SARS-CoV-2 spike cleavage and transmission

Coronaviruses enter cells via their spike glycoprotein which must be cleaved sequentially, first at the S1/S2 and then at the S2' cleavage sites to mediate membrane fusion. SARS-CoV-2 has a unique polybasic insertion at the S1/S2 cleavage site, not found in its close relatives. Here, we demonstrate that the polybasic cleavage site can be cleaved by furin. Using viral 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 cultures but selected against in Vero E6, a cell line used for passaging SARS-CoV-2. We find this selective advantage depended on express TMPRSS2, and that TMPRSS2 expression allows avoidance of the antiviral factor IFITM3. We infect ferrets with SARS-CoV-2 virus with and without the S1/S2 cleavage site and show that the polybasic insertion is vital for viral transmission. Thus, the polybasic cleavage site is likely a key determinant for efficient SARS-CoV-2 transmission.