title: Synip arrests soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE)-dependent membrane fusion as a selective target membrane SNARE-binding inhibitor.
abstract: The vesicle fusion reaction in regulated exocytosis requires the concerted action of soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) core fusion engine and a group of SNARE-binding regulatory factors. The regulatory mechanisms of vesicle fusion remain poorly understood in most exocytic pathways. Here, we reconstituted the SNARE-dependent vesicle fusion reaction of GLUT4 exocytosis in vitro using purified components. Using this defined fusion system, we discovered that the regulatory factor synip binds to GLUT4 exocytic SNAREs and inhibits the docking, lipid mixing, and content mixing of the fusion reaction. Synip arrests fusion by binding the target membrane SNARE (t-SNARE) complex and preventing the initiation of ternary SNARE complex assembly. Although synip also interacts with the syntaxin-4 monomer, it does not inhibit the pairing of syntaxin-4 with SNAP-23. Interestingly, synip selectively arrests the fusion reactions reconstituted with its cognate SNAREs, suggesting that the defined system recapitulates the biological functions of the vesicle fusion proteins. We further showed that the inhibitory function of synip is dominant over the stimulatory activity of Sec1/Munc18 proteins. Importantly, the inhibitory function of synip is distinct from how other fusion inhibitors arrest SNARE-dependent membrane fusion and therefore likely represents a novel regulatory mechanism of vesicle fusion.
authors.affiliation: Department of Molecular, Cellular, and Developmental Biology, University of Colorado, Boulder, Colorado 80309, USA.
authors.forename: Haijia
authors.initials: H
authors.lastname: Yu
authors.forename: Shailendra S
authors.initials: SS
authors.lastname: Rathore
authors.forename: Jingshi
authors.initials: J
authors.lastname: Shen
mesh_terms: D000818:Animals; D002462:Cell Membrane / Q000378:metabolism*; D005089:Exocytosis; D051275:Glucose Transporter Type 4 / Q000378:metabolism; D007313:Insecta; D008051:Lipid Bilayers / Q000737:chemistry; D008081:Liposomes / Q000378:metabolism; D008561:Membrane Fusion / Q000502:physiology*; D051379:Mice; D051938:Munc18 Proteins / Q000378:metabolism; D011485:Protein Binding; D050765:Qa-SNARE Proteins / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D050600:SNARE Proteins / Q000378:metabolism*; D033921:Vesicular Transport Proteins / Q000378:metabolism*
chemical_list: D051275:Glucose Transporter Type 4; D008051:Lipid Bilayers; D008081:Liposomes; D051938:Munc18 Proteins; D050765:Qa-SNARE Proteins; D011994:Recombinant Proteins; D050600:SNARE Proteins; C495693:Slc2a4 protein, mouse; C120000:Stxbp4 protein, mouse; D033921:Vesicular Transport Proteins
keywords: Membrane Fusion; Membrane Proteins; Membrane Reconstitution; Membrane Trafficking; Vesicles