A single inactivating amino acid change in the SARS-CoV-2 NSP3 Mac1 domain attenuates viral replication in vivo
Category
Published on
Type
journal-article
Author
Taha Y. Taha and Rahul K. Suryawanshi and Irene P. Chen and Galen J. Correy and Maria McCavitt-Malvido and Patrick C. O’Leary and Manasi P. Jogalekar and Morgan E. Diolaiti and Gabriella R. Kimmerly and Chia-Lin Tsou and Ronnie Gascon and Mauricio Montano and Luis Martinez-Sobrido and Nevan J. Krogan and Alan Ashworth and James S. Fraser and Melanie Ott
Citation
Taha, T. Y., Suryawanshi, R. K., Chen, I. P., Correy, G. J., McCavitt-Malvido, M., O’Leary, P. C., Jogalekar, M. P., Diolaiti, M. E., Kimmerly, G. R., Tsou, C.-L., Gascon, R., Montano, M., Martinez-Sobrido, L., Krogan, N. J., Ashworth, A., Fraser, J. S., & Ott, M. (2023). A single inactivating amino acid change in the SARS-CoV-2 NSP3 Mac1 domain attenuates viral replication in vivo. PLOS Pathogens, 19(8), e1011614. https://doi.org/10.1371/journal.ppat.1011614
Abstract
Despite unprecedented efforts, our therapeutic arsenal against SARS-CoV-2 remains limited. The conserved macrodomain 1 (Mac1) in NSP3 is an enzyme exhibiting ADP-ribosylhydrolase activity and a possible drug target. To determine the role of Mac1 catalytic activity in viral replication, we generated recombinant viruses and replicons encoding a catalytically inactive NSP3 Mac1 domain by mutating a critical asparagine in the active site. While substitution to alanine (N40A) reduced catalytic activity by ~10-fold, mutations to aspartic acid (N40D) reduced activity by ~100-fold relative to wild-type. Importantly, the N40A mutation rendered Mac1 unstable in vitro and lowered expression levels in bacterial and mammalian cells. When incorporated into SARS-CoV-2 molecular clones, the N40D mutant only modestly affected viral fitness in immortalized cell lines, but reduced viral replication in human airway organoids by 10-fold. In mice, the N40D mutant replicated at >1000-fold lower levels compared to the wild-type virus while inducing a robust interferon response; all animals infected with the mutant virus survived infection. Our data validate the critical role of SARS-CoV-2 NSP3 Mac1 catalytic activity in viral replication and as a promising therapeutic target to develop antivirals.
