NPC1L1 (Niemann-Pick C1-like 1) mediates sterol-specific unidirectional transport of non-esterified cholesterol in McArdle-RH7777 hepatoma cells.
Recent evidence suggests that NPC1L1 (Niemann-Pick C1-like 1) is critical for intestinal sterol absorption in mice, yet mechanisms by which NPC1L1 regulates cellular sterol transport are lacking. In the study we used a McArdle-RH7777 rat hepatoma cell line stably expressing NPC1L1 to examine the sterol-specificity and directionality of NPC1L1-mediated sterol transport. As previously described, cholesterol-depletion-driven recycling of NPC1L1 to the cell surface facilitates cellular uptake of non-esterified (free) cholesterol. However, it has no impact on the uptake of esterified cholesterol, indicating free sterol specificity. Interestingly, the endocytic recycling of NPC1L1 was also without effect on beta-sitosterol uptake, indicating that NPC1L1 can differentiate between free sterols of animal and plant origin in hepatoma cells. Furthermore, NPC1L1-driven free cholesterol transport was unidirectional, since cellular cholesterol efflux to apolipoprotein A-I, high-density lipoprotein or serum was unaffected by NPC1L1 expression or localization. Additionally, NPC1L1 facilitates mass non-esterified-cholesterol uptake only when it is located on the cell surface and not when it resides intracellularly. Finally, NPC1L1-dependent cholesterol uptake required adequate intracellular K(+), yet did not rely on intracellular Ca(2+), the cytoskeleton or signalling downstream of protein kinase A, protein kinase C or pertussis-toxin-sensitive G-protein-coupled receptors. Collectively, these findings support the notion that NPC1L1 can selectively recognize non-esterified cholesterol and promote its unidirectional transport into hepatoma cells.
Brown
J Mark
JM
Department of Pathology, Section on Lipid Sciences, Wake Forest University School of Medicine, Medical Center Blvd., Winston-Salem, NC 27157-1040, U.S.A.
Rudel
Lawrence L
LL
Yu
Liqing
L
D000818:Animals; D001692:Biological Transport; D002118:Calcium / Q000378:metabolism; D006528:Carcinoma, Hepatocellular / Q000378:metabolism*; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000378:metabolism; D002784:Cholesterol / Q000378:metabolism*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D004952:Esters / Q000378:metabolism; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D026901:Membrane Transport Proteins / Q000235:genetics / Q000378:metabolism*; D009154:Mutation / Q000235:genetics; D009542:Niemann-Pick Diseases / Q000235:genetics / Q000378:metabolism; D019301:Oleic Acid / Q000378:metabolism; D011188:Potassium / Q000378:metabolism; D011493:Protein Kinase C / Q000378:metabolism; D051381:Rats; D043562:Receptors, G-Protein-Coupled / Q000378:metabolism; D015398:Signal Transduction; D012855:Sitosterols / Q000378:metabolism
D004952:Esters; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; C482444:NPC1L1 protein, rat; D043562:Receptors, G-Protein-Coupled; D012855:Sitosterols; D019301:Oleic Acid; C025473:gamma-sitosterol; D002784:Cholesterol; D017868:Cyclic AMP-Dependent Protein Kinases; D011493:Protein Kinase C; D011188:Potassium; D002118:Calcium