title: Ion regulation of homotypic vacuole fusion in Saccharomyces cerevisiae.
abstract: Biological membrane fusion employs divalent cations as protein cofactors or as signaling ligands. For example, Mg2+ is a cofactor for the N-ethylmaleimide-sensitive factor (NSF) ATPase, and the Ca2+ signal from neuronal membrane depolarization is required for synaptotagmin activation. Divalent cations also regulate liposome fusion, but the role of such ion interactions with lipid bilayers in Rab- and soluble NSF attachment protein receptor (SNARE)-dependent biological membrane fusion is less clear. Yeast vacuole fusion requires Mg2+ for Sec18p ATPase activity, and vacuole docking triggers an efflux of luminal Ca2+. We now report distinct reaction conditions where divalent or monovalent ions interchangeably regulate Rab- and SNARE-dependent vacuole fusion. In reactions with 5 mm Mg2+, other free divalent ions are not needed. Reactions containing low Mg2+ concentrations are strongly inhibited by the rapid Ca2+ chelator BAPTA. However, addition of the soluble SNARE Vam7p relieves BAPTA inhibition as effectively as Ca2+ or Mg2+, suggesting that Ca2+ does not perform a unique signaling function. When the need for Mg2+, ATP, and Sec18p for fusion is bypassed through the addition of Vam7p, vacuole fusion does not require any appreciable free divalent cations and can even be stimulated by their chelators. The similarity of these findings to those with liposomes, and the higher ion specificity of the regulation of proteins, suggests a working model in which ion interactions with bilayer lipids permit Rab- and SNARE-dependent membrane fusion.
authors.affiliation: Department of Biochemistry, Dartmouth Medical School, Hanover, New Hampshire 03755-3844, USA.
authors.forename: Vincent J
authors.initials: VJ
authors.lastname: Starai
authors.forename: Naomi
authors.initials: N
authors.lastname: Thorngren
authors.forename: Rutilio A
authors.initials: RA
authors.lastname: Fratti
authors.forename: William
authors.initials: W
authors.lastname: Wickner
mesh_terms: D000251:Adenosine Triphosphatases / Q000737:chemistry; D002118:Calcium / Q000737:chemistry / Q000378:metabolism; D002412:Cations; D002614:Chelating Agents / Q000494:pharmacology; D002712:Chlorides / Q000737:chemistry; D004305:Dose-Response Relationship, Drug; D004492:Edetic Acid / Q000494:pharmacology; D004533:Egtazic Acid / Q000031:analogs & derivatives* / Q000494:pharmacology; D007477:Ions; D008024:Ligands; D008081:Liposomes / Q000737:chemistry; D008274:Magnesium / Q000737:chemistry; D008561:Membrane Fusion; D008565:Membrane Proteins / Q000737:chemistry; D052066:N-Ethylmaleimide-Sensitive Proteins; D009419:Nerve Tissue Proteins / Q000737:chemistry; D009474:Neurons / Q000378:metabolism; D011189:Potassium Chloride / Q000737:chemistry; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000378:metabolism*; D012492:Salts / Q000494:pharmacology; D050825:Synaptosomal-Associated Protein 25; D014617:Vacuoles / Q000737:chemistry*; D033921:Vesicular Transport Proteins / Q000737:chemistry / Q000378:metabolism*; D015032:Zinc / Q000737:chemistry; D017967:Zinc Compounds / Q000737:chemistry; D020691:rab GTP-Binding Proteins / Q000737:chemistry
chemical_list: D002412:Cations; D002614:Chelating Agents; D002712:Chlorides; D007477:Ions; D008024:Ligands; D008081:Liposomes; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D029701:Saccharomyces cerevisiae Proteins; D012492:Salts; D050825:Synaptosomal-Associated Protein 25; D033921:Vesicular Transport Proteins; D017967:Zinc Compounds; D004533:Egtazic Acid; D011189:Potassium Chloride; C016837:zinc chloride; D004492:Edetic Acid; D000251:Adenosine Triphosphatases; C064404:SEC18 protein, S cerevisiae; D052066:N-Ethylmaleimide-Sensitive Proteins; D020691:rab GTP-Binding Proteins; D008274:Magnesium; D015032:Zinc; C025603:1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid; D002118:Calcium