title: The stress-activated phosphatidylinositol 3-phosphate 5-kinase Fab1p is essential for vacuole function in S. cerevisiae.
abstract: Polyphosphoinositides have many roles in cell signalling and vesicle trafficking [1-3]. Phosphatidylinositol 3,5-bisphosphate (PI(3,5)P2), a recently discovered PIP2 isomer, is ubiquitous in eukaryotic cells and rapidly accumulates in hyperosmotically stressed yeast. PI(3,5)P2 is synthesised from PI(3)P in both yeast and mammalian cells [4,5]. A search of the Saccharomyces cerevisiae genome database identified FAB1, a gene encoding a PIP kinase homologue and potential PI(3)P 5-kinase. Fab1p shows PI(3)P 5-kinase activity both in vivo and in vitro. A yeast strain in which FAB1 had been deleted was unable to synthesise PI(3,5)P2, either in the presence or absence of osmotic shock. A loss of PI(3,5)P2 was observed also in a temperature-sensitive FAB1 strain at the non-permissive temperature. A recombinant glutathione-S-transferase (GST)-Fab1p fusion protein was shown to have selective PI(3)P 5-kinase activity in vitro. Thus, we have demonstrated that Fab1p is a PI(3)P-specific 5-kinase and represents a third class of PIP kinase activity, which we have termed type III. Deletion of the FAB1 gene produces a loss of vacuolar morphology [6]; it is therefore concluded that PI(3,5)P2, the lipid product of Fab1p, is required for normal vacuolar function.
authors.affiliation: Protein Phosphorylation Laboratory, Imperial Cancer Research Fund, London, UK.
authors.forename: F T
authors.initials: FT
authors.lastname: Cooke
authors.forename: S K
authors.initials: SK
authors.lastname: Dove
authors.forename: R K
authors.initials: RK
authors.lastname: McEwen
authors.forename: G
authors.initials: G
authors.lastname: Painter
authors.forename: A B
authors.initials: AB
authors.lastname: Holmes
authors.forename: M N
authors.initials: MN
authors.lastname: Hall
authors.forename: R H
authors.initials: RH
authors.lastname: Michell
authors.forename: P J
authors.initials: PJ
authors.lastname: Parker
mesh_terms: D016296:Mutagenesis; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism*; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D012441:Saccharomyces cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D013379:Substrate Specificity; D014617:Vacuoles
chemical_list: D019869:Phosphatidylinositol 3-Kinases; D018129:Phosphatidylinositol Phosphates; D017853:Phosphotransferases (Alcohol Group Acceptor); D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; C106336:phosphatidylinositol 3,5-diphosphate; C055525:phosphatidylinositol 3-phosphate; C037178:phosphatidylinositol 4-phosphate; C113567:phosphatidylinositol 5-phosphate; C095504:FAB1 protein, S cerevisiae; C020077:1-phosphatidylinositol-4-phosphate 5-kinase