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End date: 2019 × Start date: 2010 × DDC: 570 × Type: Text × Publisher: Berlin : Nature Publishing × WGL Institute: IPHT ×

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    2D polarization imaging as a low-cost fluorescence method to detect α-synuclein aggregation ex vivo in models of Parkinson’s disease
    (Berlin : Nature Publishing, 2018) Camacho, Rafael; Täuber, Daniela; Hansen, Christian; Shi, Juanzi; Bousset, Luc; Melki, Ronald; Li, Jia-Yi; Scheblykin, Ivan G.
    A hallmark of Parkinson’s disease is the formation of large protein-rich aggregates in neurons, where α-synuclein is the most abundant protein. A standard approach to visualize aggregation is to fluorescently label the proteins of interest. Then, highly fluorescent regions are assumed to contain aggregated proteins. However, fluorescence brightness alone cannot discriminate micrometer-sized regions with high expression of non-aggregated proteins from regions where the proteins are aggregated on the molecular scale. Here, we demonstrate that 2-dimensional polarization imaging can discriminate between preformed non-aggregated and aggregated forms of α-synuclein, and detect increased aggregation in brain tissues of transgenic mice. This imaging method assesses homo-FRET between labels by measuring fluorescence polarization in excitation and emission simultaneously, which translates into higher contrast than fluorescence anisotropy imaging. Exploring earlier aggregation states of α-synuclein using such technically simple imaging method could lead to crucial improvements in our understanding of α-synuclein-mediated pathology in Parkinson’s Disease.
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    Mechanical properties of plasma membrane vesicles correlate with lipid order, viscosity and cell density
    (Berlin : Nature Publishing, 2019) Steinkühler, Jan; Sezgin, Erdinc; Urbančič, Iztok; Eggeling, Christian; Dimova, Rumiana
    Regulation of plasma membrane curvature and composition governs essential cellular processes. The material property of bending rigidity describes the energetic cost of membrane deformations and depends on the plasma membrane molecular composition. Because of compositional fluctuations and active processes, it is challenging to measure it in intact cells. Here, we study the plasma membrane using giant plasma membrane vesicles (GPMVs), which largely preserve the plasma membrane lipidome and proteome. We show that the bending rigidity of plasma membranes under varied conditions is correlated to readout from environment-sensitive dyes, which are indicative of membrane order and microviscosity. This correlation holds across different cell lines, upon cholesterol depletion or enrichment of the plasma membrane, and variations in cell density. Thus, polarity- and viscosity-sensitive probes represent a promising indicator of membrane mechanical properties. Additionally, our results allow for identifying synthetic membranes with a few well defined lipids as optimal plasma membrane mimetics.