scholarly journals HOPS Initiates Vacuole Docking by Tethering Membranes before trans-SNARE Complex Assembly

2010 ◽  
Vol 21 (13) ◽  
pp. 2297-2305 ◽  
Author(s):  
Christopher M. Hickey ◽  
William Wickner
Keyword(s):  
Complex Formation ◽  
Present Model ◽  
Snare Complex ◽  
Snare Proteins ◽  
Membrane Association ◽  
Attachment Protein ◽  
Protein Receptor ◽  
Sensitive Factor ◽  
Rab Effector ◽  
Snare Complex Formation

Vacuole homotypic fusion has been reconstituted with all purified components: vacuolar lipids, four soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins, Sec17p, Sec18p, the Rab Ypt7p, and the hexameric homotypic fusion and vacuole protein sorting complex (HOPS). HOPS is a Rab-effector with direct affinity for SNAREs (presumably via its Sec1-Munc18 homologous subunit Vps33p) and for certain vacuolar lipids. Each of these pure vacuolar proteins was required for optimal proteoliposome clustering, raising the question of which was most directly involved. We now present model subreactions of clustering and fusion that reveal that HOPS is the direct agent of tethering. The Rab and vacuole lipids contribute to tethering by supporting the membrane association of HOPS. HOPS indirectly facilitates trans-SNARE complex formation by tethering membranes, because the synthetic liposome tethering factor polyethylene glycol can also stimulate trans-SNARE complex formation and fusion. SNAREs further stabilize the associations of HOPS-tethered membranes. HOPS then protects newly formed trans-SNARE complexes from disassembly by Sec17p/Sec18p.

scholarly journals Regulation of exocytosis by the exocyst subunit Sec6 and the SM protein Sec1

2012 ◽  
Vol 23 (2) ◽  
pp. 337-346 ◽  
Author(s):  
Francesca Morgera ◽  
Margaret R. Sallah ◽  
Michelle L. Dubuke ◽  
Pallavi Gandhi ◽  
Daniel N. Brewer ◽  
...  
Keyword(s):  
Membrane Fusion ◽  
Secretory Pathway ◽  
Secretory Vesicle ◽  
Snare Complex ◽  
Attachment Protein ◽  
Protein Receptor ◽  
Sensitive Factor ◽  
Membrane Bound ◽  
Sm Protein ◽  
Snare Complex Formation

Trafficking of protein and lipid cargo through the secretory pathway in eukaryotic cells is mediated by membrane-bound vesicles. Secretory vesicle targeting and fusion require a conserved multisubunit protein complex termed the exocyst, which has been implicated in specific tethering of vesicles to sites of polarized exocytosis. The exocyst is directly involved in regulating soluble N-ethylmaleimide–sensitive factor (NSF) attachment protein receptor (SNARE) complexes and membrane fusion through interactions between the Sec6 subunit and the plasma membrane SNARE protein Sec9. Here we show another facet of Sec6 function—it directly binds Sec1, another SNARE regulator, but of the Sec1/Munc18 family. The Sec6–Sec1 interaction is exclusive of Sec6–Sec9 but compatible with Sec6–exocyst assembly. In contrast, the Sec6–exocyst interaction is incompatible with Sec6–Sec9. Therefore, upon vesicle arrival, Sec6 is proposed to release Sec9 in favor of Sec6–exocyst assembly and to simultaneously recruit Sec1 to sites of secretion for coordinated SNARE complex formation and membrane fusion.

scholarly journals Sec1/Munc18 protein Vps33 binds to SNARE domains and the quaternary SNARE complex

2012 ◽  
Vol 23 (23) ◽  
pp. 4611-4622 ◽  
Author(s):  
Braden T. Lobingier ◽  
Alexey J. Merz
Keyword(s):  
Protein Function ◽  
Snare Complex ◽  
Snare Proteins ◽  
Missense Mutations ◽  
Attachment Protein ◽  
Protein Receptor ◽  
Biochemical Studies ◽  
Sensitive Factor ◽  
Sm Protein ◽  
Endocytic Traffic

Soluble N-ethylmaleimide–sensitive factor attachment protein receptor (SNARE) proteins catalyze membrane fusion events in the secretory and endolysosomal systems, and all SNARE-mediated fusion processes require cofactors of the Sec1/Munc18 (SM) family. Vps33 is an SM protein and subunit of the Vps-C complexes HOPS (homotypic fusion and protein sorting) and CORVET (class C core vacuole/endosome tethering), which are central regulators of endocytic traffic. Here we present biochemical studies of interactions between Saccharomyces cerevisiae vacuolar SNAREs and the HOPS holocomplex or Vps33 alone. HOPS binds the N-terminal Habc domain of the Qa-family SNARE Vam3, but Vps33 is not required for this interaction. Instead, Vps33 binds the SNARE domains of Vam3, Vam7, and Nyv1. Vps33 directly binds vacuolar quaternary SNARE complexes, and the affinity of Vps33 for SNARE complexes is greater than for individual SNAREs. Through targeted mutational analyses, we identify missense mutations of Vps33 that produce a novel set of defects, including cargo missorting and the loss of Vps33-HOPS association. Together these data suggest a working model for membrane docking: HOPS associates with N-terminal domains of Vam3 and Vam7 through Vps33-independent interactions, which are followed by binding of Vps33, the HOPS SM protein, to SNARE domains and finally to the quaternary SNARE complex. Our results also strengthen the hypothesis that SNARE complex binding is a core attribute of SM protein function.

