Eggeling, C. et al. Direct observation of the nanoscale dynamics of membrane lipids in a living cell. Nature 457, 1159-1162

Department of Nanobiophotonics, Max Planck Institute for Biophysical Chemistry, Am Fassberg 11, 37077 Göttingen, Germany.
Nature (Impact Factor: 41.46). 12/2008; 457(7233):1159-62. DOI: 10.1038/nature07596
Source: PubMed


Cholesterol-mediated lipid interactions are thought to have a functional role in many membrane-associated processes such as signalling events. Although several experiments indicate their existence, lipid nanodomains ('rafts') remain controversial owing to the lack of suitable detection techniques in living cells. The controversy is reflected in their putative size of 5-200 nm, spanning the range between the extent of a protein complex and the resolution limit of optical microscopy. Here we demonstrate the ability of stimulated emission depletion (STED) far-field fluorescence nanoscopy to detect single diffusing (lipid) molecules in nanosized areas in the plasma membrane of living cells. Tuning of the probed area to spot sizes approximately 70-fold below the diffraction barrier reveals that unlike phosphoglycerolipids, sphingolipids and glycosylphosphatidylinositol-anchored proteins are transiently ( approximately 10-20 ms) trapped in cholesterol-mediated molecular complexes dwelling within <20-nm diameter areas. The non-invasive optical recording of molecular time traces and fluctuation data in tunable nanoscale domains is a powerful new approach to study the dynamics of biomolecules in living cells.

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    • "Interestingly, syntaxin-1 molecules are only transiently associated with their nanoclusters and rapidly exchange with freely diffusing syntaxin-1 molecules . Live cell STED imaging also revealed the transient, nanoscale organization of sphingolipids and glycosylphosphatidylinositol (GPI)anchored proteins within a 20 nm cholesterol rich membrane region [53]. These studies support the notion that the PM organization is controlled by dynamic molecular interactions of associated and dissociated states. "
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    • "Technical progress facilitates today the not so long ago impossible analysis of the subtle changes in lipid composition and of the topographical distribution of individual lipid species in cellular compartments. Probes have been developed to label lipid molecules such as new generation fluorescent tags (Eggeling et al., 2009) or modified toxins with specific lipid binding abilities such as the theta-toxin or lysenin, which bind cholesterol or sphingomyelin, respectively (Abe et al., 2012). These probes together with advanced microscopy techniques that achieve sub-diffraction optical resolution (i.e., near-field scanning optical microscopy (NSOM), photoactivated localization microscopy (PALM) stochastic optical reconstruction microscopy (STORM) or stimulated depletion (STED) fluorescent microscopy) allow the direct observation of the nanoscale dynamics of membrane lipids in a living cell (Eggeling et al., 2009; van Zanten et al., 2010; Castro et al., 2013). "
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