Fig. 5

ENO binds to the RPSA IDR to form co-condensates. A and B HEK-293 T cells co-overexpressing mCherry-ENO and the indicated proteins for 36 h. A Cell lysates were immunoprecipitated with the antibody against GFP, followed by western blot analysis using an antibody against mCherry. B Co-localization of ENO and different RPSA truncations were observed by immunofluorescence. Representative images were shown (scale bar = 40 μm, left). Co-localization analysis of the selected arrowed areas used Image J (ENO and indicated protein co-localization analysis, right). C and D EGFP-ENO or ENO (20 μg/mL) protein was added to HCMEC/D3 cells. After 24 h, cells were treated with or without 1,6-Hex. C Adhesion of ENO to the cell surface was determined by immunofluorescence staining using the antibody against RPSA. Representative confocal images are shown (scale bar = 40 μm) and magnified (scale bar = 10 μm). D The amount of ENO protein acting on the cell was determined by western blotting using the antibody against his. Quantification was performed by Image J as mean ± SD (n = 3 biologically independent samples). ** for P < 0.01; one-way ANOVA with Tukey’s test