title: Roles of phosphoinositides and of Spo14p (phospholipase D)-generated phosphatidic acid during yeast sporulation.
abstract: During yeast sporulation, internal membrane synthesis ensures that each haploid nucleus is packaged into a spore. Prospore membrane formation requires Spo14p, a phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P2]-stimulated phospholipase D (PLD), which hydrolyzes phosphatidylcholine (PtdCho) to phosphatidic acid (PtdOH) and choline. We found that both meiosis and spore formation also require the phosphatidylinositol (PtdIns)/PtdCho transport protein Sec14p. Specific ablation of the PtdIns transport activity of Sec14p was sufficient to impair spore formation but not meiosis. Overexpression of Pik1p, a PtdIns 4-kinase, suppressed the sec14-1 meiosis and spore formation defects; conversely, pik1-ts diploids failed to undergo meiosis and spore formation. The PtdIns(4)P 5-kinase, Mss4p, also is essential for spore formation. Use of phosphoinositide-specific GFP-PH domain reporters confirmed that PtdIns(4,5)P2 is enriched in prospore membranes. sec14, pik1, and mss4 mutants displayed decreased Spo14p PLD activity, whereas absence of Spo14p did not affect phosphoinositide levels in vivo, suggesting that formation of PtdIns(4,5)P2 is important for Spo14p activity. Spo14p-generated PtdOH appears to have an essential role in sporulation, because treatment of cells with 1-butanol, which supports Spo14p-catalyzed PtdCho breakdown but leads to production of Cho and Ptd-butanol, blocks spore formation at concentrations where the inert isomer, 2-butanol, has little effect. Thus, rather than a role for PtdOH in stimulating PtdIns(4,5)P2 formation, our findings indicate that during sporulation, Spo14p-mediated PtdOH production functions downstream of Sec14p-, Pik1p-, and Mss4p-dependent PtdIns(4,5)P2 synthesis.
authors.affiliation: Department of Cellular and Molecular Medicine, School of Medicine, and Howard Hughes Medical Institute, University of California, San Diego, La Jolla, California 92093-0668, USA.
authors.forename: Simon A
authors.initials: SA
authors.lastname: Rudge
authors.forename: Vicki A
authors.initials: VA
authors.lastname: Sciorra
authors.forename: Michelle
authors.initials: M
authors.lastname: Iwamoto
authors.forename: Chun
authors.initials: C
authors.lastname: Zhou
authors.forename: Thomas
authors.initials: T
authors.lastname: Strahl
authors.forename: Andrew J
authors.initials: AJ
authors.lastname: Morris
authors.forename: Jeremy
authors.initials: J
authors.lastname: Thorner
authors.forename: JoAnne
authors.initials: J
authors.lastname: Engebrecht
mesh_terms: D020001:1-Butanol / Q000494:pharmacology; D019870:1-Phosphatidylinositol 4-Kinase / Q000378:metabolism; D001692:Biological Transport; D000440:Butanols / Q000494:pharmacology; D002352:Carrier Proteins / Q000378:metabolism*; D003001:Cloning, Molecular; D020404:Glycerophospholipids / Q000378:metabolism; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins; D008540:Meiosis / Q000187:drug effects; D008565:Membrane Proteins / Q000378:metabolism*; D009154:Mutation; D010712:Phosphatidic Acids / Q000378:metabolism*; D019269:Phosphatidylinositol 4,5-Diphosphate; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism; D010716:Phosphatidylinositols / Q000378:metabolism; D010739:Phospholipase D / Q000378:metabolism*; D049294:Phospholipid Transfer Proteins; D010770:Phosphotransferases; D017853:Phosphotransferases (Alcohol Group Acceptor); D011506:Proteins; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism; D013172:Spores, Fungal / Q000378:metabolism*
chemical_list: D000440:Butanols; D002352:Carrier Proteins; D020404:Glycerophospholipids; D008164:Luminescent Proteins; D008565:Membrane Proteins; D010712:Phosphatidic Acids; D019269:Phosphatidylinositol 4,5-Diphosphate; D018129:Phosphatidylinositol Phosphates; D010716:Phosphatidylinositols; D049294:Phospholipid Transfer Proteins; D011506:Proteins; C483438:SEC14 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; C068564:phosphatidylbutanol; C043958:2-butanol; D049452:Green Fluorescent Proteins; D020001:1-Butanol; D010770:Phosphotransferases; D017853:Phosphotransferases (Alcohol Group Acceptor); D019870:1-Phosphatidylinositol 4-Kinase; C441760:PIK1 protein, S cerevisiae; C489795:MSS4 protein, S cerevisiae; D010739:Phospholipase D