title: Asymmetric phospholipid distribution drives in vitro reconstituted SNARE-dependent membrane fusion.
abstract: Insulin-stimulated glucose uptake requires the fusion of GLUT4 transporter-containing vesicles with the plasma membrane, a process that depends on the SNARE (soluble N-ethylmaleimide-sensitive fusion factor attachment receptor) proteins VAMP2 (vesicle-associated membrane protein 2) and syntaxin 4 (Stx4)/SNAP23 (soluble N-ethylmaleimide-sensitive fusion factor attachment protein 23). Efficient SNARE-dependent fusion has been shown in many settings in vivo to require the generation of both phosphatidylinositol-4,5-bisphosphate (PIP2) and phosphatidic acid (PA). Addition of PA to Stx4/SNAP23 vesicles markedly enhanced the fusion rate, whereas its addition to VAMP2 vesicles was inhibitory. In contrast, addition of PIP2 to Stx4/SNAP23 vesicles inhibited the fusion reaction, and its addition to VAMP2 vesicles was stimulatory. The optimal distribution of phospholipids was found to trigger the progression from the hemifused state to full fusion. These findings reveal an unanticipated dependence of SNARE complex-mediated fusion on asymmetrically distributed acidic phospholipids and provide mechanistic insights into the roles of phospholipase D and PIP kinases in the late stages of regulated exocytosis.
authors.affiliation: Department of Pharmacological Sciences and Center for Developmental Genetics, Stony Brook University, Stony Brook, NY 11794, USA.
authors.forename: Jérôme
authors.initials: J
authors.lastname: Vicogne
authors.forename: Daniel
authors.initials: D
authors.lastname: Vollenweider
authors.forename: Jeffery R
authors.initials: JR
authors.lastname: Smith
authors.forename: Ping
authors.initials: P
authors.lastname: Huang
authors.forename: Michael A
authors.initials: MA
authors.lastname: Frohman
authors.forename: Jeffrey E
authors.initials: JE
authors.lastname: Pessin
mesh_terms: D000818:Animals; D006801:Humans; D008561:Membrane Fusion / Q000502:physiology*; D008567:Membranes, Artificial; D051379:Mice; D019269:Phosphatidylinositol 4,5-Diphosphate / Q000378:metabolism; D010739:Phospholipase D / Q000378:metabolism; D010743:Phospholipids / Q000378:metabolism*; D050765:Qa-SNARE Proteins / Q000378:metabolism*; D050766:Qb-SNARE Proteins / Q000378:metabolism*; D050767:Qc-SNARE Proteins / Q000378:metabolism*; D050988:Vesicle-Associated Membrane Protein 2 / Q000378:metabolism*
chemical_list: D008567:Membranes, Artificial; D019269:Phosphatidylinositol 4,5-Diphosphate; D010743:Phospholipids; D050765:Qa-SNARE Proteins; D050766:Qb-SNARE Proteins; D050767:Qc-SNARE Proteins; C107017:Snap23 protein, mouse; C494807:VAMP2 protein, human; D050988:Vesicle-Associated Membrane Protein 2; D010739:Phospholipase D