title: SNAP25, but not syntaxin 1A, recycles via an ARF6-regulated pathway in neuroendocrine cells.
abstract: Soluble N-ethylmaleimide-sensitive fusion protein attachment protein receptor (SNARE) proteins mediate cellular membrane fusion events and provide a level of specificity to donor-acceptor membrane interactions. However, the trafficking pathways by which individual SNARE proteins are targeted to specific membrane compartments are not well understood. In neuroendocrine cells, synaptosome-associated protein of 25 kDa (SNAP25) is localized to the plasma membrane where it functions in regulated secretory vesicle exocytosis, but it is also found on intracellular membranes. We identified a dynamic recycling pathway for SNAP25 in PC12 cells through which plasma membrane SNAP25 recycles in approximately 3 h. Approximately 20% of the SNAP25 resides in a perinuclear recycling endosome-trans-Golgi network (TGN) compartment from which it recycles back to the plasma membrane. SNAP25 internalization occurs by constitutive, dynamin-independent endocytosis that is distinct from the dynamin-dependent endocytosis that retrieves secretory vesicle constituents after exocytosis. Endocytosis of SNAP25 is regulated by ADP-ribosylation factor (ARF)6 (through phosphatidylinositol bisphosphate synthesis) and is dependent upon F-actin. SNAP25 endosomes, which exclude the plasma membrane SNARE syntaxin 1A, merge with those derived from clathrin-dependent endocytosis containing endosomal syntaxin 13. Our results characterize a robust ARF6-dependent internalization mechanism that maintains an intracellular pool of SNAP25, which is compatible with possible intracellular roles for SNAP25 in neuroendocrine cells.
authors.affiliation: Department of Biochemistry, University of Wisconsin, Madison, WI 53706, USA.
authors.forename: Yoshikatsu
authors.initials: Y
authors.lastname: Aikawa
authors.forename: Xiaofeng
authors.initials: X
authors.lastname: Xia
authors.forename: Thomas F J
authors.initials: TF
authors.lastname: Martin
mesh_terms: D000089982:ADP-Ribosylation Factor 6; D020727:ADP-Ribosylation Factors / Q000502:physiology*; D000199:Actins / Q000502:physiology; D000818:Animals; D020126:Brefeldin A / Q000494:pharmacology; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D034281:Dynamins / Q000502:physiology; D004705:Endocytosis / Q000502:physiology; D011992:Endosomes / Q000187:drug effects / Q000502:physiology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D051379:Mice; D009490:Neurosecretory Systems / Q000166:cytology / Q000378:metabolism*; D016716:PC12 Cells; D019269:Phosphatidylinositol 4,5-Diphosphate / Q000502:physiology; D051381:Rats; D050825:Synaptosomal-Associated Protein 25 / Q000032:analysis / Q000378:metabolism*; D050827:Syntaxin 1 / Q000378:metabolism*
chemical_list: D000089982:ADP-Ribosylation Factor 6; D000199:Actins; D004791:Enzyme Inhibitors; D019269:Phosphatidylinositol 4,5-Diphosphate; C494209:Snap25 protein, mouse; D050825:Synaptosomal-Associated Protein 25; D050827:Syntaxin 1; D020126:Brefeldin A; D020727:ADP-Ribosylation Factors; C000715727:Arf6 protein, mouse; C000715349:Arf6 protein, rat; D034281:Dynamins