title: Lipoprotein complex of equine lysozyme with oleic acid (ELOA) interactions with the plasma membrane of live cells.
abstract: Recent evidence supports the idea that early aggregates, protein, and lipoprotein oligomers but not large aggregates like fibrils that are formed at late stages of the aggregation process are responsible for cytotoxicity. Oligomers can interact with the cellular plasma membrane affecting its structure and/or dynamics or may be taken up by the cells. In either case, disparate cascades of molecular interactions are activated in the attempt to counteract the disturbance induced by the oligomers. If unsuccessful, cell death follows. Here, we study the molecular and cellular mechanisms underlying PC12 cell death caused by ELOA oligomers. ELOA, a lipoprotein complex formed by equine lysozyme (EL) and oleic acid (OA), induces cell death in all tested cell lines, but the actual mechanism of its action is not known. We have used methods with single-molecule sensitivity, fluorescence correlation spectroscopy (FCS), fluorescence cross-correlation spectroscopy (FCCS), and confocal laser scanning microscopy (CLSM) imaging by avalanche photodiodes (APD), so-called APD imaging, to study ELOA interactions with the plasma membrane in live PC12 cells. We detected ELOA accumulation in the cell surroundings, observed ELOA interactions with the plasma membrane, and local changes in plasma membrane lipid dynamics in the vicinity of ELOA complexes. These interactions resulted in plasma membrane rupture, followed by rapid influx and distribution of ELOA inside the already dead cell. In order to probe the ELOA-plasma membrane interaction sites at the molecular and atomic levels, the ELOA complexes were further studied by photochemically induced dynamic nuclear polarization (photo-CIDNP) spectroscopy, nuclear magnetic resonance (NMR) and atomic force microscopy (AFM). We observed a novel mechanism of oligomer toxicity-cell death induced by continuous disturbance of the plasma membrane, eventually causing permanent plasma membrane damage and identified the sites in ELOA that are potentially involved in the interactions with the plasma membrane.
authors.affiliation: Department of Clinical Neuroscience, Karolinska Institutet, 17176 Stockholm, Sweden. vladana.vukojevic@ki.se
authors.forename: Vladana
authors.initials: V
authors.lastname: Vukojević
authors.forename: Alice M
authors.initials: AM
authors.lastname: Bowen
authors.forename: Kristina
authors.initials: K
authors.lastname: Wilhelm
authors.forename: Yu
authors.initials: Y
authors.lastname: Ming
authors.forename: Zhang
authors.initials: Z
authors.lastname: Ce
authors.forename: Jürgen
authors.initials: J
authors.lastname: Schleucher
authors.forename: P J
authors.initials: PJ
authors.lastname: Hore
authors.forename: Lars
authors.initials: L
authors.lastname: Terenius
authors.forename: Ludmilla A
authors.initials: LA
authors.lastname: Morozova-Roche
mesh_terms: D000818:Animals; D001665:Binding Sites; D002462:Cell Membrane / Q000378:metabolism*; D002470:Cell Survival; D003116:Color; D005456:Fluorescent Dyes / Q000378:metabolism; D006736:Horses; D008074:Lipoproteins / Q000378:metabolism*; D009113:Muramidase / Q000378:metabolism*; D019301:Oleic Acid / Q000378:metabolism*; D016716:PC12 Cells; D011485:Protein Binding; D051381:Rats; D013050:Spectrometry, Fluorescence
chemical_list: D005456:Fluorescent Dyes; D008074:Lipoproteins; D019301:Oleic Acid; D009113:Muramidase