Denys Khaperskyy, Dalhousie University

Madeleine Stolz, Caleb Galbraith, Scott Tersteeg, Emily Andrews, Trushar R. Patel, and Denys A. Khaperskyy Department of Microbiology and Immunology, Faculty of Medicine, Dalhousie University, Halifax, Nova Scotia B3H 4R2, Canada; Alberta RNA Research and Training Institute, Department of Chemistry and Biochemistry, Faculty of Arts and Science, University of Lethbridge, Lethbridge, Alberta B3H 4R2, Canada

Suppressing & Conquering

Host shutoff is a process by which viruses suppress host gene expression. Non-structural protein 1 (Nsp1) of SARS-CoV-2 binds the ribosome and drives host shutoff by blocking mRNA access to the ribosome's RNA entry channel and by degrading host transcripts. At the same time, Nsp1 shutoff is selective and permits translation of viral transcripts and select host mRNAs, such as the stress granule-binding TIA1-related protein (TIAR), and many host mRNAs containing terminal oligopyrimidine (TOP) motifs. In SARS-CoV-2, the 5' leader sequence shared between all viral transcripts confers Nsp1 resistance, however the mechanism thereby viral or select host transcripts escape Nsp1 shutoff is poorly understood. We used reporter assays to examine the features of TIAR mRNA that allow it to resist Nsp1 shutoff and showed that the absence of guanosines (Gs) from a window 10-18 nucleotides downstream from the 5' end of both TIAR mRNA and viral mRNAs is sufficient and necessary to confer Nsp1 resistance. We also showed that removal of Gs from the 10-18 nucleotide window in the 5' untranslated region (UTR) of a susceptible control mRNA similarly conferred Nsp1 resistance. The TOP motif-containing transcript encoding eukaryotic elongation factor 2 (EEF2), however, displayed Nsp1 resistance despite lacking a G-less window in its 5' UTR, suggesting that transcripts can escape Nsp1 shutoff by multiple mechanisms. Our findings are consistent with a model that argues that the Nsp1 escape is not determined by the secondary structure, but by sequence features of mRNA 5' UTRs.