Autophagy inhibitor 3 methyladenine

Hakai antibody (Invitrogen, Carlsbad, CA, USA) was used for western-blotting, and Hakai-2498 antibody used for immunohistochemistry was kindly provided by Dr. Fujita and Hakai antibody (Bethyl, Montgomery, TX, USA) was used for immunoprecipitation^{12 (link)}. The rest of antibodies used for western-blotting are FASN antibody (Santa Cruz, Dallas, TX, USA), LC3 A/B antibody (Cell Signaling, Leiden, The Netherlands), E-cadherin antibody (BD Trans Lab, Franklin Lakes, NJ, USA), β-catenin antibody (Cell Signaling, Leiden, The Netherlands), GAPDH antibody (Invitrogene) and mouse and rabbit secondary antibodies (GE Healthcare, Chicago, IL, USA). Proteasome inhibitor MG132 (Sigma-Aldrich, St. Louis, MO, USA) was added for 6 h using 10 µM and 30 µM. Lysosome degradation inhibitor Chloroquine (Sigma-Aldrich, St. Louis, MO, USA), was added for 24 h at 50 µM and for 6 h at 100 µM. Autophagy inhibitor 3-Methyladenine (Sigma-Aldrich, St. Louis, MO, USA) was added for 24 h at 5 mM and 10 mM. Protein synthesis inhibitor cycloheximide (Sigma-Aldrich) was used at 10 μg/mL for the indicated times.