Both cholesterol and a-tocopherol are essential lipophilic nutrients for humans and animals. Although cholesterol in excess causes severe problems such as coronary heart disease, it is a necessary component of cell membranes and is the precursor for the biosynthesis of steroid hormones and bile acids. Niemann Pick Cl-like 1 (NPC1L1) is a cholesterol transporter that is highly expressed in the small intestine and liver in humans and plays an important role in cholesterol homeostasis. Cholesterol promotes NPC1L1 endocytosis, which is an early step in cholesterol uptake. Furthermore, alpha-tocopherol is the most active form of vitamin E, and sufficient amounts of vitamin E are critical for health. It has been reported that NPC1L1 mediates alpha-tocopherol absorption; however, the mechanisms underlying this process are unknown. In this study, we found that treatment of cells that stably express NPC1L1-GFP with alpha-tocopherol promotes NPC1L1 endocytosis, and the NPC1L1 inhibitor, ezetimibe, efficiently prevents the alpha-tocopherol-induced endocytosis of NPC1L1. Cholesterol binding to the N-terminal domain (NTD) of NPC1L1 (NPC1L1-NTD) is essential for NPC1L1-mediated cholesterol absorption. We found that alpha-tocopherol competitively binds NPC1L1-NTD with cholesterol. Furthermore, when cells stably expressed NPC1L1ANTD-GFP, alpha-tocopherol could not induce the endocytosis of NPC1L1ANTD. Taken together, these results demonstrate that NPC1L1 recognizes alpha-tocopherol via its NTD and mediates alpha-tocopherol uptake through the same mechanism as cholesterol absorption. (C) 2017 Elsevier Inc. All rights reserved.