Monitoring fusion kinetics of viral and target cell membranes in living cells using a SARS-CoV-2 spike-protein-mediated membrane fusion assay

Hesham Nasser; Ryo Shimizu; Jumpei Ito; Genotype to Phenotype Japan (G2P-Japan) Consortium; Akatsuki Saito; Kei Sato; Terumasa Ikeda

doi:10.1016/j.xpro.2022.101773

Monitoring fusion kinetics of viral and target cell membranes in living cells using a SARS-CoV-2 spike-protein-mediated membrane fusion assay

STAR Protoc. 2022 Sep 26;3(4):101773. doi: 10.1016/j.xpro.2022.101773. eCollection 2022 Dec 16.

Authors

Hesham Nasser¹, Ryo Shimizu², Jumpei Ito³; Genotype to Phenotype Japan (G2P-Japan) Consortium; Akatsuki Saito⁴, Kei Sato⁵, Terumasa Ikeda⁶

Collaborators

Genotype to Phenotype Japan (G2P-Japan) Consortium:
Keita Matsuno, Naganori Nao, Hirofumi Sawa, Mai Kishimoto, Shinya Tanaka, Masumi Tsuda, Lei Wang, Yoshikata Oda, Marie Kato, Zannatul Ferdous, Hiromi Mouri, Kenji Shishido, Takasuke Fukuhara, Tomokazu Tamura, Rigel Suzuki, Hayato Ito, Daichi Yamasoba, Izumi Kimura, Naoko Misawa, Keiya Uriu, Yusuke Kosugi, Shigeru Fujita, Mai Suganami, Mika Chiba, Ryo Yoshimura, So Nakagawa, Jiaqi Wu, Akifumi Takaori-Kondo, Kotaro Shirakawa, Kayoko Nagata, Yasuhiro Kazuma, Ryosuke Nomura, Yoshihito Horisawa, Yusuke Tashiro, Yugo Kawai, Takashi Irie, Ryoko Kawabata, Mst Monira Begum, Otowa Takahashi, Kimiko Ichihara, Takamasa Ueno, Chihiro Motozono, Mako Toyoda, Yuri L Tanaka, Erika P Butlertanaka, Maya Shofa, Kazuo Takayama, Rina Hashimoto, Sayaka Deguchi, Takao Hashiguchi, Tateki Suzuki, Kanako Kimura, Jiei Sasaki, Yukari Nakajima, Kaori Tabata

Affiliations

¹ Division of Molecular Virology and Genetics, Joint Research Center for Human Retrovirus Infection, Kumamoto University, Kumamoto 8600811, Japan; Department of Clinical Pathology, Faculty of Medicine, Suez Canal University, Ismailia 41511, Egypt.
² Division of Molecular Virology and Genetics, Joint Research Center for Human Retrovirus Infection, Kumamoto University, Kumamoto 8600811, Japan; Graduate School of Medical Sciences, Kumamoto University, Kumamoto 8600811, Japan.
³ Division of Systems Virology, Department of Microbiology and Immunology, The Institute of Medical Science, The University of Tokyo, Tokyo 1088639, Japan.
⁴ Department of Veterinary Science, Faculty of Agriculture, University of Miyazaki, Miyazaki 8892192, Japan; Center for Animal Disease Control, University of Miyazaki, Miyazaki 8892192, Japan; Graduate School of Medicine and Veterinary Medicine, University of Miyazaki, Miyazaki 8891692, Japan.
⁵ Division of Systems Virology, Department of Microbiology and Immunology, The Institute of Medical Science, The University of Tokyo, Tokyo 1088639, Japan; International Research Center for Infectious Diseases, The Institute of Medical Science, The University of Tokyo, Tokyo 1088639, Japan; International Vaccine Design Center, The Institute of Medical Science, The University of Tokyo, Tokyo 1088639, Japan; Graduate School of Medicine, The University of Tokyo, Tokyo 1138654, Japan; Graduate School of Frontier Sciences, The University of Tokyo, Kashiwa 2778561, Japan; Collaboration Unit for Infection, Joint Research Center for Human Retrovirus Infection, Kumamoto University, Kumamoto 8600811, Japan; CREST, Japan Science and Technology Agency, Kawaguchi 3320012, Japan.
⁶ Division of Molecular Virology and Genetics, Joint Research Center for Human Retrovirus Infection, Kumamoto University, Kumamoto 8600811, Japan. Electronic address: ikedat@kumamoto-u.ac.jp.

Abstract

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike protein mediates membrane fusion between the virus and the target cells, triggering viral entry into the latter. Here, we describe a SARS-CoV-2 spike-protein-mediated membrane fusion assay using a dual functional split reporter protein to quantitatively monitor the fusion kinetics of the viral and target cell membranes in living cells. This approach can be applied in various cell types, potentially predicting the pathogenicity of newly emerging variants. For complete details on the use and execution of this protocol, please refer to Kimura et al. (2022b), Kimura et al. (2022c), Motozono et al. (2021), Saito et al. (2022a), Saito et al. (2022b), Suzuki et al. (2022), and Yamasoba et al. (2022).

Keywords: Cell-based Assays; Microbiology.

Publication types

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

MeSH terms

COVID-19*
Cell Membrane / metabolism
Humans
Membrane Fusion
SARS-CoV-2 / genetics
Spike Glycoprotein, Coronavirus* / genetics

Substances

spike protein, SARS-CoV-2
Spike Glycoprotein, Coronavirus