Genome-scale CRISPR Screens Identify Host Factors that Promote Human Coronavirus Infection

Abstract

The COVID-19 pandemic has resulted in 153 million infections and 3.2 million deaths as of May 2021. While effective vaccines are being administered globally, there is still a great need for antiviral therapies as potentially antigenically distinct SARS-CoV-2 variants continue to emerge across the globe. Viruses require host factors at every step in their life cycle, representing a rich pool of candidate targets for antiviral drug design. To identify host factors that promote SARS-CoV-2 infection with potential for broad-spectrum activity across the coronavirus family, we performed genome-scale CRISPR knockout screens in two cell lines (Vero E6 and HEK293T ectopically expressing ACE2) with SARS-CoV-2 and the common cold-causing human coronavirus OC43. While we identified multiple genes and functional pathways that have been previously reported to promote human coronavirus replication, we also identified a substantial number of novel genes and pathways. Of note, host factors involved in cell cycle regulation were enriched in our screens as were several key components of the programmed mRNA decay pathway. Finally, we identified novel candidate antiviral compounds targeting a number of factors revealed by our screens. Overall, our studies substantiate and expand the growing body of literature focused on understanding key human coronavirus-host cell interactions and exploit that knowledge for rational antiviral drug development.

A proteome-scale map of the SARS-CoV-2–human contactome

“…Although the antiviral activity of AZ1 was independently identified in Relative NF-κB induction (firefly/renillla) Vector ctrl. + TNF-α NSP14 a small-molecule screen 55 , our results inform mechanistic studies by identifying NSP16 as a viral interaction partner. NSP16 and associated complexes methylate viral RNA to prevent its detection and destruction by the innate immune system 56,57 .…”

Kinase-independent activity of DYRK1A promotes viral entry of highly pathogenic human coronaviruses

“…We previously identified DYRK1A as a pro-viral gene for both SARS-CoVs and MERS-CoV in Vero-E6 cells and Calu-3 cells 22,23 . Additional recent independent screens in Vero-E6 and Calu-3 cells have confirmed our initial finding of DYRK1A as a host dependency factor for SARS-CoV-2 33,34 . Numerous other SARS-CoV-2 genome-wide CRISPR knockout screens failed to identify DYRK1A – a disparity likely attributable to their reliance on cells that ectopically overexpress ACE2.…”

“…We previously identified DYRK1A as a pro-viral gene for both SARS-CoVs and MERS-CoV in Vero-E6 cells and Calu-3 cells 22,23 . Additional recent independent screens in Vero-E6 and Calu-3 cells have confirmed our initial finding of DYRK1A as a host dependency factor for SARS-CoV-2 33,34 .…”

“…While reproducibility between screens was excellent when performed on the same cell type, there was otherwise limited overlap across hits in these loss of function screens [27][28][29][30][31][32] . One notable exception was DYRK1A, which was a top enriched hit in both Vero-E6 and Calu-3 cells 22,23,33,34 . Interestingly, DYRK1A was not a hit in cell lines exogenously overexpressing ACE2, such as A549, Huh7.5, or HeLa cells [27][28][29][30][31][32] .…”

Phenotype-driven identification of drug targets for post-COVID-19 anosmia

“…Notably, one study recently screened a number of drugs in vitro against human coronaviruses, including an inhibitor of FGFR1-3 approved for various lung disease indications such as idiopathic pulmonary fibrosis, which showed only "moderate" inhibition of SARS-CoV-2 cytotoxicity 29 . However, we do not draw conclusions either way from this result, as the mechanism of action to address acute infection may be different from the mechanism of action needed to address post-acute symptoms (as in the case of NRP1, above).…”