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Generation of a Sleeping Beauty Transposon-Based Cellular System for Rapid and Sensitive Screening for Compounds and Cellular Factors Limiting SARS-CoV-2 Replication. | LitMetric

AI Article Synopsis

  • - SARS-CoV-2, the virus causing COVID-19, is highly infectious, prompting the need for efficient methods to study its infectivity in lab settings, which traditionally are complex and time-intensive.
  • - Researchers developed a new cellular infection model using the Sleeping Beauty transposase system with A549 lung cells, which enhance virus susceptibility through the introduction of ACE2 and TMPRSS2 proteins.
  • - This innovative model allows for quick and effective monitoring of SARS-CoV-2 infections, enabling high-throughput experiments and the potential to identify key host factors that aid in viral replication.

Article Abstract

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of the acute respiratory disease COVID-19, which has become a global concern due to its rapid spread. The common methods to monitor and quantitate SARS-CoV-2 infectivity in cell culture are so far time-consuming and labor-intensive. Using the Sleeping Beauty transposase system, we generated a robust and versatile cellular infection model that allows SARS-CoV-2 infection experiments compatible for high-throughput and live cell imaging. The model is based on lung derived A549 cells, which show a profound interferon response and convenient cell culture characteristics. ACE2 and TMPRSS2 were introduced for constitutive expression (A549-AT). Subclones with varying levels of ACE2/TMPRSS2 were screened for optimal SARS-CoV-2 susceptibility. Furthermore, extensive evaluation demonstrated that SARS-CoV-2 infected A549-AT cells were distinguishable from mock-infected cells and already showed approximately 12 h post infection a clear signal to noise ratio in terms of cell roughness, fluorescence and a profound visible cytopathic effect. Moreover, due to the high transfection efficiency and proliferation capacity, Sleeping Beauty transposase-based overexpression cell lines with a second inducible fluorescence reporter cassette (eGFP) can be generated in a very short time, enabling the investigation of host and restriction factors in a doxycycline-inducible manner. Thus, the novel model cell line allows rapid and sensitive monitoring of SARS-CoV-2 infection and the screening for host factors essential for viral replication.

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Source
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8362758PMC
http://dx.doi.org/10.3389/fmicb.2021.701198DOI Listing

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