Functional evaluation of proteolytic activation for the SARS-CoV-2 variant B.1.1.7: role of the P681H mutation

Lubinski, Bailey; Tang, Tiffany; Daniel, Susan; Jaimes, Javier A.; Whittaker, Gary R.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the agent behind the current COVID-19 pandemic having emerged in Wuhan China in late 2019 from a yet to be determined animal reservoir. SARS-CoV-2 B.1.1.7, a variant identified in the UK in late 2020, contains a higher than typical level of point mutants across its genome, including P681H in the spike S1/S2 cleavage site. Here, we performed assays using fluorogenic peptides mimicking the S1/S2 sequence from Wuhan-Hu1 and B.1.1.7 and observed no definitive difference in furin cleavage between Wuhan-Hu1 and B.1.1.7 in vitro. We performed functional assays using pseudo-typed particles harboring SARS-CoV-2 spike proteins and observed no significant differences between Wuhan-Hu1, Wuhan-Hu1 P681H or B.1.1.7 spike-carrying pseudo-typed particles in VeroE6 or Vero-TMPRSS2 cells, despite the spikes containing P681H being more efficiently cleaved. Likewise, we or show no differences in cell-cell fusion assays using the spike P681H-expressing cells. Our findings suggest that while the introduction of P681H in the SARS-CoV-2 B.1.1.7 variant may increase spike cleavage by furin-like proteases, this does not significantly impact viral entry or cell-cell spread. We consider that other factors are at play to account for the increased in transmission and disease severity attributed to this variant of concern (VOC).