title: Dynamic Lipid-dependent Modulation of Protein Topology by Post-translational Phosphorylation.
abstract: Membrane protein topology and folding are governed by structural principles and topogenic signals that are recognized and decoded by the protein insertion and translocation machineries at the time of initial membrane insertion and folding. We previously demonstrated that the lipid environment is also a determinant of initial protein topology, which is dynamically responsive to post-assembly changes in membrane lipid composition. However, the effect on protein topology of post-assembly phosphorylation of amino acids localized within initially cytoplasmically oriented extramembrane domains has never been investigated. Here, we show in a controlled in vitro system that phosphorylation of a membrane protein can trigger a change in topological arrangement. The rate of change occurred on a scale of seconds, comparable with the rates observed upon changes in the protein lipid environment. The rate and extent of topological rearrangement were dependent on the charges of extramembrane domains and the lipid bilayer surface. Using model membranes mimicking the lipid compositions of eukaryotic organelles, we determined that anionic lipids, cholesterol, sphingomyelin, and membrane fluidity play critical roles in these processes. Our results demonstrate how post-translational modifications may influence membrane protein topology in a lipid-dependent manner, both along the organelle trafficking pathway and at their final destination. The results provide further evidence that membrane protein topology is dynamic, integrating for the first time the effect of changes in lipid composition and regulators of cellular processes. The discovery of a new topology regulatory mechanism opens additional avenues for understanding unexplored structure-function relationships and the development of optimized topology prediction tools.
authors.affiliation: From the Department of Biochemistry and Molecular Biology and Center for Membrane Biology, University of Texas McGovern Medical School, Houston, Texas 77030. Electronic address: Heidi.Vitrac@uth.tmc.edu.
authors.forename: Heidi
authors.initials: H
authors.lastname: Vitrac
authors.affiliation: From the Department of Biochemistry and Molecular Biology and Center for Membrane Biology, University of Texas McGovern Medical School, Houston, Texas 77030.
authors.forename: David M
authors.initials: DM
authors.lastname: MacLean
authors.affiliation: the Department of Internal Medicine, Division of Cardiology, University of Texas McGovern Medical School, Houston, Texas 77030.
authors.forename: Anja
authors.initials: A
authors.lastname: Karlstaedt
authors.affiliation: the Department of Internal Medicine, Division of Cardiology, University of Texas McGovern Medical School, Houston, Texas 77030.
authors.forename: Heinrich
authors.initials: H
authors.lastname: Taegtmeyer
authors.affiliation: From the Department of Biochemistry and Molecular Biology and Center for Membrane Biology, University of Texas McGovern Medical School, Houston, Texas 77030.
authors.forename: Vasanthi
authors.initials: V
authors.lastname: Jayaraman
authors.affiliation: From the Department of Biochemistry and Molecular Biology and Center for Membrane Biology, University of Texas McGovern Medical School, Houston, Texas 77030.
authors.forename: Mikhail
authors.initials: M
authors.lastname: Bogdanov
authors.affiliation: From the Department of Biochemistry and Molecular Biology and Center for Membrane Biology, University of Texas McGovern Medical School, Houston, Texas 77030. Electronic address: william.dowhan@uth.tmc.edu.
authors.forename: William
authors.initials: W
authors.lastname: Dowhan
mesh_terms: D002462:Cell Membrane / Q000737:chemistry* / Q000378:metabolism; D008051:Lipid Bilayers / Q000737:chemistry* / Q000378:metabolism; D008563:Membrane Lipids / Q000737:chemistry* / Q000378:metabolism; D008565:Membrane Proteins / Q000737:chemistry* / Q000378:metabolism; D010766:Phosphorylation; D000072417:Protein Domains; D011499:Protein Processing, Post-Translational
chemical_list: D008051:Lipid Bilayers; D008563:Membrane Lipids; D008565:Membrane Proteins
keywords: fluorescence resonance energy transfer (FRET); glycerophospholipid; lipid-protein interaction; membrane protein; phosphorylation; post-translational modification (PTM)