Abstract: Acta Biochim Biophys Sin 2023, 55(11): 1840–1843
https://doi.org/10.3724/abbs.2023129
Advance Access Publication Date: 6 July 2023
Lab Note
Lab Note
A pseudovirus-based method to dynamically mimic
SARS-CoV-2-associated cell-to-cell fusion and transmission
Xiangpeng Sheng1,2,3,†,*, Yi Yang4,†, Fang Zhu5,†, Fan Yang2,†, Honghua Wang1, and
Ronggui Hu1,2,*
1Key Laboratory of Systems Health Science of Zhejiang Province, School of Life Science, Hangzhou Institute for Advanced Study, University of
Chinese Academy of Sciences, Hangzhou 310024, China, 2State Key Laboratory of Molecular Biology, Shanghai Institute of Biochemistry and
Cell Biology, Center for Excellence in Molecular Cell Science, University of Chinese Academy of Sciences, Chinese Academy of Sciences,
Shanghai 200031, China, 3State Key Laboratory of Animal Disease Control, Harbin Veterinary Research Institute, Chinese Academy of
Agricultural Sciences, Harbin 150069, China, 4Department of Thoracic Surgery, Ruijin Hospital, Shanghai Jiaotong University School of
Medicine, Shanghai 200025, China, and 5School of Medicine, Guizhou University, Guiyang 550025, China
†These authors contributed equally to this work.
*Correspondence address. Tel: +86-17717541320; E-mail: coryhu@sibcb.ac.cn (R.H.) / Tel: +86-13701790678; E-mail: shengxiangpeng@ucas.ac.cn (X.S.)
Received 8 June 2023 Accepted 25 June 2023
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2),
responsible for the COVID-19 pandemic, has caused tremendous
global loss and continues to evolve to generate variants. Entry of
SARS-CoV-2 into target host cells is primarily mediated by spike (S),
which binds to the host receptor hACE2 and initiates virus-cell
membrane fusion [1]. Most COVID-19 patients show pneumocyte
syncytia in the lungs [2]. Cell fusion contributes to viral entry, cellto-cell transmission and tissue damage, and thus attracts much
attention. Because authentic SARS-CoV-2 live virions can only be
handled in biosafety level-3 (BSL-3) facilities, many researchers
have developed different assays to study cell fusion in BSL-1/2 by
directly expressing S and hACE2 on mammalian cells [2‒5]. Briefly,
in regular cell fusion assays, S-expressing cells and hACE2-positive
cells are cocultured at approximately a 1:1 ratio, which induces cellcell fusion and usually activates a fusion reporter. Although these
strategies are useful, they cannot efficiently simulate cell-cell fusion
and transmission in SARS-CoV-2 infection, in which virions from
one target cell are transmitted to neighboring cells, resulting in
syncytia. Here, we design a pseudovirus-based method to dynamically and highly mimic cell-to-cell fusion and virus transmission of
SARS-CoV-2.
First, we generated spike-pseudotyped virions (S pseudovirions)
in HEK293FT cells by co-transfecting three plasmids, including
psPAX2, pCDH-sfGFP, and a plasmid expressing SARS-CoV-2 S into
cells (Figure 1A), and collected the viral supernatant, which is
similar to a previous report [6]. S pseudovirus was found to
efficiently infect hACE2-positive 293T cells (293T-hACE2) but not
the control 293T or 293FT cell lines (Supplementary Figure S1A,B),
suggesting that S indeed envelops the pseudovirions. However,
fluorescence microscopy revealed that the infection of 293T-hACE2
cells by pseudovirus supernatant could not trigger cell-cell
membrane fusion events (Supplementary Figure S1C). Given that
the authentic SARS-CoV-2-infected host cells can continue to
generate live virions to..
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