1. The palmitoylation state of PMP22 modulates epithelial cell morphology and migration.
- Author
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Zoltewicz SJ, Lee S, Chittoor VG, Freeland SM, Rangaraju S, Zacharias DA, and Notterpek L
- Subjects
- Animals, Bacterial Proteins genetics, Caveolins metabolism, Cell Movement drug effects, Cells, Cultured, Cicatrix metabolism, Cicatrix pathology, Contactin 1 metabolism, Cysteine genetics, Cysteine metabolism, Dogs, Dose-Response Relationship, Drug, Epithelial Cells drug effects, Fatty Acids, Unsaturated pharmacology, Green Fluorescent Proteins metabolism, Humans, Lectins metabolism, Lipoylation drug effects, Lipoylation genetics, Luminescent Proteins genetics, Madin Darby Canine Kidney Cells, Mutation genetics, Myelin Proteins genetics, Radioimmunoprecipitation Assay, Rats, Schwann Cells cytology, Schwann Cells drug effects, Signal Transduction drug effects, Signal Transduction genetics, Transfection, Wounds and Injuries pathology, rac GTP-Binding Proteins metabolism, Cell Movement genetics, Cell Size, Epithelial Cells cytology, Epithelial Cells physiology, Lipoylation physiology, Myelin Proteins metabolism
- 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.
- Published
- 2012
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