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Dual reporter cells expressing the SARS-CoV-2 receptors ACE2 & TMPRSS2

Reporter systems in A549-Dual™-derived cells

InvivoGen offers a new series of A549-Dual™ cell lines, specifically designed for COVID-19 studies:

— A549-Dual™ hACE2-TMPRSS2 cells
— A549-Dual™ KO-MDA5 hACE2-TMPRSS2 cells
— A549-Dual™ KO-RIG-I hACE2-TMPRSS2 cells

These cells derive from the human A549 lung carcinoma cell line. They stably overexpress the genes encoding for the SARS-CoV-2 receptors, human ACE2, and TMPRSS2.

These cells also express two inducible reporter genes, allowing the concomitant study of the IRF and NF-κB pathways, by monitoring the Lucia luciferase and SEAP (secreted embryonic alkaline phosphatase) activities, respectively. In addition, these cells either express MDA5 (Melanoma Differentiation Associated gene 5) and RIG-I (Retinoic Acid Inducible protein 1) endogenously or are knockout for each gene, individually. MDA5 and RIG-I are sensors of viral RNA that have been reported to participate in the immune response to SARS-CoV-2 infection [1-3].

The A549 cell line is commonly used for the study of respiratory infections. SARS-CoV-2, the causative agent of coronavirus disease-19 (COVID-19), gains entry into the human lung epithelium through the interaction of the virus Spike protein with the host ACE2 and TMPRSS2 receptors [3-5]. A549 cells express negligible levels of ACE2 and no TMPRSS2 and thus are poorly permissive to infection by SARS-CoV-2 or Spike pseudotyped lentiviral particles [4-6, in-house data]. To increase their permissivity, A549-Dual™ cells have been stably transfected with the human ACE2 and TMPRSS2 genes. In contrast to A549-Dual™ cells, A549-Dual™ hACE2-TMPRSS2 cells are highly permissive to Spike pseudotyped lentiviral particles as visualized by the expression of the lentiviral GFP transgene (see Figures).

Key Features:

Overexpression of human ACE2 and TMPRSS2
Highly permissive to SARS-CoV-2 Spike pseudotyped lentiviral particles
Readily assessable Lucia luciferase and SEAP reporter activity for the IRF and NF-κB activation, respectively
Endogenous or knockout expression of human MDA5 or human RIG-I

Applications:

Study of SARS-CoV-2 RNA sensing pathways
Screening of small molecule inhibitors and/or neutralizing antibodies of the ACE2‑Spike interaction
Screening of small molecule inhibitors and/or neutralizing antibodies of the TMPRSS2 surface protease
Comparative studies of the effects of drugs targeting ACE2 and/or TMPRSS2 on the SARS-CoV-2 infection and cellular signaling outcomes

Learn more about SARS-CoV-2 infection cycle, immune responses, and potential therapeutics.

References

1. Ying X. et al., 2021. MDA5 governs the innate immune response to SARS-CoV-2 in lung epithelial cells. Cell Reports. 34:108628.
2. Rebendenne A. et al., 2021. SARS-CoV-2 triggers MDA-5-dependent interferon response which is unable to control replication in lung epithelial cells. J. Virol. doi:10.1128/JVI.02415-20.
3. Wu J, et al., 2021. SARS-CoV-2 ORF9b inhibits RIG-I MAVS antiviral signaling by interrupting K63-linked ubiquitination of NEMO. Cell Reports. 34(7):108761.
4. Chen H. et al., 2020. SARS-CoV-2 activated lung epithelia cell proinflammatory signaling and leads to immune dysregulation in COVID-19 patients by single-cell sequencing. medRxiv: DOI 10.1101/2020.05.08.20096024.
5. Hoffmann M. et al., 2020. SARS-CoV-2 cell entry depends on ACE2 and TMPRSS2 and is blocked by a clinically proven protease inhibitor. Cell. 181:1-16.
6. Matsuyama S. et al., 2020. Enhanced isolation of SARS-CoV-2 by TMPRSS2-expressing cells. PNAS. 117(13):7001-7003.

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