The homozygous NPC1L1-EGFP mice have normal NPC1L1 expression pattern as well as cholesterol homeostasis on chow or high-cholesterol diets. Here, we genetically labelled endogenous NPC1L1 protein with EGFP in a knock-in mouse model, and demonstrated fluorescent visualization and evaluation of the endocytic vesicles of NPC1L1-cago during intestinal cholesterol absorption. Niemann-Pick C1-like 1 (NPC1L1) is a major membrane protein responsible for cholesterol absorption, in which the physiological role of vesicular endocytosis is still controversial, and it lacks a feasible tool to visualize and evaluate the endocytosis of NPC1L1 vesicles in vivo. Įxcessive cholesterol absorption from intestinal lumen contributes to the pathogenesis of hypercholesterolemia, which is an independent risk factor for atherosclerotic cardiovascular disease. Specific binding data were fit to a single class of AS-binding sites and are presented relative to the maximum receptor occupancy dog NPC1L1, 9, 10 and 11. TsA-201 cells were transfected with dog NPC1L1 (F) and chimeras 9 (), 10 (■), or 11 (OE) and incubated with increasing concentrations of AS, as indicated in Experimental Procedures.
(E) AS binding to dog/mouse loop C exchange chimeras 9-11. Specific binding data were fit to a single class of AS-binding sites and are presented relative to the maximum receptor occupancy dog NPC1L1, 7 and 8. TsA-201 cells were transfected with dog NPC1L1 (F) and loop C exchange chimeras 7 (OE) or 8 (■) and incubated with increasing concentrations of AS, as indicated in Experimental Procedures. (D) AS binding to dog/mouse loop C exchange chimeras 7-8. Specific binding was fit to a single class of AS-binding sites and is presented relative to the maximum receptor occupancy dog NPC1L1, 4, 5 and 6. TsA-201 cells transfected with dog NPC1L1 (F) and chimeras 4 (■), 5 () or 6 (F) were incubated with increasing concentrations of AS, as indicated in Experimental Procedures. (C) AS binding to dog/mouse chimeras 4-6.
Specific binding data were fit to a single class of AS-binding sites and are presented relative to the maximum receptor occupancy dog NPC1L1, 1, 2 and chimera 3. TsA-201 cells transfected with dog NPC1L1 (F) and chimeras 1 (), 2 (), or 3 (■) were incubated with increasing concentrations of AS, as indicated in Experimental Procedures. (B) AS binding to dog/mouse chimeras 1-3. SSD, between amino acids 629 and 806 is indicated by an arrow. Dog (orange) and mouse (blue) NPC1L1 sequences are shown, with their predicted TMDs indicated in dark color. Binding of AS to dog/mouse NPC1L1 chimeras.
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