title: Sng1 associates with Nce102 to regulate the yeast Pkh-Ypk signalling module in response to sphingolipid status.
abstract: All cells are delimited by biological membranes, which are consequently a primary target of stress-induced damage. Cold alters membrane functionality by decreasing lipid fluidity and the activity of membrane proteins. In Saccharomyces cerevisiae, evidence links sphingolipid homeostasis and membrane phospholipid asymmetry to the activity of the Ypk1/2 proteins, the yeast orthologous of the mammalian SGK1-3 kinases. Their regulation is mediated by different protein kinases, including the PDK1 orthologous Pkh1/2p, and requires the function of protein effectors, among them Nce102p, a component of the sphingolipid sensor machinery. Nevertheless, the mechanisms and the actors involved in Pkh/Ypk regulation remain poorly defined. Here, we demonstrate that Sng1, a transmembrane protein, is an effector of the Pkh/Ypk module and identify the phospholipid asymmetry as key for yeast cold adaptation. Overexpression of SNG1 impairs phospholipid flipping, reduces reactive oxygen species (ROS) and improves, in a Pkh-dependent manner, yeast growth in myriocin-treated cells, suggesting that excess Sng1p stimulates the Pkh/Ypk signalling. Furthermore, we link these effects to the association of Sng1p with Nce102p. Indeed, we found that Sng1p interacts with Nce102p both physically and genetically. Moreover, mutant nce102∆ sng1∆ cells show features of impaired Pkh/Ypk signalling, including increased ROS accumulation, reduced life span and defects in Pkh/Ypk-controlled regulatory pathways. Finally, myriocin-induced hyperphosphorylation of Ypk1p and Orm2p, which controls sphingolipid homeostasis, does not occur in nce102∆ sng1∆ cells. Hence, both Nce102p and Sng1p participate in a regulatory circuit that controls the activity of the Pkh/Ypk module and their function is required in response to sphingolipid status.
authors.affiliation: Department of Biotechnology, Instituto de Agroquímica y Tecnología de los Alimentos, Consejo Superior de Investigaciones Científicas, Avda. Agustín Escardino, 7. 46980, Paterna, Valencia, Spain.
authors.forename: Sara
authors.initials: S
authors.lastname: García-Marqués
authors.affiliation: Department of Biotechnology, Instituto de Agroquímica y Tecnología de los Alimentos, Consejo Superior de Investigaciones Científicas, Avda. Agustín Escardino, 7. 46980, Paterna, Valencia, Spain.
authors.forename: Francisca
authors.initials: F
authors.lastname: Randez-Gil
authors.affiliation: UMR Procédés Alimentaires et Microbiologiques (PAM), AgroSup Dijon/Université de Bourgogne 1, Esplanade Erasme, 21000, Dijon, France.
authors.forename: Sebastien
authors.initials: S
authors.lastname: Dupont
authors.affiliation: Department of Biotechnology, Instituto de Agroquímica y Tecnología de los Alimentos, Consejo Superior de Investigaciones Científicas, Avda. Agustín Escardino, 7. 46980, Paterna, Valencia, Spain.
authors.forename: Elena
authors.initials: E
authors.lastname: Garre
authors.affiliation: Department of Biotechnology, Instituto de Agroquímica y Tecnología de los Alimentos, Consejo Superior de Investigaciones Científicas, Avda. Agustín Escardino, 7. 46980, Paterna, Valencia, Spain. Electronic address: prieto@iata.csic.es.
authors.forename: Jose A
authors.initials: JA
authors.lastname: Prieto
mesh_terms: D064413:3-Phosphoinositide-Dependent Protein Kinases / Q000235:genetics / Q000378:metabolism*; D001430:Bacteriocins / Q000494:pharmacology; D003080:Cold Temperature; D005229:Fatty Acids, Monounsaturated / Q000494:pharmacology; D005454:Fluorescence Polarization; D038362:Glycogen Synthase Kinase 3 / Q000235:genetics / Q000378:metabolism*; D006706:Homeostasis / Q000187:drug effects; D015151:Immunoblotting; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D018613:Microscopy, Confocal; D008954:Models, Biological; D009154:Mutation; D010455:Peptides / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D011485:Protein Binding; D017382:Reactive Oxygen Species; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction / Q000235:genetics; D013107:Sphingolipids / Q000037:antagonists & inhibitors / Q000378:metabolism*
chemical_list: D001430:Bacteriocins; D005229:Fatty Acids, Monounsaturated; D008565:Membrane Proteins; C525535:NCE102 protein, S cerevisiae; D010455:Peptides; D017382:Reactive Oxygen Species; C093629:SNG1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D013107:Sphingolipids; C040522:duramycin; D064413:3-Phosphoinositide-Dependent Protein Kinases; C118385:PKH1 protein, S cerevisiae; D038362:Glycogen Synthase Kinase 3; C068124:MCK1 protein, S cerevisiae; C001996:thermozymocidin
keywords: Cold stress; Membrane properties; Myriocin; Orm2; Phospholipid flipping; Signalling; Yeast