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  3. Replicon-based genome-wide CRISPR knockout screening for the identification of host factors involved in viral replication

Replicon-based genome-wide CRISPR knockout screening for the identification of host factors involved in viral replication

  • Nat Commun. 2025 Dec 10;16(1):11028. doi: 10.1038/s41467-025-65979-3.
Karen W Cheng 1,  Madhura Bhave 1,  Andrew L Markhard 1 2,  Duo Peng 1,  Karan D Bhatt 1 3,  Katherine A Travisano 1 4,  Josette V Medicielo 1 5,  Astrid Anaya 1,  Sanae Lembirik 6,  Leila Njoya 1,  Manu Anantpadma 6,  Jens H Kuhn 6,  Andreas S Puschnik 1,  Amy L Kistler 7
Affiliations

Affiliations

  • 1 Chan Zuckerberg Biohub, San Francisco, CA, USA.
  • 2 New York University Grossman School of Medicine, New York, NY, USA.
  • 3 Washington University School of Medicine, St. Louis, MO, USA.
  • 4 Stanford University, Palo Alto, CA, USA.
  • 5 Medical College of Wisconsin, Milwaukee, WI, USA.
  • 6 Integrated Research Facility at Fort Detrick, Division of Clinical Research, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Fort Detrick, Frederick, MD, USA.
  • 7 Chan Zuckerberg Biohub, San Francisco, CA, USA. [email protected].
PMID: 41372121 DOI: 10.1038/s41467-025-65979-3
Abstract

We describe a viral replicon-based CRISPR knockout (KO) screening approach to specifically identify host factors essential for viral replication which are often missed in live virus screens. We benchmark the replicon screening using a stable fluorescent Dengue Virus type 2 (DENV-2) replicon cell line and successfully identify host genes known to be required for viral DENV-2 replication (e.g., endoplasmic reticulum membrane complex and oligosaccharyltransferase complex components), along with additional genes that have not been reported in prior CRISPR KO screens with DENV-2. We extend this replicon screening approach to chikungunya virus (CHIKV), a positive-sense RNA virus, and Ebola virus (EBOV), a negative-sense RNA virus, and identify distinct sets of genes required for replication of each virus. Our findings indicate that viral replicon-based CRISPR screens are a useful approach to identify host factors essential for replication of diverse viruses and to elucidate potential novel targets for host-directed medical countermeasures.

Figures
Products
  • Cat. No.
    Product Name
    Description
    Target
    Research Area
  • HY-10244
    99.81%, HCV Replication Inhibitor
    HCV