title: Effect of receptor dimerization on membrane lipid raft structure continuously quantified on single cells by camera based fluorescence correlation spectroscopy.
abstract: Membrane bound cell signaling is modulated by the membrane ultra-structure, which itself may be affected by signaling. However, measuring the interaction of membrane proteins with membrane structures in intact cells in real-time poses considerable challenges. In this paper we present a non-destructive fluorescence method that quantifies these interactions in single cells, and is able to monitor the same cell continuously to observe small changes. This approach combines total internal fluorescence microscopy with fluorescence correlation spectroscopy to measure the protein's diffusion and molecular concentration in different sized areas simultaneously. It correctly differentiates proteins interacting with membrane fences from proteins interacting with cholesterol-stabilized domains, or lipid rafts. This method detects small perturbations of the membrane ultra-structure or of a protein's tendency to dimerize. Through continuous monitoring of single cells, we demonstrate how dimerization of GPI-anchored proteins increases their association with the structural domains. Using a dual-color approach we study the effect of dimerization of one GPI-anchored protein on another type of GPI-anchored protein expressed in the same cell. Scans over the cell surface reveal a correlation between cholesterol stabilized domains and membrane cytoskeleton.
authors: Department of Physics, University at Buffalo the State University of New York, Buffalo, New York, United States of America.
authors: Heng
authors: H
authors: Huang
authors: Department of Physics, University at Buffalo the State University of New York, Buffalo, New York, United States of America.
authors: M Fethullah
authors: MF
authors: Simsek
authors: Department of Physics, University at Buffalo the State University of New York, Buffalo, New York, United States of America.
authors: Weixiang
authors: W
authors: Jin
authors: Department of Physics, University at Buffalo the State University of New York, Buffalo, New York, United States of America; Department of Biophysics and Physiology, University at Buffalo the State University of New York, Buffalo, New York, United States of America.
authors: Arnd
authors: A
authors: Pralle
mesh_terms: D000185:Actinin / Q000378:metabolism; D002784:Cholesterol / Q000378:metabolism; D004058:Diffusion; D058973:Folate Receptors, GPI-Anchored / Q000737:chemistry / Q000378:metabolism*; D049452:Green Fluorescent Proteins / Q000378:metabolism; D021962:Membrane Microdomains / Q000737:chemistry*; D008565:Membrane Proteins / Q000378:metabolism; D010781:Photography / Q000295:instrumentation*; D055503:Protein Multimerization; D017434:Protein Structure, Tertiary; D059010:Single-Cell Analysis / Q000295:instrumentation*; D013050:Spectrometry, Fluorescence / Q000295:instrumentation*
chemical_list: D058973:Folate Receptors, GPI-Anchored; D008565:Membrane Proteins; D000185:Actinin; D049452:Green Fluorescent Proteins; D002784:Cholesterol