Cytoskeletal Control of CD36 Diffusion Promotes Its Receptor and Signaling Function

Department of Systems Biology, Harvard Medical School, Boston, MA 02115, USA.
Cell (Impact Factor: 32.24). 08/2011; 146(4):593-606. DOI: 10.1016/j.cell.2011.06.049
Source: PubMed


The mechanisms that govern receptor coalescence into functional clusters--often a critical step in their stimulation by ligand--are poorly understood. We used single-molecule tracking to investigate the dynamics of CD36, a clustering-responsive receptor that mediates oxidized LDL uptake by macrophages. We found that CD36 motion in the membrane was spatially structured by the cortical cytoskeleton. A subpopulation of receptors diffused within linear confinement regions whose unique geometry simultaneously facilitated freedom of movement along one axis while increasing the effective receptor density. Co-confinement within troughs enhanced the probability of collisions between unligated receptors and promoted their clustering. Cytoskeleton perturbations that inhibited diffusion in linear confinement regions reduced receptor clustering in the absence of ligand and, following ligand addition, suppressed CD36-mediated signaling and internalization. These observations demonstrate a role for the cytoskeleton in controlling signal transduction by structuring receptor diffusion within membrane regions that increase their collision frequency.

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Available from: Nicolas Touret, Jan 29, 2015
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    • "This has led to the " picket-fence " picture of membrane organization wherein certain membrane components are corralled by the stable actin mesh (Kusumi et al., 2012). Recent studies have also shown the cortical actin and microtubule based cytoskeleton can spatially organize the diffusion of oxidized-LDL receptor CD36 and regulate its signaling ability (Jaqaman et al., 2011). Fluorescence correlation spectroscopy (FCS) measurements in cells have also suggested that mobilities of sphingolipids, ganglioside GM1 and GPI-anchored proteins shows a signature of transient trapping arising from their engagement with nanoscale lipid domains (Lenne et al., 2006) and/or the underlying cortical actin meshwork (Mueller et al., 2011). "
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    ABSTRACT: Molecular diffusion at the surface of living cells is thought to be predominantly driven by thermal kicks. However, there is growing evidence that certain cell surface molecules are driven by the fluctuating dynamics of cortical cytoskeleton. Using fluorescence correlation spectroscopy (FCS) we measure the diffusion coefficient of a variety of cell-surface molecules over a temperature range 24-37°C. Predictably, exogenously incorporated fluorescent lipids with short acyl chains exhibit the expected increase of diffusion coefficient over this temperature range. In contrast, we find that GPI-anchored proteins exhibit temperature independent diffusion over this range, and revert to temperature-dependent diffusion on cell membrane blebs, in cells depleted of cholesterol, and upon acute perturbation of actin dynamics and myosin activity. A model transmembrane protein with a cytosolic actin-binding domain also exhibits the temperature independent behavior, thereby directly implicating the role of cortical actin. We show that diffusion of GPI-anchored proteins also becomes temperature-dependent when the filamentous dynamic actin nucleator, formin, is inhibited. However, changes in cortical actin mesh size or perturbation of branched actin nucleator Arp2/3 do not affect this behavior. Thus, the cell surface diffusion of GPI-anchored proteins and transmembrane protein that associate with actin, are driven by the active fluctuations of dynamic cortical actin filaments in addition to thermal fluctuations, consistent with expectations from an "active actin-membrane composite" cell surface.
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    • "Second, the plasma membrane is attached to an actin cortex, whose role in influencing local membrane composition is poorly understood. Finally, the organization and dynamics of a variety of plasma membrane molecules such as membrane proteins (Gowrishankar et al., 2012; Jaqaman et al., 2011), lipid-anchored proteins (Goswami et al., 2008; Prior et al., 2003; Sharma et al., 2004), and glycolipids (Fujita et al., 2007) into nanometer sized clusters cannot be derived from equilibrium-based mechanisms. "
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    ABSTRACT: Understanding how functional lipid domains in live cell membranes are generated has posed a challenge. Here, we show that transbilayer interactions are necessary for the generation of cholesterol-dependent nanoclusters of GPI-anchored proteins mediated by membrane-adjacent dynamic actin filaments. We find that long saturated acyl-chains are required for forming GPI-anchor nanoclusters. Simultaneously, at the inner leaflet, long acyl-chain-containing phosphatidylserine (PS) is necessary for transbilayer coupling. All-atom molecular dynamics simulations of asymmetric multicomponent-membrane bilayers in a mixed phase provide evidence that immobilization of long saturated acyl-chain lipids at either leaflet stabilizes cholesterol-dependent transbilayer interactions forming local domains with characteristics similar to a liquid-ordered (lo) phase. This is verified by experiments wherein immobilization of long acyl-chain lipids at one leaflet effects transbilayer interactions of corresponding lipids at the opposite leaflet. This suggests a general mechanism for the generation and stabilization of nanoscale cholesterol-dependent and actin-mediated lipid clusters in live cell membranes. Copyright © 2015 Elsevier Inc. All rights reserved.
    No preview · Article · Apr 2015 · Cell
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    • "In all other cases, p values were calculated using Student's t test. Mathematical and statistical analyses for SPT and subdiffraction imaging experiments were performed using custom scripts written in Matlab software (MathWorks, Natick, MA; Jaqaman et al., 2008, 2011). Data are presented as mean ± SEM unless otherwise stated. "
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