Cholesterol-regulated translocation of NPC1L1 to the cell surface facilitates free cholesterol uptake.
Although NPC1L1 is required for intestinal cholesterol absorption, data demonstrating mechanisms by which this protein facilitates the process are few. In this study, a hepatoma cell line stably expressing human NPC1L1 was established, and cholesterol uptake was studied. A relationship between NPC1L1 intracellular trafficking and cholesterol uptake was apparent. At steady state, NPC1L1 proteins localized predominantly to the transferrin-positive endocytic recycling compartment, where free cholesterol also accumulated as revealed by filipin staining. Interestingly, acute cholesterol depletion induced with methyl-beta-cyclodextrin stimulated relocation of NPC1L1 to the plasma membrane, preferentially to a newly formed "apical-like" subdomain. This translocation was associated with a remarkable increase in cellular cholesterol uptake, which in turn was dose-dependently inhibited by ezetimibe, a novel cholesterol absorption inhibitor that specifically binds to NPC1L1. These findings define a cholesterol-regulated endocytic recycling of NPC1L1 as a novel mechanism regulating cellular cholesterol uptake.
Yu
Liqing
L
Department of Pathology, Wake Forest University School of Medicine, Winston-Salem, North Carolina 27157-1040, USA. lyu@wfubmc.edu
Bharadwaj
Shantaram
S
Brown
J Mark
JM
Ma
Yinyan
Y
Du
Wei
W
Davis
Matthew A
MA
Michaely
Peter
P
Liu
Pingsheng
P
Willingham
Mark C
MC
Rudel
Lawrence L
LL
D000818:Animals; D000924:Anticholesteremic Agents / Q000494:pharmacology; D001384:Azetidines / Q000494:pharmacology; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000378:metabolism*; D002784:Cholesterol / Q000737:chemistry* / Q000378:metabolism*; D004705:Endocytosis / Q000502:physiology; D000069438:Ezetimibe; D017930:Genes, Reporter; D006056:Golgi Apparatus / Q000378:metabolism; D000882:Haplorhini; D006801:Humans; D008099:Liver / Q000378:metabolism; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase / Q000502:physiology; D008565:Membrane Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D026901:Membrane Transport Proteins; D021381:Protein Transport / Q000502:physiology; D011506:Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014168:Transferrin / Q000378:metabolism
D000924:Anticholesteremic Agents; D001384:Azetidines; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; C410800:NPC1L1 protein, human; D011506:Proteins; D011993:Recombinant Fusion Proteins; D014168:Transferrin; D002784:Cholesterol; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase; D000069438:Ezetimibe