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NF-κB inhibitor alpha controls SARS-CoV-2 infection in ACE2-overexpressing human airway organoids

Affiliations
  • 1Gladstone Institute of Virology, San Francisco, CA, USA.
  • 2Biomedical Sciences Graduate Program, University of California San Francisco, San Francisco, CA, USA.
  • 3Chan Zuckerberg Biohub, San Francisco, CA, USA.
  • 4Center for Computational Biology, University of California, Berkeley, Berkeley, CA, USA.
  • 5Medical Scientist Training Program, University of California San Francisco, San Francisco, CA, USA.
  • 6ImmunoX CoLabs, University of California San Francisco, San Francisco, CA, USA.
  • 7Department of Microbiology and Immunology, University of California, San Francisco, San Francisco, CA, USA.
  • 8Medical Service, San Francisco VA Healthcare System, San Francisco, CA, USA.
  • 9Department of Medicine and Department of Anesthesia, Cardiovascular Research Institute, University of California San Francisco, San Francisco, CA, USA.
  • 10Department of Anatomy, University of California San Francisco, San Francisco, CA, USA.
  • 11Division of Pulmonary and Critical Care, Department of Medicine, University of California San Francisco, San Francisco, CA, USA.
  • 12Center for Computational Biology, University of California, Berkeley, Berkeley, CA, USA. nir.yosef@weizmann.ac.il.
  • 13Department of Systems Immunology, Weizmann Institute of Science, Rehovot, Israel. nir.yosef@weizmann.ac.il.
  • 14Gladstone Institute of Virology, San Francisco, CA, USA. melanie.ott@gladstone.ucsf.edu.
  • 15Chan-Zuckerberg Biohub, San Francisco, CA, USA. melanie.ott@gladstone.ucsf.edu.
  • 16Department of Medicine, University of California, San Francisco, San Francisco, CA, USA. melanie.ott@gladstone.ucsf.edu.

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July 3, 2024

Abstract

As SARS-CoV-2 continues to spread worldwide, tractable primary airway cell models that recapitulate the cell-intrinsic response to arising viral variants are needed. Here we describe an adult stem cell-derived human airway organoid model overexpressing the ACE2 receptor (ACE2-OE) that supports robust viral replication while maintaining 3D architecture and cellular diversity of the airway epithelium. ACE2-OE organoids were infected with SARS-CoV-2 variants and subjected to single-cell RNA-sequencing. Interferon-lambda was upregulated in cells with low-level infection while the NF-kB inhibitor alpha gene (encoding IkBa) was consistently upregulated in infected cells, and its expression positively correlated with infection levels. Confocal microscopy showed more IkBa expression in infected than bystander cells, but found concurrent nuclear translocation of NF-kB that IkBa usually prevents. Overexpressing a nondegradable IkBa mutant reduced NF-kB translocation and increased viral infection. These data demonstrate the functionality of ACE2-OE organoids in SARS-CoV-2 research and underscore that the strength of the NF-kB feedback loop in infected cells controls viral replication.