Figure 2.

Introduction of human ACE2 residues into rat ACE2 converts rat ACE2 to an efficient SARS-CoV receptor. (A) HEK293T cells transfected with plasmids encoding the indicated human and rat ACE2 variants or with vector alone were analyzed as in Figure 1 (light gray), or by flow cytometry (dark gray). Flow cytometry values indicate the ratio of mean fluorescence intensity (m.f.i.) observed using S1-Ig to that using the 9E10 antibody, which recognizes a tag present at the amino-terminus of each ACE2 variant. Error bars indicate the range of two or more experiments. (B) Representative example of an immunoprecipitation experiment used in (A). (C) Murine leukemia viruses (MLV) expressing green fluorescent protein (GFP), lacking its endogenous envelope glycoprotein (MLV-GFP), and pseudotyped with the S protein of SARS-CoV (TOR2 isolate) were incubated with HEK293T cells transfected with plasmids encoding the indicated human or rat ACE2 variants. Amount of ACE2-expressing plasmid was adjusted to maintain comparable receptor expression levels, as indicated, by flow cytometry using the antibody 9E10 that recognizes an amino-terminal myc tag on these receptors. GFP expression in cells was quantified by flow cytometry to measure infection of cells by pseudotyped viruses. Error bars indicate range of two experiments.