CRISPR activation as a platform to identify interferon stimulated genes with anti-viral function

Emily N Kirby; Xavier B Montin; Timothy P Allen; Jaslan Densumite; Brooke N Trowbridge; Michael R Beard

doi:10.1177/17534259231225611

CRISPR activation as a platform to identify interferon stimulated genes with anti-viral function

Innate Immun. 2024 Jan 23:17534259231225611. doi: 10.1177/17534259231225611. Online ahead of print.

Authors

Emily N Kirby^{1

2}, Xavier B Montin^{1

2}, Timothy P Allen², Jaslan Densumite³, Brooke N Trowbridge^{1

2}, Michael R Beard^{1

2}

Affiliations

¹ Research Centre for Infectious Diseases, The University of Adelaide, Adelaide, South Australia, Australia.
² Discipline of Molecular and Biomedical Science, The University of Adelaide, Adelaide, South Australia, Australia.
³ Department of Immunology, Faculty of Medicine, Siriraj Hospital, Mahidol University, Bangkok, Thailand.

PMID: 38258394
DOI: 10.1177/17534259231225611

Abstract

Interferon Stimulated Gene (ISG) expression plays a key role in the control of viral replication and development of a robust adaptive response. Understanding this dynamic relationship between the pathogen and host is critical to our understanding of viral life-cycles and development of potential novel anti-viral strategies. Traditionally, plasmid based exogenous prompter driven expression of ISGs has been used to investigate anti-viral ISG function, however there are deficiencies in this approach. To overcome this, we investigated the utility of CRISPR activation (CRISPRa), which allows for targeted transcriptional activation of a gene from its endogenous promoter. Using the CRISPRa-SAM system to induce targeted expression of a panel of anti-viral ISGs we showed robust induction of mRNA and protein expression. We then employed our CRISPRa-SAM ISG panel in several antiviral screen formats to test for the ability of ISGs to prevent viral induced cytopathic cell death (CPE) and replication of Dengue Virus (DENV), Zika Virus (ZIKV), West Nile Virus Kunjin (WNV_KUN), Hepatitis A Virus (HAV) and Human Coronavirus 229E (HCoV-229E). Our CRISPRa approach confirmed the anti-viral activity of ISGs like IFI6, IFNβ and IFNλ2 that prevented viral induced CPE, which was supported by high-content immunofluorescence imaging analysis. This work highlights CRISPRa as a rapid, agile, and powerful methodology to identify and characterise ISGs and viral restriction factors.

Keywords: Antiviral; CRISPR activation; interferon stimulated genes; virus.