Figure 2.

AMDE-1-induced NCA does not require the participation of the canonical autophagy-specific class III PI-3 kinase. (a) WT-MEFs were treated by AMDE-1 (10 μM) with or without 3-MA (10 mM) or WM (1 μM) for 6 h, and then analyzed by immunoblotting. (b) A549 cells were treated as indicated (Rap, 1 μM; AMDE-1, 10 μM; 3-MA, 10 mM) for 6 h, followed by immunostaining with an anti-ATG16L1 antibody. Arrows indicate ATG16-positive dots. (c) Atg14-siRNA was transfected to wild- type MEFs for 48 h, followed by treatment with AMDE-1 or EBSS. (d and e) Wild-type and Beclin1 KD U251 cells expressing GFP-LC3 were treated with or without AMDE-1 (10 μM) for 6 h or EBSS for 2 h and then assessed for LC3-II formation by immunoblotting (d), or immunostaining (e). Bar=25 μm