title: Breaking the chain at the membrane: paraoxonase 2 counteracts lipid peroxidation at the plasma membrane.
abstract: Lipid peroxidation through electrophilic molecules of extracellular origin is involved in the pathogenesis of many inflammatory conditions. To counteract free radical actions at the plasma membrane, cells host a variety of antioxidative enzymes. Here we analyzed localization, membrane topology, and trafficking of PON2 a member of the paraoxonase family of 3 enzymatically active proteins (PON1-3) found to have antiatherogenic properties. Immunohistochemistry localized PON2 to the villous tip of human intestinal epithelial cells. Employing membrane preparations, surface biotinylation experiments, and mutational analyses in HEK 293T and HeLa cells, we demonstrate that PON2 is a type II transmembrane protein. A hydrophobic stretch in the N terminus was identified as single transmembrane domain of PON2. The enzymatically active domain faced the extracellular compartment, where it suppressed lipid peroxidation (P<0.05) and regulated the glucosylceramide content, as demonstrated by mass spectrometry (P<0.05). PON2 translocation to the plasma membrane was dependent on intracellular calcium responses and could be induced to >10-fold as compared to baseline (P=0.0001) by oxidative stress. Taken together, these data identify the paraoxonase protein PON2 as a type II transmembrane protein, which is dynamically translocated to the plasma membrane in response to oxidative stress to counteract lipid peroxidation.
authors.affiliation: 1Department II of Internal Medicine and Center for Molecular Medicine, University of Cologne, Kerpener Str. 62, 50937 Cologne, Germany. P.T.B., paul.brinkkoetter@uk-koeln.de.
authors.forename: Henning
authors.initials: H
authors.lastname: Hagmann
authors.forename: Alexander
authors.initials: A
authors.lastname: Kuczkowski
authors.forename: Michael
authors.initials: M
authors.lastname: Ruehl
authors.forename: Tobias
authors.initials: T
authors.lastname: Lamkemeyer
authors.forename: Susanne
authors.initials: S
authors.lastname: Brodesser
authors.forename: Sven
authors.initials: S
authors.lastname: Horke
authors.forename: Stuart
authors.initials: S
authors.lastname: Dryer
authors.forename: Bernhard
authors.initials: B
authors.lastname: Schermer
authors.forename: Thomas
authors.initials: T
authors.lastname: Benzing
authors.forename: Paul Thomas
authors.initials: PT
authors.lastname: Brinkkoetter
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D043303:Aryldialkylphosphatase / Q000235:genetics / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism*; D004847:Epithelial Cells / Q000201:enzymology; D005963:Glucosylceramides / Q000378:metabolism; D057809:HEK293 Cells; D006367:HeLa Cells; D006801:Humans; D015151:Immunoblotting; D007150:Immunohistochemistry; D007422:Intestines / Q000166:cytology / Q000201:enzymology; D015227:Lipid Peroxidation; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D018613:Microscopy, Confocal; D008969:Molecular Sequence Data; D018384:Oxidative Stress; D021381:Protein Transport; D034622:RNA Interference
chemical_list: D005963:Glucosylceramides; D008565:Membrane Proteins; D043303:Aryldialkylphosphatase; D002118:Calcium
keywords: arylesterase; atherosclerosis; inflammation; oxidative stress