Characterising proteolysis during SARS-CoV-2 infection identifies viral cleavage sites and cellular targets with therapeutic potential

Bjoern Meyer (1) , Jeanne Chiaravalli (2) , Stacy Gellenoncourt (3) , Philip Brownridge (4) , Dominic Bryne (4) , Leonard Daly (4) , Arturas Grauslys (4) , Marius Walter (5) , Fabrice Agou (2) , Lisa A. Chakrabarti (3) , Charles Craik (6) , Claire Eyers (4) , Patrick Eyers (4) , Yann Gambin (7) , Andrew Jones (4) , Emma Sierecki (7) , Eric Verdin (5) , Marco Vignuzzi (1) , Edward Emmott (4)

1 Populations virales et Pathogenèse - Viral Populations and Pathogenesis
2 PF-CCB - Criblage chémogénomique et biologique (Plateforme) - Chemogenomic and Biological Screening Platform
3 CNRS-UMR3569 - Virus et Immunité - Virus and immunity
4 University of Liverpool
5 Buck Institute for Research on Aging
6 UC San Francisco - University of California [San Francisco]
7 UNSW - University of New South Wales [Sydney]

Abstract SARS-CoV-2 is the causative agent behind the COVID-19 pandemic, responsible for over 170 million infections, and over 3.7 million deaths worldwide. Efforts to test, treat and vaccinate against this pathogen all benefit from an improved understanding of the basic biology of SARS-CoV-2. Both viral and cellular proteases play a crucial role in SARS-CoV-2 replication. Here, we study proteolytic cleavage of viral and cellular proteins in two cell line models of SARS-CoV-2 replication using mass spectrometry to identify protein neo-N-termini generated through protease activity. We identify previously unknown cleavage sites in multiple viral proteins, including major antigens S and N: the main targets for vaccine and antibody testing efforts. We discover significant increases in cellular cleavage events consistent with cleavage by SARS-CoV-2 main protease, and identify 14 potential high-confidence substrates of the main and papain-like proteases. We show that siRNA depletion of these cellular proteins inhibits SARS-CoV-2 replication, and that drugs targeting two of these proteins: the tyrosine kinase SRC and Ser/Thr kinase MYLK, show a dose-dependent reduction in SARS-CoV-2 titres. Overall, our study provides a powerful resource to understand proteolysis in the context of viral infection, and to inform the development of targeted strategies to inhibit SARS-CoV-2 and treat COVID-19.

Domains: Virology

Bjoern Meyer, Jeanne Chiaravalli, Stacy Gellenoncourt, Philip Brownridge, Dominic Bryne, et al.. Characterising proteolysis during SARS-CoV-2 infection identifies viral cleavage sites and cellular targets with therapeutic potential. Nature Communications, 2021, 12 (1), pp.5553. ⟨10.1038/s41467-021-25796-w⟩. ⟨pasteur-03654295⟩