Fig. 2: mGluR2 is required for internalization of SARS-CoV-2 but not for cell binding.

a, b SARS-CoV-2 binding (a) and internalization (b) assays were performed in mGluR2-silenced Vero-E6 cells and Caco-2 cells. Viral binding or internalization was quantified by normalization to the respective scrambled siRNA-transfected cells. c Vero-E6 cells were treated as described in a, except they were not treated with acid buffer/trypsin. Cell nuclei (blue), SARS-CoV-2 S protein (green). Left: representative images are shown, and the dashed box is magnified at the indicated location of the same image. Right: the fluorescence signal of SARS-CoV-2 particles was quantified by immunofluorescence staining using ZEN software. The relative fluorescence of cell-bound HRB25 under permeabilized or unpermeabilized conditions was quantified by normalization to the corresponding scrambled siRNA-transfected cells. The circles represent individual data points. n, the number of quantified cells. Data represent the sum of three independent experiments. d Surface expression level of mGluR2 was detected by using flow cytometry after infection with HRB25 at 37 °C for 30 min under unpermeabilized conditions in Vero-E6 cells. e, f Multiplex immunofluorescence was performed in Vero-E6 cells (e) and HPAE cells (f). Colocalization of mGluR2 (green), SARS-CoV-2 N protein (red), and clathrin (purple) was observed and quantified. The yellow arrows indicate the representative colocalization of mGluR2 (green), SARS-CoV-2 N protein (red), and clathrin (purple), shown in three dimensions. The data shown are representative results from three independent experiments (a, b, d n = 3), means ± SD, Student’s t-test, ns, not significant, *P < 0.05, ***P < 0.001.