A reporter cell line for the automated quantification of SARS-CoV-2 infection in living cells

• Published in: Frontiers in microbiology Vol. 13; p. 1031204
• Main Authors: Desmarets, Lowiese, Callens, Nathalie, Hoffmann, Eik, Danneels, Adeline, Lavie, Muriel, Couturier, Cyril, Dubuisson, Jean, Belouzard, Sandrine, Rouillé, Yves
• Format: Journal Article
• Language: English
• Published: Switzerland Frontiers Media 29.09.2022; Frontiers Media S.A
• Subjects: 3CLpro; antivirals; Cellular Biology; coronavirus; high content imaging; Life Sciences; Microbiology; Microbiology and Parasitology; SARS-CoV-2; screening; Subcellular Processes; Virology; antivirals; SARS-CoV-2; screening; Mpro; coronavirus; nsp5; 3CLpro; high content imaging; M pro; 3CL pro
• ISSN: 1664-302X; 1664-302X
• DOI: 10.3389/fmicb.2022.1031204

The SARS-CoV-2 pandemic and the urgent need for massive antiviral testing highlighted the lack of a good cell-based assay that allowed for a fast, automated screening of antivirals in high-throughput content with minimal handling requirements in a BSL-3 environment. The present paper describes the construction of a green fluorescent substrate that, upon cleavage by the SARS-CoV-2 main protease, re-localizes from the cytoplasm in non-infected cells to the nucleus in infected cells. The construction was stably expressed, together with a red fluorescent nuclear marker, in a highly susceptible clone derived from Vero-81 cells. With this fluorescent reporter cell line, named F1G-red, SARS-CoV-2 infection can be scored automatically in living cells by comparing the patterns of green and red fluorescence signals acquired by automated confocal microscopy in a 384-well plate format. We show the F1G-red system is sensitive to several SARS-CoV-2 variants of concern and that it can be used to assess antiviral activities of compounds in dose–response experiments. This high-throughput system will provide a reliable tool for antiviral screening against SARS-CoV-2.