{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,9,26]],"date-time":"2026-09-26T23:57:51Z","timestamp":1790467071214,"version":"4.1.0"},"posted":{"date-parts":[[2026,9,23]]},"group-title":"Microbiology","reference-count":48,"publisher":"openRxiv","license":[{"start":{"date-parts":[[2026,9,23]],"date-time":"2026-09-23T00:00:00Z","timestamp":1790121600000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by-nc-nd\/4.0\/"}],"funder":[{"name":"NIH","award":["AI171000"],"award-info":[{"award-number":["AI171000"]}]},{"name":"NIH","award":["AI183793"],"award-info":[{"award-number":["AI183793"]}]},{"name":"NIH","award":["AI179722"],"award-info":[{"award-number":["AI179722"]}]},{"id":[{"id":"https:\/\/ror.org\/018da3y22","id-type":"ROR","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2026,9,23]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                <jats:p>Human coronaviruses (HCoVs) cause pathogenic outcomes ranging from mild illness to severe respiratory disease. Determining how respiratory cells coordinate an early antiviral response is necessary to understand successful control of HCoV replication. The Type I Interferon (IFN) response is a major component of innate viral immunity. IFN signaling leads to the upregulation of hundreds of interferon-stimulated genes (ISGs) that can have antiviral activity, though the individual ISGs that are responsible for restricting mildly pathogenic HCoVs in the respiratory epithelium are not clear. Here, we developed a targeted CRISPR-Cas9 knockout sgRNA library (termed the respISG library), focusing on ISGs that are upregulated across a panel of respiratory cells, to screen for innate host factors that restrict the mildly pathogenic human coronavirus, HCoV-OC43. We executed cell death-based screens in immortalized human small airway epithelial cells and identified lymphocyte antigen 6E (LY6E) as a top hit. We confirmed LY6E activity in validation studies and found it regulates entry of various SARS-CoV-2 Spike-pseudotyped lentiviruses. In addition to demonstrating a key role for LY6E in blocking a variety of HCoVs in small airway epithelial cells, this study establishes a loss-of-function CRISPR-Cas9 screening platform for identification of HCoV antiviral ISGs in respiratory cells.<\/jats:p>\n                <jats:sec>\n                  <jats:title>Importance<\/jats:title>\n                  <jats:p>Interferon-stimulated genes (ISGs) are upregulated during the innate immune response and contribute to early antiviral defense. High-throughput genetic screening approaches to identify functional ISGs have revolutionized our understanding of interferon-mediated control of several viruses, including coronaviruses. However, screens that focus on cells most relevant for replication of the target virus tend to be less common. Here, we develop a loss-of-function screen based on ISGs from a panel of respiratory cell models and use it to probe antiviral genes that target HCoV-OC43 in human small airway epithelial cells. We find LY6E, a well-known viral effector, to be a dominant antiviral ISG and demonstrate that its activity, in a cell type that has broad relevance to HCoV pathogenesis, also extends to SARS-CoV-2 variants.<\/jats:p>\n                <\/jats:sec>","DOI":"10.64898\/2026.09.21.751853","type":"posted-content","created":{"date-parts":[[2026,9,24]],"date-time":"2026-09-24T04:05:23Z","timestamp":1790222723000},"source":"Crossref","is-referenced-by-count":0,"title":["A knockout screening platform for interferon-stimulated genes in respiratory cells identifies LY6E as a dominant antiviral effector against coronaviruses"],"prefix":"10.64898","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-2750-3228","authenticated-orcid":false,"given":"Caitlin I.","family":"Stoddard","sequence":"first","affiliation":[{"name":"Human Biology Division, Fred Hutchinson Cancer Center, Seattle, Washington, USA"},{"name":"Center for Vaccine Research, University of Pittsburgh, Pittsburgh, Pennsylvania, USA"},{"name":"Department of Microbiology and Molecular Genetics, University of Pittsburgh, Pittsburgh, Pennsylvania, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Morgan L.","family":"Litchford","sequence":"additional","affiliation":[{"name":"Human Biology Division, Fred Hutchinson Cancer Center, Seattle, Washington, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kevin","family":"Sung","sequence":"additional","affiliation":[{"name":"Public Health Sciences Division, Fred Hutchinson Cancer Center, Seattle, Washington, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Rebecca L.","family":"Hutcheson","sequence":"additional","affiliation":[{"name":"Center for Vaccine Research, University of Pittsburgh, Pittsburgh, Pennsylvania, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Joshua","family":"Marceau","sequence":"additional","affiliation":[{"name":"Human Biology Division, Fred Hutchinson Cancer Center, Seattle, Washington, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Taylor","family":"Johnson","sequence":"additional","affiliation":[{"name":"Institute for Public Health Genetics, School of Public Health, University of Washington, Seattle, Washington, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Akinobu","family":"Ota","sequence":"additional","affiliation":[{"name":"Department of Immunology, University of Washington, Seattle, WA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Nell E.","family":"Baumgarten","sequence":"additional","affiliation":[{"name":"Human Biology Division, Fred Hutchinson Cancer Center, Seattle, Washington, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"suffix":"Jr.","given":"Michael","family":"Gale","sequence":"additional","affiliation":[{"name":"Department of Immunology, University of Washington, Seattle, WA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Julie","family":"Overbaugh","sequence":"additional","affiliation":[{"name":"Human Biology Division, Fred Hutchinson Cancer Center, Seattle, Washington, USA"},{"name":"Public Health Sciences Division, Fred Hutchinson Cancer Center, Seattle, Washington, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"54368","reference":[{"key":"2026092616000729000_2026.09.21.751853v1.1","doi-asserted-by":"publisher","DOI":"10.1128\/JCM.00636-10"},{"key":"2026092616000729000_2026.09.21.751853v1.2","doi-asserted-by":"publisher","DOI":"10.1016\/j.ejim.2004.01.006"},{"key":"2026092616000729000_2026.09.21.751853v1.3","doi-asserted-by":"crossref","first-page":"567","DOI":"10.1146\/annurev-virology-092818-015756","article-title":"Interferon-Stimulated Genes: What Do They All Do?","volume":"6","year":"2019","journal-title":"Annu Rev Virol"},{"key":"2026092616000729000_2026.09.21.751853v1.4","unstructured":"Bastard P , Rosen LB , Zhang Q , Michailidis E , Hoffmann HH , Zhang Y , Dorgham K , Philippot Q , Rosain J , Beziat V , Manry J , Shaw E , Haljasmagi L , Peterson P , Lorenzo L , Bizien L , Trouillet- Assant S , Dobbs K , de Jesus AA , Belot A , Kallaste A , Catherinot E , Tandjaoui-Lambiotte Y , Le Pen J , Kerner G , Bigio B , Seeleuthner Y , Yang R , Bolze A , Spaan AN , Delmonte OM , Abers MS , Aiuti A , Casari G , Lampasona V , Piemonti L , Ciceri F , Bilguvar K , Lifton RP , Vasse M , Smadja DM , Migaud M , Hadjadj J , Terrier B , Duffy D , Quintana-Murci L , van de Beek D , Roussel L , Vinh DC , Tangye SG , et al. 2020. 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Protocol and Reagents for Pseudotyping Lentiviral Particles with SARS-CoV-2 Spike Protein for Neutralization Assays. 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