title: Homotypic fusion of immature secretory granules during maturation requires syntaxin 6.
abstract: Homotypic fusion of immature secretory granules (ISGs) gives rise to mature secretory granules (MSGs), the storage compartment in endocrine and neuroendocrine cells for hormones and neuropeptides. With the use of a cell-free fusion assay, we investigated which soluble N-ethylmaleimide-sensitive fusion protein attachment receptor (SNARE) molecules are involved in the homotypic fusion of ISGs. Interestingly, the SNARE molecules mediating the exocytosis of MSGs in neuroendocrine cells, syntaxin 1, SNAP-25, and VAMP2, were not involved in homotypic ISG fusion. Instead, we have identified syntaxin 6 as a component of the core machinery responsible for homotypic ISG fusion. Subcellular fractionation studies and indirect immunofluorescence microscopy show that syntaxin 6 is sorted away during the maturation of ISGs to MSGs. Although, syntaxin 6 on ISG membranes is associated with SNAP-25 and SNAP-29/GS32, we could not find evidence that these target (t)-SNARE molecules are involved in homotypic ISG fusion. Nor could we find any involvement for the vesicle (v)-SNARE VAMP4, which is known to be associated with syntaxin 6. Importantly, we have shown that homotypic fusion requires the function of syntaxin 6 on both donor as well as acceptor membranes, which suggests that t-t-SNARE interactions, either direct or indirect, may be required during fusion of ISG membranes.
authors.affiliation: Secretory Pathway Laboratory, Imperial Cancer Research Fund, London WC2A 3PX, UK.
authors.forename: F
authors.initials: F
authors.lastname: Wendler
authors.forename: L
authors.initials: L
authors.lastname: Page
authors.forename: S
authors.initials: S
authors.lastname: Urbé
authors.forename: S A
authors.initials: SA
authors.lastname: Tooze
mesh_terms: D000818:Animals; D000954:Antigens, Surface / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism; D002474:Cell-Free System; D002850:Chromatography, Gel; D004305:Dose-Response Relationship, Drug; D004703:Endocrine System / Q000378:metabolism; D019084:Fluorescent Antibody Technique, Indirect; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism / Q000502:physiology*; D008856:Microscopy, Fluorescence; D009419:Nerve Tissue Proteins / Q000737:chemistry / Q000378:metabolism; D016716:PC12 Cells; D011233:Precipitin Tests; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D050765:Qa-SNARE Proteins; D050683:R-SNARE Proteins; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D050600:SNARE Proteins; D022142:Secretory Vesicles / Q000378:metabolism*; D013347:Subcellular Fractions; D050825:Synaptosomal-Associated Protein 25; D050827:Syntaxin 1; D033921:Vesicular Transport Proteins
chemical_list: D000954:Antigens, Surface; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D050765:Qa-SNARE Proteins; D050683:R-SNARE Proteins; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D050600:SNARE Proteins; C494210:Snap25 protein, rat; C494213:Stx1a protein, rat; D050825:Synaptosomal-Associated Protein 25; D050827:Syntaxin 1; D033921:Vesicular Transport Proteins