title: Interaction of the Spo20 membrane-sensor motif with phosphatidic acid and other anionic lipids, and influence of the membrane environment.
abstract: The yeast protein Spo20 contains a regulatory amphipathic motif that has been suggested to recognize phosphatidic acid, a lipid involved in signal transduction, lipid metabolism and membrane fusion. We have investigated the interaction of the Spo20 amphipathic motif with lipid membranes using a bioprobe strategy that consists in appending this motif to the end of a long coiled-coil, which can be coupled to a GFP reporter for visualization in cells. The resulting construct is amenable to in vitro and in vivo experiments and allows unbiased comparison between amphipathic helices of different chemistry. In vitro, the Spo20 bioprobe responded to small variations in the amount of phosphatidic acid. However, this response was not specific. The membrane binding of the probe depended on the presence of phosphatidylethanolamine and also integrated the contribution of other anionic lipids, including phosphatidylserine and phosphatidyl-inositol-(4,5)bisphosphate. Inverting the sequence of the Spo20 motif neither affected the ability of the probe to interact with anionic liposomes nor did it modify its cellular localization, making a stereo-specific mode of phosphatidic acid recognition unlikely. Nevertheless, the lipid binding properties and the cellular localization of the Spo20 alpha-helix differed markedly from that of another amphipathic motif, Amphipathic Lipid Packing Sensor (ALPS), suggesting that even in the absence of stereo specific interactions, amphipathic helices can act as subcellular membrane targeting determinants in a cellular context.
authors.affiliation: Institut de Pharmacologie Moléculaire et Cellulaire, Université de Nice Sophia Antipolis et CNRS, Valbonne, France.
authors.forename: Habib
authors.initials: H
authors.lastname: Horchani
authors.affiliation: Institut de Pharmacologie Moléculaire et Cellulaire, Université de Nice Sophia Antipolis et CNRS, Valbonne, France.
authors.forename: Maud
authors.initials: M
authors.lastname: de Saint-Jean
authors.affiliation: Institut de Pharmacologie Moléculaire et Cellulaire, Université de Nice Sophia Antipolis et CNRS, Valbonne, France.
authors.forename: Hélène
authors.initials: H
authors.lastname: Barelli
authors.affiliation: Institut de Pharmacologie Moléculaire et Cellulaire, Université de Nice Sophia Antipolis et CNRS, Valbonne, France.
authors.forename: Bruno
authors.initials: B
authors.lastname: Antonny
mesh_terms: D020816:Amino Acid Motifs; D002460:Cell Line; D004847:Epithelial Cells / Q000166:cytology / Q000378:metabolism; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D008024:Ligands; D008081:Liposomes / Q000737:chemistry / Q000378:metabolism; D015335:Molecular Probes; D008969:Molecular Sequence Data; D010712:Phosphatidic Acids / Q000378:metabolism*; D010714:Phosphatidylethanolamines / Q000737:chemistry / Q000378:metabolism; D018129:Phosphatidylinositol Phosphates / Q000737:chemistry / Q000378:metabolism; D010718:Phosphatidylserines / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D050766:Qb-SNARE Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D050767:Qc-SNARE Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction
chemical_list: D008024:Ligands; D008081:Liposomes; D015335:Molecular Probes; D010712:Phosphatidic Acids; D010714:Phosphatidylethanolamines; D018129:Phosphatidylinositol Phosphates; D010718:Phosphatidylserines; D050766:Qb-SNARE Proteins; D050767:Qc-SNARE Proteins; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; C497084:Spo20 protein, S cerevisiae; D049452:Green Fluorescent Proteins; C483858:phosphatidylethanolamine