title: Collaboration of AMPK and PKC to induce phosphorylation of Ser413 on PIP5K1B resulting in decreased kinase activity and reduced PtdIns(4,5)P2 synthesis in response to oxidative stress and energy restriction.
abstract: The spatial and temporal regulation of the second messenger PtdIns(4,5)P2 has been shown to be crucial for regulating numerous processes in the cytoplasm and in the nucleus. Three isoforms of PIP5K1 (phosphatidylinositol 4-phosphate 5-kinase), A, B and C, are responsible for the regulation of the major pools of cellular PtdIns(4,5)P2. PIP5K1B is negatively regulated in response to oxidative stress although it remains unclear which pathways regulate its activity. In the present study, we have investigated the regulation of PIP5K1B by protein phosphorylation. Using MS analysis, we identified 12 phosphorylation sites on PIP5K1B. We developed a phospho-specific antibody against Ser413 and showed that its phosphorylation was increased in response to treatment of cells with phorbol ester, H2O2 or energy restriction. Using inhibitors, we define a stress-dependent pathway that requires the activity of the cellular energy sensor AMPK (AMP-activated protein kinase) and PKC (protein kinase C) to regulate Ser413 phosphorylation. Furthermore, we demonstrate that PKC can directly phosphorylate Ser413 in vitro. Mutation of Ser413 to aspartate to mimic serine phosphorylation decreased both PIP5K1B activity in vitro and PtdIns(4,5)P2 synthesis in vivo. Our studies show that collaboration between AMPK and PKC dictates the extent of Ser413 phosphorylation on PIP5K1B and regulates PtdIns(4,5)P2 synthesis.
authors.affiliation: *Inositide laboratory, The Paterson Institute for Cancer Research, Wilmslow Road, Manchester M20 4BX, U.K.
authors.forename: Iman
authors.initials: I
authors.lastname: van den Bout
authors.forename: David R
authors.initials: DR
authors.lastname: Jones
authors.forename: Zahid H
authors.initials: ZH
authors.lastname: Shah
authors.forename: Jonathan R
authors.initials: JR
authors.lastname: Halstead
authors.forename: Willem-Jan
authors.initials: WJ
authors.lastname: Keune
authors.forename: Shabaz
authors.initials: S
authors.lastname: Mohammed
authors.forename: Clive S
authors.initials: CS
authors.lastname: D'Santos
authors.forename: Nullin
authors.initials: N
authors.lastname: Divecha
mesh_terms: D055372:AMP-Activated Protein Kinases / Q000378:metabolism*; D004734:Energy Metabolism; D057809:HEK293 Cells; D006367:HeLa Cells; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism; D009154:Mutation; D018384:Oxidative Stress; D019269:Phosphatidylinositol 4,5-Diphosphate / Q000378:metabolism; D010766:Phosphorylation; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000378:metabolism; D012694:Serine / Q000235:genetics* / Q000378:metabolism
chemical_list: D019269:Phosphatidylinositol 4,5-Diphosphate; D012694:Serine; D006861:Hydrogen Peroxide; D017853:Phosphotransferases (Alcohol Group Acceptor); C020077:1-phosphatidylinositol-4-phosphate 5-kinase; D055372:AMP-Activated Protein Kinases