Hnrnp f

Antibodies used for Western blotting and immunofluorescence assays were as follows: hnRNPM (OriGene), hnRNPF (Santa Cruz Biotechnology, Inc.), hnRNPH1/H2 (Santa Cruz Biotechnology, Inc.), p-Smad3 (Cell Signaling), Smad3 (Cell Signaling), CD44 (R&D Systems and Santa Cruz Biotechnology, Inc.), E-cadherin (Cell Signaling Technology), γ-catenin (Cell Signaling Technology), occludin (Santa Cruz Biotechnology, Inc.), N-cadherin (BD), vimentin (NEOMarkers), β-actin (Sigma-Aldrich), and GAPDH (Millipore). The ESRP1 antibody was a gift from Dr. R. Carstens (University of Pennsylvania).

Cells were lysed with SDS sample buffer (50 mM Tris at pH 7.4, 100 mM NaCl, 1% Triton X-100, 0.1% SDS, 1% sodium deoxycholate, 1× proteinase inhibitor cocktail [Biotools]), resolved by SDS-PAGE, transferred to nitrocellulose membranes, and incubated with primary antibodies detecting CDH1 (1:1000; Cell Signaling), hnRNP H1 (1:1000; Bethyl Laboratories), hnRNP F (1:1500; Santa Cruz Biotechnology), GAPDH (1:20,000; Sigma), or ACTB (1:30,000; Sigma) overnight at 4°C. Secondary antibodies HRP-conjugated anti-mouse and anti-rabbit IgG (Sigma) were used at 1:10,000 for 2 h at room temperature. Following washes with PBST (1× PBS, 0.1% Tween-20), ECL-Prime (GE Healthcare) was used to visualize the chemiluminescence signal, and ImageJ software was used for quantification.