Smarce1

Lab-Tek Chamber Slide w/Cover Glass Slide Sterile (Thermo, Waltham, MA, USA) was used to culture and transfect Caco2 cells, with 100 μL of medium per well. Primary antibodies of SMARCE1 (Abcam, Cambridge, UK, 1:100) and tfec (Santa Cruz, USA, 1:50) were used for double immunofluorescence. The cells were fixed with 4% formalin and incubated with specific primary antibodies overnight at 4°C. Primary antibody binding was visualized using fluorescein-labeled donkey anti-rabbit or anti-mouse IgG (Invitrogen, Carlsbad, CA, USA, 1:500) for 1 h at 25°C. DAPI (Solarbio, Beijing, China) was used for nuclear staining. Images were captured using a confocal laser scanning microscope (Olympus FV1000, Japan).

The transfected control Caco2 cells were lysed in RIPA buffer (150 mM NaCl; 50 mM Tris-Cl, pH 8; 1% NP-40; 0.5% deoxycholate; 0.1% SDS) (Beyotime, Shanghai, China) with Protease Inhibitor Cocktail (100×) (Thermo Scientific, MA, USA). The resulting samples were separated using 10% SDS-PAGE (Invitrogen, Carlsbad, CA, USA) and transferred to polyvinylidene fluoride (PVDF) membranes. After blocking for 1 h in 5% non-fat milk in TBST at 25°C, the membranes were incubated overnight at 4 °C with specific primary antibodies, including SMARCE1 (Abcam, Cambridge, UK, 1:1000), tfec (Santa Cruz, USA,1:500), and β-tubulin (Sigma, USA, 1:10,000). Primary antibody binding was visualized using horseradish peroxidase-conjugated goat anti-rabbit or anti-mouse IgG (Sigma, USA, 1:10,000) for 1 h at 25°C. Signal intensities were measured using a Chemiluminescent HRP substrate (Yamei, Shanghai, China) and imaged using ImageJ v1.8.0 (NIH, Bethesda, MD, USA).