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PUBLISHED ONLINE: 26 DECEMBER 2016 | http://dx.doi.org/10.1038/nchembio.2268
Web End =DOI: 10.1038/NCHEMBIO.2268
Orthogonal lipid sensors identify transbilayer asymmetry of plasma membrane cholesterol
Shu-Lin Liu1,6, Ren Sheng1,6, Jae Hun Jung2, Li Wang1, Ewa Stec1, Matthew J OConnor1, Seohyoen Song1, Rama Kamesh Bikkavilli3, Robert A Winn3, Daesung Lee1, Kwanghee Baek4, Kazumitsu Ueda5,
Irena Levitan3, Kwang-Pyo Kim2 & Wonhwa Cho1,4
Controlled distribution of lipids across various cell membranes is crucial for cell homeostasis and regulation. We developed an imaging method that allows simultaneous in situ quantification of cholesterol in two leaflets of the plasma membrane (PM) using tunable orthogonal cholesterol sensors. Our imaging revealed marked transbilayer asymmetry of PM cholesterol (TAPMC) in various mammalian cells, with the concentration in the inner leaflet (IPM) being ~12-fold lower than that in the outer leaflet (OPM). The asymmetry was maintained by active transport of cholesterol from IPM to OPM and its chemical retention at OPM. Furthermore, the increase in the IPM cholesterol level was triggered in a stimulus-specific manner, allowing cholesterol to serve as a signaling lipid. We found excellent correlation between the IPM cholesterol level and cellular Wnt signaling activity, suggesting that TAPMC and stimulus-induced PM cholesterol redistribution are crucial for tight regulation of cellular processes under physiological conditions.
2017 Nature America, Inc., part of Springer Nature. All rights reserved.
Cell membranes contain a wide variety of lipid molecules with different structural and functional properties. Many lipids are asymmetrically distributed across various cell mem
branes, most notably in the PM of mammalian cells13. Transbilayer asymmetry of membrane lipids is maintained by complex mechanisms involving lipid transporters and is crucial for cell homeostasis and regulation2. However, conventional methods for determining transbilayer distribution of lipids fail to provide spatiotempo-rally resolved quantitative information because they depend on lipid extraction, use of labeled lipids or indirect estimation13. The
unavailability of direct real-time quantification methods makes it difficult to accurately determine the degree of dynamic transbilayer asymmetry of lipids and to elucidate when and how changes in transbilayer distribution of lipids modulate cellular processes.
Cholesterol is a major lipid in the PM of mammalian cells4 and has diverse structural and functional roles5,6. Cellular unesterified cholesterol, which is either derived from low-density lipoprotein-receptor-mediated endocytosis or synthesized de novo in the endoplasmic reticulum57, is primarily (up to...