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Flotillins play an essential role in Niemann-Pick C1-like 1-mediated cholesterol uptake.

ABSTRACT: Dietary absorption is a major way for mammals to obtain cholesterol, which is mediated by Niemann-Pick C1-like 1 (NPC1L1) via vesicular endocytosis. One fundamental question in this process is how free cholesterol is efficiently taken up through the internalization of NPC1L1. Using exogenously expressed NPC1L1-EGFP, we show that the lipid raft proteins flotillins associate with NPC1L1 and their localization is regulated by NPC1L1 during intracellular trafficking. Furthermore, flotillins are essential for NPC1L1-mediated cellular cholesterol uptake, biliary cholesterol reabsorption, and the regulation of lipid levels in mice. Together with NPC1L1, they form cholesterol-enriched membrane microdomains, which function as carriers for bulk of cholesterol. The hypocholesterolemic drug ezetimibe disrupts the association between NPC1L1 and flotillins, which blocks the formation of the cholesterol-enriched microdomains. Our findings reveal a functional role of flotillins in NPC1L1-mediated cholesterol uptake and elucidate the formation of NPC1L1-flotillins-postive cholesterol-enriched membrane microdomains as a mechanism for efficient cholesterol absorption.

SUBMITTER: Ge L

PROVIDER: S-EPMC3021008 | biostudies-literature | 2011 Jan

REPOSITORIES: biostudies-literature

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Flotillins play an essential role in Niemann-Pick C1-like 1-mediated cholesterol uptake.

Ge Liang L Qi Wei W Wang Li-Juan LJ Miao Hong-Hua HH Qu Yu-Xiu YX Li Bo-Liang BL Song Bao-Liang BL

Proceedings of the National Academy of Sciences of the United States of America 20101227 2

Dietary absorption is a major way for mammals to obtain cholesterol, which is mediated by Niemann-Pick C1-like 1 (NPC1L1) via vesicular endocytosis. One fundamental question in this process is how free cholesterol is efficiently taken up through the internalization of NPC1L1. Using exogenously expressed NPC1L1-EGFP, we show that the lipid raft proteins flotillins associate with NPC1L1 and their localization is regulated by NPC1L1 during intracellular trafficking. Furthermore, flotillins are esse ...[more]

PMID: 21187433

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Project description:UNLABELLED:Recent work demonstrated that the Niemann-Pick C1 (NPC1) protein is an essential entry receptor for filoviruses. While previous studies focused on filovirus entry requirements of NPC1 in vitro, its roles in filovirus replication and pathogenesis in vivo remain unclear. Here, we evaluated the importance of NPC1, and its partner in cholesterol transport, NPC2, by using a mouse model of Ebolavirus (EBOV) disease. We found that, whereas wild-type mice had high viral loads and succumbed to EBOV infection, Npc1(-/-) mice were entirely free of viral replication and completely protected from EBOV disease. Interestingly, Npc1(+/-) mice transiently developed high levels of viremia, but were nevertheless substantially protected from EBOV challenge. We also found Npc2(-/-) mice to be fully susceptible to EBOV infection, while Npc1(-/-) mice treated to deplete stored lysosomal cholesterol remained completely resistant to EBOV infection. These results provide mechanistic evidence that NPC1 is directly required for EBOV infection in vivo, with little or no role for NPC1/NPC2-dependent cholesterol transport. Finally, we assessed the in vivo antiviral efficacies of three compounds known to inhibit NPC1 function or NPC1-glycoprotein binding in vitro. Two compounds reduced viral titers in vivo and provided a modest, albeit not statistically significant, degree of protection. Taken together, our results show that NPC1 is critical for replication and pathogenesis in animals and is a bona fide target for development of antifilovirus therapeutics. Additionally, our findings with Npc1(+/-) mice raise the possibility that individuals heterozygous for NPC1 may have a survival advantage in the face of EBOV infection. IMPORTANCE:Researchers have been searching for an essential filovirus receptor for decades, and numerous candidate receptors have been proposed. However, none of the proposed candidate receptors has proven essential in all in vitro scenarios, nor have they proven essential when evaluated using animal models. In this report, we provide the first example of a knockout mouse that is completely refractory to EBOV infection, replication, and disease. The findings detailed here provide the first critical in vivo data illustrating the absolute requirement of NPC1 for filovirus infection in mice. Our work establishes NPC1 as a legitimate target for the development of anti-EBOV therapeutics. However, the limited success of available NPC1 inhibitors to protect mice from EBOV challenge highlights the need for new molecules or approaches to target NPC1 in vivo.

Dataset Information

Flotillins play an essential role in Niemann-Pick C1-like 1-mediated cholesterol uptake.

Publications

Flotillins play an essential role in Niemann-Pick C1-like 1-mediated cholesterol uptake.

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