A pseudovirus-based method to dynamically mimic SARS-CoV-2-associated cell-to-cell fusion and transmission

SARS-CoV-2 has caused the global tremendous loss and continues to evolve to generate variants. Entry of SARS-CoV-2 into the host cells is primarily mediated by Spike (S), which binds to the host receptor hACE2 and initiates virus-cell membrane fusion. Cell fusion contributes to viral entry, cell-to-cell transmission and tissue damage in COVID-19 patients. Many reporter assays have been developed to study S-mediated cell fusion by equally coculturing S-expressing cells and hACE2-positive cells. However, these strategies cannot fully simulate cell-to-cell fusion and transmission of SARS-CoV-2 infection, in which virions from a single target cell transmit to the neighbor cells and induce syncytia formation. Here, we design a pseudovirus-based method to dynamically mimic cell-to-cell fusion and transmission of SARS-CoV-2. We coculture a small number of pseudovirus-producing 293FT cells and a large number of hACE2-expressing 293T cells, and demonstrate that a single cell producing S-pseudotyped virions can induce significant syncytia of hACE2-positive cells. This pseudovirus-based method is a powerful tool to screen and estimate potential inhibitors of S-driven syncytia. Moreover, this strategy can also be utilized to explore fusogenic ability of SARS-CoV-2 variants. Together, the pseudovirus-based method we report here will be beneficial to drug screening and scientific research against SARS-CoV-2 or future emerging coronavirus.