Fig. 7: SARS-CoV-2 Spike, ORF7a and SERINC5 co-localize at the ERGIC and physically interact.
From: SARS-CoV-2 ORF7a potently inhibits the antiviral effect of the host factor SERINC5
a SARS-CoV-2 Spike (S), ORF7a, SERINC5 co-localize at the ERGIC. AD-293 cells cotransfected with plasmids expressing SERINC5-HA, EGFP-ERGIC53, SARS-CoV-2 ORF7a and SARS-CoV-2 S were subjected to immunostaining. Images were acquired using 63×/1.4 oil-immersion objective with a Leica TCS SP8 confocal microscope. Arrow heads indicate areas of colocalization, and the dotted line outlines the edges of the cells. Shown are confocal images revealing co-localization of SERINC5, SARS-CoV-2 S and ORF7a with ERGIC53 cellular marker (arrows). Scale bar = 10 µm. Representative deconvolved single Z-section images are shown. Table on the right shows quantitative analyses for co-localization between SERINC5, SARS-CoV-2 S or ORF7a and ERGIC53. Spearman’s rank correlation values (rs, mean ± SD) of a region of interest (ROI) defined by the presence of EGFP-ERGIC53 signal were calculated using the ImageJ (FIJI) Coloc2 plugin from 3 different images each from a different experiment. b SARS-CoV-2 S, ORF7a, SERINC5 form a complex. 293T cells were cotransfected with SARS-CoV-2 S, SERINC5, SARS-CoV-2 ORF7a or empty vector as indicated. Cells were harvested 48 h post transfection and lysates were immunoprecipitated with anti-SARS-CoV 2 S, anti-HA and anti-V5 antibodies followed by immunoblot analyses probing with anti-SARS-CoV-2 S, anti-HA (SERINC5), anti-V5 (SARS-CoV-2 ORF7a), and anti-Actin antibodies. Representative immunoblotting results are shown. Uncropped blots are in Source Data. All results are shown for n = 3 independent experiments. (SARS-CoV-2 ORF7a, ORF7a; SARS-CoV-2 S, S; SARS-CoV-2 Spike S2 subunit, S2).
