부산대학교 도서관

Interferons establish an antiviral state through the induction of hundreds of interferon-stimulated genes (ISGs). The mechanisms and viral specificities for most ISGs remain incompletely understood. To enable high-throughput interrogation of ISG antiviral functions in pooled genetic screens while mitigating potentially confounding effects of endogenous interferon and antiproliferative/proapoptotic ISG activities, we adapted a CRISPR-activation (CRISPRa) system for inducible ISG expression in isogenic cell lines with and without the capacity to respond to interferons. We used this platform to screen for ISGs that restrict SARS-CoV-2. Results included ISGs previously described to restrict SARS-CoV-2 and novel candidate antiviral factors. We validated a subset of these by complementary CRISPRa and cDNA expression experiments. OAS1, a top-ranked hit across multiple screens, exhibited strong antiviral effects against SARS-CoV-2, which required OAS1 catalytic activity. These studies demonstrate a high-throughput approach to assess antiviral functions within the ISG repertoire, exemplified by identification of multiple SARS-CoV-2 restriction factors. Author summary: To counteract viral infection, interferon induces the expression of antiviral effectors collectively termed Interferon Stimulated Genes (ISGs). Different effector genes restrict different viruses through a variety of mechanisms. Identifying the particular ISGs that restrict specific viruses can uncover viral vulnerabilities and inform therapeutic strategies. Here, we developed a CRISPR-activation (CRISPRa) experimental strategy for screening hundreds of pooled ISGs for antiviral activity in parallel while accounting for the ability of some ISGs to inhibit cell cycle and to promote cell death. We applied this approach to detect ISGs that restrict SARS-CoV-2, the virus that causes COVID-19. In a series of controlled experiments, we identified multiple ISGs that counteract SARS-CoV-2 infection, and further validated a subset of these ISGs through focused CRISPRa and cDNA expression studies. Our results validate several previously identified ISGs, and identify multiple novel antiviral effectors with activity against SARS-CoV-2. Overall, our work demonstrates an experimental system for the controlled assessment of ISG antiviral activities, and expands the present understanding of innate antiviral defense against SARS-CoV-2. [ABSTRACT FROM AUTHOR]