scholarly journals Dual inhibition of SNARE complex formation by tomosyn ensures controlled neurotransmitter release

2008 ◽  
Vol 183 (2) ◽  
pp. 323-337 ◽  
Author(s):  
Toshiaki Sakisaka ◽  
Yasunori Yamamoto ◽  
Sumiko Mochida ◽  
Michiko Nakamura ◽  
Kouki Nishikawa ◽  
...  
Keyword(s):  
Complex Formation ◽  
Neurotransmitter Release ◽  
Snare Complex ◽  
Protein Receptor ◽  
Repeat Domain ◽  
Dual Inhibition ◽  
Presynaptic Nerve Terminals ◽  
Synaptic Vesicle Fusion ◽  
Deficient Mice ◽  
Snare Complex Formation

Neurotransmitter release from presynaptic nerve terminals is regulated by soluble NSF attachment protein receptor (SNARE) complex–mediated synaptic vesicle fusion. Tomosyn inhibits SNARE complex formation and neurotransmitter release by sequestering syntaxin-1 through its C-terminal vesicle-associated membrane protein (VAMP)–like domain (VLD). However, in tomosyn-deficient mice, the SNARE complex formation is unexpectedly decreased. In this study, we demonstrate that the N-terminal WD-40 repeat domain of tomosyn catalyzes the oligomerization of the SNARE complex. Microinjection of the tomosyn N-terminal WD-40 repeat domain into neurons prevented stimulated acetylcholine release. Thus, tomosyn inhibits neurotransmitter release by catalyzing oligomerization of the SNARE complex through the N-terminal WD-40 repeat domain in addition to the inhibitory activity of the C-terminal VLD.

Distinct Initial SNARE Configurations Underlying the Diversity of Exocytosis

2012 ◽  
Vol 92 (4) ◽  
pp. 1915-1964 ◽  
Author(s):  
Haruo Kasai ◽  
Noriko Takahashi ◽  
Hiroshi Tokumaru
Keyword(s):  
Membrane Trafficking ◽  
Synaptic Vesicles ◽  
Cell Types ◽  
Snare Proteins ◽  
Attachment Protein ◽  
Protein Receptor ◽  
Large Dense Core Vesicles ◽  
Sensitive Factor ◽  
The Common ◽  
Kinetics Of

The dynamics of exocytosis are diverse and have been optimized for the functions of synapses and a wide variety of cell types. For example, the kinetics of exocytosis varies by more than five orders of magnitude between ultrafast exocytosis in synaptic vesicles and slow exocytosis in large dense-core vesicles. However, in all cases, exocytosis is mediated by the same fundamental mechanism, i.e., the assembly of soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins. It is often assumed that vesicles need to be docked at the plasma membrane and SNARE proteins must be preassembled before exocytosis is triggered. However, this model cannot account for the dynamics of exocytosis recently reported in synapses and other cells. For example, vesicles undergo exocytosis without prestimulus docking during tonic exocytosis of synaptic vesicles in the active zone. In addition, epithelial and hematopoietic cells utilize cAMP and kinases to trigger slow exocytosis of nondocked vesicles. In this review, we summarize the manner in which the diversity of exocytosis reflects the initial configurations of SNARE assembly, including trans-SNARE, binary-SNARE, unitary-SNARE, and cis-SNARE configurations. The initial SNARE configurations depend on the particular SNARE subtype (syntaxin, SNAP25, or VAMP), priming proteins (Munc18, Munc13, CAPS, complexin, or snapin), triggering proteins (synaptotagmins, Doc2, and various protein kinases), and the submembraneous cytomatrix, and they are the key to determining the kinetics of subsequent exocytosis. These distinct initial configurations will help us clarify the common SNARE assembly processes underlying exocytosis and membrane trafficking in eukaryotic cells.

