title: The palmitoylation state of PMP22 modulates epithelial cell morphology and migration.
abstract: PMP22 (peripheral myelin protein 22), also known as GAS 3 (growth-arrest-specific protein 3), is a disease-linked tetraspan glycoprotein of peripheral nerve myelin and constituent of intercellular junctions in epithelia. To date, our knowledge of the post-translational modification of PMP22 is limited. Using the CSS-Palm 2.0 software we predicted that C85 (cysteine 85), a highly conserved amino acid located between the second and third transmembrane domains, is a potential site for palmitoylation. To test this, we mutated C85S (C85 to serine) and established stable cells lines expressing the WT (wild-type) or the C85S-PMP22. In Schwann and MDCK (Madin-Darby canine kidney) cells mutating C85 blocked the palmitoylation of PMP22, which we monitored using 17-ODYA (17-octadecynoic acid). While palmitoylation was not necessary for processing the newly synthesized PMP22 through the secretory pathway, overexpression of C85S-PMP22 led to pronounced cell spreading and uneven monolayer thinning. To further investigate the functional significance of palmitoylated PMP22, we evaluated MDCK cell migration in a wound-healing assay. While WT-PMP22 expressing cells were resistant to migration, C85S cells displayed lamellipodial protrusions and migrated at a similar rate to vector control. These findings indicate that palmitoylation of PMP22 at C85 is critical for the role of the protein in modulating epithelial cell shape and motility.
authors.affiliation: Department of Neuroscience, College of Medicine, McKnight Brain Institute, University of Florida, Gainesville, FL 32610, USA.
authors.forename: Susie J
authors.initials: SJ
authors.lastname: Zoltewicz
authors.forename: Sooyeon
authors.initials: S
authors.lastname: Lee
authors.forename: Vinita G
authors.initials: VG
authors.lastname: Chittoor
authors.forename: Steven M
authors.initials: SM
authors.lastname: Freeland
authors.forename: Sunitha
authors.initials: S
authors.lastname: Rangaraju
authors.forename: David A
authors.initials: DA
authors.lastname: Zacharias
authors.forename: Lucia
authors.initials: L
authors.lastname: Notterpek
mesh_terms: D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics; D022461:Caveolins / Q000378:metabolism; D002465:Cell Movement / Q000187:drug effects / Q000235:genetics*; D048429:Cell Size; D002478:Cells, Cultured; D002921:Cicatrix / Q000378:metabolism / Q000473:pathology; D058967:Contactin 1 / Q000378:metabolism; D003545:Cysteine / Q000235:genetics / Q000378:metabolism; D004285:Dogs; D004305:Dose-Response Relationship, Drug; D004847:Epithelial Cells / Q000166:cytology* / Q000187:drug effects / Q000502:physiology*; D005231:Fatty Acids, Unsaturated / Q000494:pharmacology; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006801:Humans; D037102:Lectins / Q000378:metabolism; D054878:Lipoylation / Q000187:drug effects / Q000235:genetics / Q000502:physiology*; D008164:Luminescent Proteins / Q000235:genetics; D061985:Madin Darby Canine Kidney Cells; D009154:Mutation / Q000235:genetics; D009185:Myelin Proteins / Q000235:genetics / Q000378:metabolism*; D015531:Radioimmunoprecipitation Assay; D051381:Rats; D012583:Schwann Cells / Q000166:cytology / Q000187:drug effects; D015398:Signal Transduction / Q000187:drug effects / Q000235:genetics; D014162:Transfection; D014947:Wounds and Injuries / Q000473:pathology; D020744:rac GTP-Binding Proteins / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D022461:Caveolins; D058967:Contactin 1; D005231:Fatty Acids, Unsaturated; D037102:Lectins; D008164:Luminescent Proteins; D009185:Myelin Proteins; C073687:PMP22 protein, human; C054562:yellow fluorescent protein, Bacteria; D049452:Green Fluorescent Proteins; C047008:17-octadecynoic acid; D020744:rac GTP-Binding Proteins; D003545:Cysteine