The furin cleavage site of SARS-CoV-2 spike protein is a key determinant for transmission due to enhanced replication in airway cells

Peacock, Thomas P.; Goldhill, Daniel H.; Zhou, Jie; Baillon, Laury; Frise, Rebecca; Swann, Olivia C.; Kugathasan, Ruthiran; Penn, Rebecca; Brown, Jonathan C.; Sanchez-David, Raul Y.; Braga, Luca; Williamson, Maia Kavanagh; Hassard, Jack A.; Staller, Ecco; Hanley, Brian; Osborn, Michael; Giacca, Mauro; Davidson, Andrew D.; Matthews, David A.; Barclay, Wendy S.

SARS-CoV-2 enters cells via its spike glycoprotein which must be cleaved sequentially at the S1/S2, then the S2’ cleavage sites (CS) to mediate membrane fusion. SARS-CoV-2 has a unique polybasic insertion at the S1/S2 CS, which we demonstrate can be cleaved by furin. Using lentiviral pseudotypes and a cell-culture adapted SARS-CoV-2 virus with a S1/S2 deletion, we show that the polybasic insertion is selected for in lung cells and primary human airway epithelial cultures but selected against in Vero E6, a cell line used for passaging SARS-CoV-2. We find this selective advantage depends on expression of the cell surface protease, TMPRSS2, that allows virus entry independent of endosomes thus avoiding antiviral IFITM proteins. SARS-CoV-2 virus lacking the S1/S2 furin CS was shed to lower titres from infected ferrets and was not transmitted to cohoused sentinel animals. Thus, the polybasic CS is a key determinant for efficient SARS-CoV-2 transmission.