title: Oxidized Phospholipids Inhibit the Formation of Cholesterol-Dependent Plasma Membrane Nanoplatforms.
abstract: We previously developed a single-molecule microscopy method termed TOCCSL (thinning out clusters while conserving stoichiometry of labeling), which allows for direct imaging of stable nanoscopic platforms with raft-like properties diffusing in the plasma membrane. As a consensus raft marker, we chose monomeric GFP linked via a glycosylphosphatidylinositol (GPI) anchor to the cell membrane (mGFP-GPI). With this probe, we previously observed cholesterol-dependent homo-association to nanoplatforms diffusing in the plasma membrane of live CHO cells. Here, we report the release of this homo-association upon addition of 1-palmitoyl-2-(5-oxovaleroyl)-sn-glycero-3-phosphocholine (POVPC) or 1-palmitoyl-2-glutaroyl-sn-glycero-3-phosphocholine, two oxidized phospholipids (oxPLs) that are typically present in oxidatively modified low-density lipoprotein. We found a dose-response relationship for mGFP-GPI nanoplatform disintegration upon addition of POVPC, correlating with the signal of the apoptosis marker Annexin V-Cy3. Similar concentrations of lysolipid showed no effect, indicating that the observed phenomena were not linked to properties of the lipid bilayer itself. Inhibition of acid sphingomyelinase by NB-19 before addition of POVPC completely abolished nanoplatform disintegration by oxPLs. In conclusion, we were able to determine how oxidized lipid species disrupt mGFP-GPI nanoplatforms in the plasma membrane. Our results favor an indirect mechanism involving acid sphingomyelinase activity rather than a direct interaction of oxPLs with nanoplatform constituents.
authors.affiliation: Institute of Applied Physics, TU Wien, Vienna, Austria. Electronic address: brameshuber@iap.tuwien.ac.at.
authors.forename: Mario
authors.initials: M
authors.lastname: Brameshuber
authors.affiliation: Institute of Applied Physics, TU Wien, Vienna, Austria.
authors.forename: Eva
authors.initials: E
authors.lastname: Sevcsik
authors.affiliation: Institute of Applied Physics, TU Wien, Vienna, Austria.
authors.forename: Benedikt K
authors.initials: BK
authors.lastname: Rossboth
authors.affiliation: Institute of Applied Physics, TU Wien, Vienna, Austria.
authors.forename: Christina
authors.initials: C
authors.lastname: Manner
authors.affiliation: Faculty of Medical and Life Sciences, Institute of Precision Medicine, Furtwangen University, Villingen-Schwenningen, Germany; Fraunhofer Institute IZI/EXIM, Furtwangen, Germany.
authors.forename: Hans-Peter
authors.initials: HP
authors.lastname: Deigner
authors.affiliation: Institute of Biochemistry, Biological Research Centre, Hungarian Academy of Sciences, Szeged, Hungary.
authors.forename: Begüm
authors.initials: B
authors.lastname: Peksel
authors.affiliation: Institute of Biochemistry, Biological Research Centre, Hungarian Academy of Sciences, Szeged, Hungary.
authors.forename: Mária
authors.initials: M
authors.lastname: Péter
authors.affiliation: Institute of Biochemistry, Biological Research Centre, Hungarian Academy of Sciences, Szeged, Hungary.
authors.forename: Zsolt
authors.initials: Z
authors.lastname: Török
authors.affiliation: Institute of Biochemistry, Graz University of Technology, Graz, Austria.
authors.forename: Albin
authors.initials: A
authors.lastname: Hermetter
authors.affiliation: Institute of Applied Physics, TU Wien, Vienna, Austria. Electronic address: schuetz@iap.tuwien.ac.at.
authors.forename: Gerhard J
authors.initials: GJ
authors.lastname: Schütz
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D016466:CHO Cells; D002462:Cell Membrane / Q000187:drug effects* / Q000378:metabolism*; D002784:Cholesterol / Q000378:metabolism*; D006224:Cricetinae; D003412:Cricetulus; D017261:Glycosylphosphatidylinositols / Q000378:metabolism; D006801:Humans; D008853:Microscopy; D036103:Nanotechnology; D010084:Oxidation-Reduction; D010742:Phospholipid Ethers / Q000494:pharmacology*
chemical_list: C400184:1-palmitoyl-2-(5-oxovaleroyl)-sn-glycero-3-phosphorylcholine; C400185:1-palmitoyl-2-glutaroyl-sn-glycero-3-phosphorylcholine; D017261:Glycosylphosphatidylinositols; D010742:Phospholipid Ethers; D002784:Cholesterol