Clathrin-mediated endocytosis is required for ANE 30-100K-induced autophagy

Background We identified an autophagy-inducing areca nut (AN) ingredient (AIAI) in the 30-100 kDa fraction of AN extract (ANE 30-100K). This study was to analyze the role of endocytosis in ANE 30-100K-induced autophagy. Methods We used benzyl alcohol, dynasore, and shRNA of clathrin and dynamin to assess whether ANE 30-100K-induced cytotoxicity and accumulation of microtubule-associated protein 1 light chain 3 (LC3)-II were affected in oral (OECM-1) and esophageal (CE81T/VGH) carcinoma cells. Results Both benzyl alcohol and dynasore effectively reduced ANE 30-100K-induced cytotoxicity and LC3-II accumulation in OECM-1 and CE81T/VGH cells. Downregulated protein expression of both clathrin and dynamin by their shRNA also significantly attenuated ANE 30-100K-induced elevation of LC3-II levels in CE81T/VGH cells. Conclusions These results indicate that AIAI may be engulfed by cells through clathrin-mediated endocytosis, which promotes the execution of the following autophagy program. This article is protected by copyright. All rights reserved.