Nkolola¹, Ralph S Baric⁴, Dan H Barouch^{5

2

3}

Affiliations

¹ Center for Virology and Vaccine Research, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA 02215, USA.
² Harvard Medical School, Boston, MA, 02115.
³ Ragon Institute of MGH, MIT, and Harvard, Cambridge, MA 02139, USA.
⁴ University of North Carolina at Chapel Hill, Chapel Hill, NC, 27599, USA.
⁵ Center for Virology and Vaccine Research, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA 02215, USA dbarouch@bidmc.harvard.edu.

Abstract

Pseudotyped viruses are valuable tools for studying virulent or lethal viral pathogens that need to be handled in biosafety level 3 (BSL-3) or higher facilities. With the explosive spread of the coronavirus disease 2019 (COVID-19) pandemic, the establishment of a BSL-2 adapted SARS-CoV-2 pseudovirus neutralization assay is needed to facilitate the development of countermeasures. Here we describe an approach to generate a single-round lentiviral vector-based SARS-CoV-2 pseudovirus, which produced a signal more than 2 logs above background. Specifically, a SARS-CoV-2 spike variant with a cytoplasmic tail deletion of 13 amino acids, termed SΔCT13, conferred enhanced spike incorporation into pseudovirions and increased viral entry into cells as compared with full-length spike (S). We further compared S and SΔCT13 in terms of their sensitivity to vaccine sera, purified convalescent IgG, hACE2-mIgG, and the virus entry inhibitor BafA1. We developed a SΔCT13-based pseudovirus neutralization assay and defined key assay characteristics, including linearity, limit of detection, and intra- and intermediate-assay precision. Our data demonstrate that the SΔCT13-based pseudovirus shows enhanced infectivity in target cells, which will facilitate the assessment of humoral immunity to SARS-CoV-2 infection, antibody therapeutics, and vaccination. This pseudovirus neutralization assay can also be readily adapted to SARS-CoV-2 variants that emerge.IMPORTANCESARS-CoV-2 is the etiologic agent of the COVID-19 pandemic. The development of a high throughput pseudovirus neutralization assay is critical for the development of vaccines and immune-based therapeutics. In this study, we show that deletion of the cytoplasmic tail of the SARS-CoV-2 spike leads to pseudoviruses with enhanced infectivity. This SΔCT13-based pseudovirus neutralization assay should be broadly useful for the field.

Abstract

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