Fig. 4.

Transduction assays with pseudotyped virus of the interactions between viral spike proteins and human ACE2. Retroviral MLVs expressing β-galactosidase and pseudotyped with the NL63-CoV or SARS-CoV spike protein were used to infect hACE2-expressing HEK293T cells. Transduction efficiency of the pseudotyped viruses was measured by β-galactosidase assays. After mutations were introduced into the spike proteins or hACE2, the corresponding transduction efficiency was normalized against the transduction efficiency of viruses pseudotyped with wild-type spike proteins in cells expressing wild-type hACE2. Each experiment was repeated 6 times. The corresponding standard errors are shown. (A) Transduction of MLVs pseudotyped with NL63-CoV spike protein in hACE2-expressing cells. (B) Transduction of MLVs pseudotyped with SARS-CoV spike protein in hACE2-expressing cells. (C) Western blotting of coronavirus spike proteins and hACE2. The NL63-CoV and SARS-CoV spike proteins packaged in pseudotyped retroviruses both contained a C-terminal C9 tag, and the hACE2 expressed on the HEK293T cell surface contained a C-terminal HA tag. The expression levels of the spike proteins and hACE2 were detected by Western blotting using anti-C9 and anti-HA antibodies, respectively. The protein bands were quantified using software Image J (version 1.6).