title: The sphingoid long chain base phytosphingosine activates AGC-type protein kinases in Saccharomyces cerevisiae including Ypk1, Ypk2, and Sch9.
abstract: The Pkh1 protein kinase of Saccharomyces cerevisiae, a homolog of the mammalian 3-phosphoinositide-dependent kinase (PDK1), regulates downstream AGC-type protein kinases including Ypk1/2 and Pkc1, which control cell wall integrity, growth, and other processes. Phytosphingosine (PHS), a sphingoid long chain base, is hypothesized to be a lipid activator of Pkh1 and thereby controls the activity of Ypk1/2. Here we present biochemical evidence supporting this hypothesis, and in addition we demonstrate that PHS also stimulates autophosphorylation and activation of Ypk1/2. Greatest stimulation of Ypk1/2 phosphorylation and activity are achieved by inclusion of both PHS and Pkh1 in an in vitro kinase reaction. We also demonstrate for the first time that Pkh1 phosphorylates the Sch9 protein kinase in vitro and that such phosphorylation is stimulated by PHS. This is the first biochemical demonstration of Sch9 activators, and the results further support roles for long chain bases in heat stress resistance in addition to implying roles in chronological aging and cell size determination, since Sch9 functions in these processes. Thus, our data support a model in which PHS, rather than simply being an upstream activator of Pkh1, also activates kinases that are downstream targets of Pkh1 including Ypk1/2 and Sch9.
authors.affiliation: Department of Molecular and Cellular Biochemistry, University of Kentucky College of Medicine, Lexington, 40536, USA.
authors.forename: Ke
authors.initials: K
authors.lastname: Liu
authors.forename: Xiping
authors.initials: X
authors.lastname: Zhang
authors.forename: Robert L
authors.initials: RL
authors.lastname: Lester
authors.forename: Robert C
authors.initials: RC
authors.lastname: Dickson
mesh_terms: D064413:3-Phosphoinositide-Dependent Protein Kinases; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation; D015966:Gene Expression Regulation, Fungal; D005816:Genetic Complementation Test; D038362:Glycogen Synthase Kinase 3; D006639:Histidine / Q000737:chemistry; D006358:Hot Temperature; D047468:Immunoprecipitation; D007700:Kinetics; D050356:Lipid Metabolism; D008954:Models, Biological; D009154:Mutation; D010641:Phenotype; D010766:Phosphorylation; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D011494:Protein Kinases / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases; D017434:Protein Structure, Tertiary; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000502:physiology*; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism* / Q000502:physiology; D013110:Sphingosine / Q000031:analogs & derivatives* / Q000737:chemistry*; D013997:Time Factors
chemical_list: D029701:Saccharomyces cerevisiae Proteins; D006639:Histidine; D011494:Protein Kinases; C071569:SCH9 protein kinase; C060388:YPK2 protein, S cerevisiae; D011505:Protein-Tyrosine Kinases; D064413:3-Phosphoinositide-Dependent Protein Kinases; C118385:PKH1 protein, S cerevisiae; D017346:Protein Serine-Threonine Kinases; D038362:Glycogen Synthase Kinase 3; C068124:MCK1 protein, S cerevisiae; C012491:phytosphingosine; D013110:Sphingosine