Fig 1: MFSD1 promotes recycling and stabilization of inactive ß1 integrin. (A) Western blot of ß1 integrin immunoprecipitated by HMb1-1 or 9EG7 antibodies from total cell lysates or intact cell surface (n = 2). (B) Western blot of cell-surface biotinylated and immunoprecipitated ß1 integrin for biotinylation (streptavidin-HRP) at 0 and 12 h time point post-treatment (biotinylation). ITGb1 blot displays the input of ß1 integrin (n = 4). (C) Densitometric quantification of biotinylated ß1 integrin (n = 4). (D) Western blot of ß1 integrin upon brefeldin A treatment for 12 h (rate of ß1 integrin degradation experiment). GAPDH serves as a loading control. A representative experiment is shown. (E) Densitometric quantification of ITGb1 from brefeldin A-treated cells for 12 h (n = 4). (F) Flow cytometric analysis of MC-38 WT and MFSD1-/- cells treated with recycling inhibitor primaquine (PQ) or control (n = 7). (G) Co-immunofluorescence of MC-38 MFSD1-eGFP cells with an antibody detecting either the total (Itgb1) (n = 14) or the active conformation (9EG7) (n = 6) of ß1 integrin, respectively. The area indicated by a dashed square is shown magnified to the right. Arrows indicate co-localization. Bar = 10 µm. (H) Co-localization analysis of MFSD1-eGFP with the indicated ß1 integrin antibodies (n = 10). *p < 0.05; ****p < 0.0001; ns, not significant.