Fig. 4

Immunoblotting of ACE2 as well as S and S-related proteins in HEK293(S + ACE2) cells treated with quercetin. (A) Diagrammatic representation of the S-GFP fusion protein and its proteolytic fragment. Acronyms: NTD, N-terminal domain; RBD, receptor binding domain; FP, fusion peptide; HR, heptad region, TM, transmembrane domain, GFP, green fluorescent protein. (B) Representative Immunoblot of cells extracts transfected with empty vector (EV) (lane 1), vector for expression of S-GFP (lane 8), ACE2 (lane 9), or both S-GFP and ACE2 (lanes 2–7) with DMSO (lane 2) or increasing concentrations of quercetin (lanes 3–7); (a) The blot was immuno-probed successively for GFP and β-actin; (b) a parallel blot of the same cell extracts was immune-probed successively for ACE2 and β-actin antibody. (C) Densitometry of immunoreactive bands in three separate experiments was conducted. S, S2 and S2’and ACE2 values were normalized by those of β-actin (a-c, f). The ratios S2/S (d) and S2’/S2 (e) were computed as measures of the proteolytic conversion S to S2 and S2 to S2’, respectively. Values are expressed as means ± SD. * < 0.05, ** < 0.005