negative regulation of syntaxin4snap-23vamp2-mediated membrane fusion by munc18c in vitro负调节syntaxin4snap-23vamp2-mediated munc18c体外膜融合.pdfVIP

Negative Regulation of Syntaxin4/SNAP-23/VAMP2- Mediated Membrane Fusion by Munc18c In Vitro 1. 1. 2 2 1 1 Fiona M. Brandie , Veronica Aran , Avani Verma , James A. McNew , Nia J. Bryant , Gwyn W. Gould * 1 Henry Wellcome Laboratory of Cell Biology, Division of Biochemistry and Molecular Biology, Institute of Biomedical and Life Sciences, University of Glasgow, Glasgow, United States of America, 2 Department of Biochemistry and Cell Biology, Rice University, Houston, Texas, United States of America Abstract Background: Translocation of the facilitative glucose transporter GLUT4 from an intracellular store to the plasma membrane is responsible for the increased rate of glucose transport into fat and muscle cells in response to insulin. This represents a specialised form of regulated membrane trafficking. Intracellular membrane traffic is subject to multiple levels of regulation by conserved families of proteins in all eukaryotic cells. Notably, all intracellular fusion events require SNARE proteins and Sec1p/Munc18 family members. Fusion of GLUT4-containing vesicles with the plasma membrane of insulin-sensitive cells involves the SM protein Munc18c, and is regulated by the formation of syntaxin 4/SNAP23/VAMP2 SNARE complexes. Methodology/Principal Findings: Here we have used biochemical approaches to characterise the interaction(s) of Munc18c with its cognate SNARE proteins and to examine the role of Munc18c in regulating liposome fusion catalysed by syntaxin 4/ SNAP23/VAMP2 SNARE complex formation. We demonstrate that Munc18c makes contacts with both t- and v-SNARE proteins of this complex, and directly inhibits bilayer fusion mediated by the syntaxin 4/SNAP23/VAMP2 SNARE complex. Conclusion/Significance