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THREE DECADES OF QUESTIONS ON LIPID RAFTS Whitney Stutts 1.

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Presentation on theme: "THREE DECADES OF QUESTIONS ON LIPID RAFTS Whitney Stutts 1."— Presentation transcript:

1 THREE DECADES OF QUESTIONS ON LIPID RAFTS Whitney Stutts 1

2 Outline  What are lipid rafts?  Why do they form?  What methods are used to study lipid rafts?  What effects do they have on eukaryotes?  Why all the controversy? 2

3 What are lipid rafts?  “Lipid rafts are small (10-200nm), heterogeneous, highly dynamic, sterol- and sphingolipid-enriched domains that compartmentalize cellular processes.” ~ 2006 Keystone Symposium 3

4 What are lipid rafts?  Cholesterol and sphingolipid-enriched membrane microdomains or platforms  Cholesterol levels double  Sphingomyelin levels elevated by 50%  Concentrate and segregate proteins within the plane of the bilayer  More ordered and tightly packed than surrounding bilayer  Float freely in L c bilayer 4

5 Video  Lipid Rafts ml 5

6 Two types of lipid rafts  Caveolae: small, flask- shaped invaginations of the plasma membrane enriched in caveolin  Planar lipid rafts: found in neurons and enriched in flotillin  Caleolin and flotillin recruit signaling proteins  Signaling can be promoted or dampened 6

7 Raft Proteins  “True resident proteins”  GPI-anchored proteins-prion protein (PrPc)  Caveolin  Flotillin  Signaling proteins  G-protein, non-receptor tyrosine kinases  Cytoskeletal/Adhesion proteins  actin, myosin, vinculin, cofilin, cadherin, ezrin 7

8 Why do they form? 8

9 Why do they form? Cholesterol  Cholesterol is the dynamic “glue” that holds the raft together  Saturation  Hydroxyl H-bonding with amide  Up to 25% of cholesterol is found in the brain…CNS?  When removed, most proteins dissociate from rafts 9

10 Misconception  Rafts contain only phospholipids with fully saturated acyl chains….FALSE!  Most glycerophospholipids in membrane rafts contain at least one monounsaturated acyl chain  Sphygomyelin- usually saturated chains but when unsaturated the DB is located at C15 10

11 Why do rafts form?  Driving force- line tension: energy required to create a boundary between the raft and the surrounding membrane  Raft thickness Hydrophobic mismatch  contrast in thickness line tension  Results in larger, more circular rafts which reduce line tension and energetic cost of the raft interface length 11

12 What methods are used to study lipid rafts?  DRM Isolation - lipid rafts are insoluble in cold non-ionic detergents (Triton X-100)  Electon microscopy - determines location of raft components and can detect clustering of proteins  FRET – used to determine whether two raft components are spatially close  FRAP - probe for the association of proteins to lipid rafts and study diffusion of proteins in lipid rafts  Manipulation of cholesterol  Sequestration  Depletion or removal  Inhibition of cholesterol synthesis 12

13 What effects do rafts have on eukaryotes?  Organizing centers-assembly of signaling molecules  Signaling can be promoted or dampened  Effects membrane fluidity  Acyl chain fluidity  Lateral diffusion  Involved in trafficking of membrane proteins  Regulation of neurotransmission and receptor trafficking  Neurotrophin receptors embedded in rafts 13

14 Hijacking Viruses  HIV virus  Budding may occur from lipid rafts  Influenza virus  Raft-associated glycoproteins in envelope 14

15 Disorders & Diseases  Mood disorders  Therapeutic efficacy of antidepressants  Alzheimer’s disease  Platforms for production of amyloid- β (neurotoxic protein)  Prion disorder  Normal prion protein (PrPc) is converted to abnormal proteins (PrPsc) in lipid rafts (GPI anchor required) 15

16 Why all the controversy?  Problems with biomembranes  Lipid rafts are too small to be resolved by light microscopy  Difficult to study lipid rafts in intact cells  Not in thermodynamic equilibrium 16

17 Why all the controversy?  Problems with synthetic membranes  Lower concentration of proteins  Difficult to model membrane-cytoskeletal interactions  Lack natural lipid asymmetry  Studied under equilibrium conditions 17

18 More Questions  What are the effects of membrane protein levels?  What is the physiological function of lipid rafts?  What effect does flux of membrane lipids have on raft formation?  What effect do diet and drugs have on lipid rafts?  What effect do proteins located at raft boundaries have on lipid rafts? 18

19 Works Cited  Allen, John A. "Lipid raft microdomains and neurotransmitter signaling." Nature 8 (2007):  Benarroch, Eduardo E. "Lipid rafts, protein scaffolds, and neurologic disease." Neurology 69 (2007):  Hamasaki, Dr. Toshikazu. "Tutorial 2, Plasma Membrane." UCLA. 22 Feb  Jacobson, Ken. "Lipid rafts: at a crossroad between cell biology and physics." Nature Cell Biology 9 (2007):  Jacques Fantini, Nicolas Garmy, Radhia Mahfoud and Nouara Yahi (2002) Lipid rafts: structure, function and role in HIV, Alzheimer’s and prion diseases. Exp. Rev. Mol. Med. 20 December,  Korade, Zeljka. "Lipid rafts, cholesterol, and the brain." Neuropharmacology 55 (2008):  Luckey, Mary. Membrane Structural Biology : With Biochemical and Biophysical Foundations. New York: Cambridge UP,  Pike, Linda J. "The Challenge of Lipid Rafts." Journal of Lipid Research Oct (2008):  Simons, Kai, and Ehehalt, R. "Cholesterol, lipid rafts, and disease." The Journal of Clinical Invesigation 110 (2002):  Simons, Kai. "Lipid Rafts and Signal Transduction." Nature Reviews 1 (2000):  Simons, Kai. "Model Systems, Lipid Rafts, and Cell Membranes." Annu. Rev. Biophys. Biomol. Struct. 33 (2004):  Video: Viel, A., Lue R.A., “Inner life of the cell.” The president and Fellows of Harvard College (2007) 19


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