title: Scanning STED-FCS reveals spatiotemporal heterogeneity of lipid interaction in the plasma membrane of living cells.
abstract: The interaction of lipids and proteins plays an important role in plasma membrane bioactivity, and much can be learned from their diffusion characteristics. Here we present the combination of super-resolution STED microscopy with scanning fluorescence correlation spectroscopy (scanning STED-FCS, sSTED-FCS) to characterize the spatial and temporal heterogeneity of lipid interactions. sSTED-FCS reveals transient molecular interaction hotspots for a fluorescent sphingolipid analogue. The interaction sites are smaller than 80 nm in diameter and lipids are transiently trapped for several milliseconds in these areas. In comparison, newly developed fluorescent phospholipid and cholesterol analogues with improved phase-partitioning properties show more homogenous diffusion, independent of the preference for liquid-ordered or disordered membrane environments. Our results do not support the presence of nanodomains based on lipid-phase separation in the basal membrane of our cultured nonstimulated cells, and show that alternative interactions are responsible for the strong local trapping of our sphingolipid analogue.
authors.affiliation: Department of NanoBiophotonics, Max-Planck-Institute for Biophysical Chemistry, Am Fassberg 11, Goettingen 37077, Germany.
authors.forename: Alf
authors.initials: A
authors.lastname: Honigmann
authors.affiliation: Department of NanoBiophotonics, Max-Planck-Institute for Biophysical Chemistry, Am Fassberg 11, Goettingen 37077, Germany.
authors.forename: Veronika
authors.initials: V
authors.lastname: Mueller
authors.affiliation: Department of NanoBiophotonics, Max-Planck-Institute for Biophysical Chemistry, Am Fassberg 11, Goettingen 37077, Germany.
authors.forename: Haisen
authors.initials: H
authors.lastname: Ta
authors.affiliation: Department of NanoBiophotonics, Max-Planck-Institute for Biophysical Chemistry, Am Fassberg 11, Goettingen 37077, Germany.
authors.forename: Andreas
authors.initials: A
authors.lastname: Schoenle
authors.affiliation: MRC Human Immunology Unit, Weatherall Institute of Molecular Medicine, University of Oxford, Headley Way, Oxford OX3 9DS, UK.
authors.forename: Erdinc
authors.initials: E
authors.lastname: Sezgin
authors.affiliation: Department of NanoBiophotonics, Max-Planck-Institute for Biophysical Chemistry, Am Fassberg 11, Goettingen 37077, Germany.
authors.forename: Stefan W
authors.initials: SW
authors.lastname: Hell
authors.affiliation: 1] Department of NanoBiophotonics, Max-Planck-Institute for Biophysical Chemistry, Am Fassberg 11, Goettingen 37077, Germany [2] MRC Human Immunology Unit, Weatherall Institute of Molecular Medicine, University of Oxford, Headley Way, Oxford OX3 9DS, UK.
authors.forename: Christian
authors.initials: C
authors.lastname: Eggeling
mesh_terms: D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism*; D002784:Cholesterol / Q000378:metabolism; D008563:Membrane Lipids / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism*; D008853:Microscopy; D010743:Phospholipids / Q000378:metabolism; D048328:Potoroidae; D013050:Spectrometry, Fluorescence; D013107:Sphingolipids / Q000378:metabolism
chemical_list: D008563:Membrane Lipids; D008565:Membrane Proteins; D010743:Phospholipids; D013107:Sphingolipids; D002784:Cholesterol