{
  "abstract": "Respiratory viruses, including coronaviruses, generally initiate infection in the upper respiratory tract, where mechanistic studies of host-virus interactions require models amenable to genetic manipulation. Human nasal air-liquid interface (ALI) cultures are realistic models of the nasal epithelium, but robust CRISPR/Cas9 engineering workflows for this primary cell system have been lacking. Here, we established an integration-free CRISPR/Cas9 workflow for genetic engineering of human nasal basal cells, which are subsequently differentiated into gene-edited ALI cultures. GFP-based sorting of RNP-transfected nasal basal cells substantially increases editing, achieving knockout efficiencies of 70-90% across gene targets (B2M, SLC35A1, ACE2, and TMPRSS2). Genetic disruption of ACE2 markedly reduces infection by SARS-CoV-2 and hCoV-NL63, confirming its important role for both viruses in the human nasal epithelium. TMPRSS2 knockout substantially reduces replication of both viruses, demonstrating SARS-CoV-2 depending on TMPRSS2 in the human nasal epithelium and providing genetic evidence in primary human cell cultures that hCoV-NL63 relies on TMPRSS2. This platform enables systematic genetic perturbation in a physiologically relevant model of the primary site of respiratory virus infection, validates ACE2 and TMPRSS2 as targets for medical countermeasures against SARS-CoV-2 and hCoV-NL63, and provides a versatile approach to identify host dependency factors for both circulating and emerging respiratory viruses.",
  "abstract_shown": true,
  "affiliations": [
    "Duke-NUS Medical School"
  ],
  "article_type": "New results",
  "arxiv": null,
  "author_total": 14,
  "authors": [
    {
      "name": "S. W. van Leur"
    },
    {
      "name": "J. T. S. Chu"
    },
    {
      "name": "X. Wu"
    },
    {
      "name": "A. Rawat"
    },
    {
      "name": "N. K.-W. Yeo"
    },
    {
      "name": "T. W. X. Lee"
    },
    {
      "name": "M. G. Mah"
    },
    {
      "name": "Y.-J. Chen"
    },
    {
      "name": "C. S. Y. Ng"
    },
    {
      "name": "C. Trupin"
    },
    {
      "name": "A. C.-Y. Hsu"
    },
    {
      "name": "G. J. Smith"
    },
    {
      "name": "Y. S. Ooi"
    },
    {
      "affs": [
        0
      ],
      "name": "M. M. Lamers"
    }
  ],
  "bibtex": "https://research.draconitic.com/paper/fwtjqbar5ce67.bib",
  "cited_by": 0,
  "doi": "10.64898/2026.10.06.756366",
  "fulltext_url": "https://www.biorxiv.org/content/10.64898/2026.10.06.756366v1.full",
  "id": "fwtjqbar5ce67",
  "issns": [],
  "landing_url": "https://www.biorxiv.org/content/10.64898/2026.10.06.756366v1",
  "language": null,
  "licence": "CC-BY",
  "licence_label": "CC BY",
  "licence_source": "article",
  "licence_url": "https://creativecommons.org/licenses/by/4.0/",
  "metadata_licence": null,
  "native_subjects": [
    [
      "biorxiv",
      "microbiology"
    ],
    [
      "openalex",
      "1312|CRISPR and Genetic Engineering"
    ]
  ],
  "openalex": "W7221135812",
  "pdf_url": "https://www.biorxiv.org/content/10.64898/2026.10.06.756366v1.full.pdf",
  "pmcid": null,
  "pmid": null,
  "published": "2026-10-06",
  "publisher": null,
  "ris": "https://research.draconitic.com/paper/fwtjqbar5ce67.ris",
  "sources": [
    {
      "first_seen": "2026-10-30T12:26:42Z",
      "native_id": "10.64898/2026.10.06.756366",
      "source": "biorxiv",
      "url": "https://www.biorxiv.org/content/10.64898/2026.10.06.756366v1"
    }
  ],
  "subjects": [
    {
      "field": "biology",
      "origin": "biorxiv:microbiology",
      "subfield": "microbiology"
    }
  ],
  "title": "An integration-free CRISPR platform for genetic dissection of respiratory virus host dependencies in human nasal epithelium",
  "title_text": "An integration-free CRISPR platform for genetic dissection of respiratory virus host dependencies in human nasal epithelium",
  "type": "preprint",
  "updated_at": "2026-10-30T12:26:42Z",
  "url": "https://research.draconitic.com/paper/fwtjqbar5ce67",
  "venue": "bioRxiv",
  "versions": []
}