title: Microdomain Formation Controls Spatiotemporal Dynamics of Cell-Surface Glycoproteins.
abstract: The effect of galectin-mediated microdomain formation on the spatiotemporal dynamics of glycosylated membrane proteins in human microvascular endothelial cells (HMEC-1) was studied qualitatively and quantitatively by high-resolution fluorescence microscopy and artificially mimicked by metabolic glycoprotein engineering. Two types of membrane proteins, sialic acid-bearing proteins (SABPs) and mucin-type proteins (MTPs), were investigated. For visualization they were metabolically labeled with azido sugars and then coupled to a cyclooctyne-conjugated fluorescent dye by click chemistry. Both spatial (diffusion) and temporal (residence time) dynamics of SABPs and MTPs on the membrane were investigated after treatment with exogenous galectin-1 or -3. Strong effects of galectin-mediated lattice formation were observed for MTPs (decreased spatial mobility), but not for SABPs. Lattice formation also strongly decreased the turnover of MTPs (increased residence time on the cell membrane). The effects of galectin-mediated crosslinking was accurately mimicked by streptavidin-mediated crosslinking of biotin-tagged glycoproteins and verified by single-molecule tracking. This technique allows the induction of crosslinking of membrane proteins under precisely controlled conditions, thereby influencing membrane residence time and the spatial dynamics of glycans on the cell membrane in a controlled way.
authors: Department of Physical Chemistry, Ludwig Maximilian University Munich, Butenandtstrasse 11, 81377, Munich, Germany.
authors: Leonhard
authors: L
authors: Möckl
authors: Institute for Experimental Immunology and Hepatology, University Medical Center Hamburg, Martinistrasse 52, 20246, Hamburg, Germany.
authors: Andrea K
authors: AK
authors: Horst
authors: Otto Diels Institute of Organic Chemistry, Christiana Albertina University of Kiel, Otto-Hahn-Platz 3-4, 24098, Kiel, Germany.
authors: Katharina
authors: K
authors: Kolbe
authors: Otto Diels Institute of Organic Chemistry, Christiana Albertina University of Kiel, Otto-Hahn-Platz 3-4, 24098, Kiel, Germany.
authors: Thisbe K
authors: TK
authors: Lindhorst
authors: Department of Physical Chemistry, Ludwig Maximilian University Munich, Butenandtstrasse 11, 81377, Munich, Germany. christoph.braeuchle@cup.lmu.de.
authors: Christoph
authors: C
authors: Bräuchle
mesh_terms: D002460:Cell Line; D004058:Diffusion; D042783:Endothelial Cells / Q000166:cytology / Q000378:metabolism*; D037161:Galectins / Q000378:metabolism*; D006801:Humans; D008562:Membrane Glycoproteins / Q000032:analysis / Q000378:metabolism*; D021962:Membrane Microdomains / Q000378:metabolism* / Q000648:ultrastructure; D009077:Mucins / Q000032:analysis / Q000378:metabolism; D012794:Sialic Acids / Q000032:analysis / Q000378:metabolism
chemical_list: D037161:Galectins; D008562:Membrane Glycoproteins; D009077:Mucins; D012794:Sialic Acids
keywords: galectin lattice; glycosylation; lattice mimicking; membrane proteins; single-molecule tracking