VAMP-8 segregates mast cell–preformed mediator exocytosis from cytokine trafficking pathways

Blood ◽  
2008 ◽  
Vol 111 (7) ◽  
pp. 3665-3674 ◽  
Author(s):  
Neeraj Tiwari ◽  
Cheng-Chun Wang ◽  
Cristiana Brochetta ◽  
Gou Ke ◽  
Francesca Vita ◽  
...  
Keyword(s):  
Mast Cells ◽  
Inflammatory Responses ◽  
Secretory Granules ◽  
Snare Complex ◽  
Protein Receptor ◽  
Sensitive Factor ◽  
Syntaxin 4 ◽  
Deficient Mice ◽  
Snare Complex Formation ◽  
Vesicular Compartment

Abstract Inflammatory responses by mast cells are characterized by massive exocytosis of prestored granular mediators followed by cytokine/chemokine release. The vesicular trafficking mechanisms involved remain poorly understood. Vesicular-associated membrane protein-8 (VAMP-8), a member of the soluble N-ethylmaleimide–sensitive factor (NSF) attachment protein receptor (SNARE) family of fusion proteins initially characterized in endosomal and endosomal-lysosomal fusion, may also function in regulated exocytosis. Here we show that in bone marrow–derived mast cells (BMMCs) VAMP-8 partially colocalized with secretory granules and redistributed upon stimulation. This was associated with increased SNARE complex formation with the target t-SNAREs, SNAP-23 and syntaxin-4. VAMP-8–deficient BMMCs exhibited a markedly reduced degranulation response after IgE+ antigen-, thapsigargin-, or ionomycin-induced stimulation. VAMP-8–deficient mice also showed reduced plasma histamine levels in passive systemic anaphylaxis experiments, while cytokine/chemokine release was not affected. Unprocessed TNF accumulated at the plasma membrane where it colocalized with a VAMP-3–positive vesicular compartment but not with VAMP-8. The findings demonstrate that VAMP-8 segregates secretory lysosomal granule exocytosis in mast cells from cytokine/chemokine molecular trafficking pathways.

scholarly journals Endosomal and Phagosomal SNAREs

2018 ◽  
Vol 98 (3) ◽  
pp. 1465-1492 ◽  
Author(s):  
Ilse Dingjan ◽  
Peter T. A. Linders ◽  
Danielle R. J. Verboogen ◽  
Natalia H. Revelo ◽  
Martin ter Beest ◽  
...  
Keyword(s):  
Intracellular Transport ◽  
Snare Proteins ◽  
Intracellular Pathogens ◽  
Snare Protein ◽  
Attachment Protein ◽  
System A ◽  
Protein Receptor ◽  
Sensitive Factor ◽  
Organelle Communication ◽  
Factor Attachment Protein Receptor

The soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) protein family is of vital importance for organelle communication. The complexing of cognate SNARE members present in both the donor and target organellar membranes drives the membrane fusion required for intracellular transport. In the endocytic route, SNARE proteins mediate trafficking between endosomes and phagosomes with other endosomes, lysosomes, the Golgi apparatus, the plasma membrane, and the endoplasmic reticulum. The goal of this review is to provide an overview of the SNAREs involved in endosomal and phagosomal trafficking. Of the 38 SNAREs present in humans, 30 have been identified at endosomes and/or phagosomes. Many of these SNAREs are targeted by viruses and intracellular pathogens, which thereby reroute intracellular transport for gaining access to nutrients, preventing their degradation, and avoiding their detection by the immune system. A fascinating picture is emerging of a complex transport network with multiple SNAREs being involved in consecutive trafficking routes.

scholarly journals How Could SNARE Proteins Open a Fusion Pore?

Physiology ◽  
2014 ◽  
Vol 29 (4) ◽  
pp. 278-285 ◽  
Author(s):  
Qinghua Fang ◽  
Manfred Lindau
Keyword(s):  
Conformational Change ◽  
Pore Formation ◽  
Vesicle Fusion ◽  
Neurosecretory Cells ◽  
Snare Complex ◽  
Snare Proteins ◽  
Fusion Pore ◽  
Receptor Complex ◽  
Attachment Protein ◽  
Protein Receptor

The SNARE (Soluble NSF Attachment protein REceptor) complex, which in mammalian neurosecretory cells is composed of the proteins synaptobrevin 2 (also called VAMP2), syntaxin, and SNAP-25, plays a key role in vesicle fusion. In this review, we discuss the hypothesis that, in neurosecretory cells, fusion pore formation is directly accomplished by a conformational change in the SNARE complex via movement of the transmembrane domains.

scholarly journals Localization, Dynamics, and Protein Interactions Reveal Distinct Roles for ER and Golgi SNAREs

1998 ◽  
Vol 141 (7) ◽  
pp. 1489-1502 ◽  
Author(s):  
Jesse C. Hay ◽  
Judith Klumperman ◽  
Viola Oorschot ◽  
Martin Steegmaier ◽  
Christin S. Kuo ◽  
...  
Keyword(s):  
Membrane Fusion ◽  
Protein Interactions ◽  
The Other ◽  
Snare Proteins ◽  
Attachment Protein ◽  
Golgi Transport ◽  
Protein Receptor ◽  
Sensitive Factor ◽  
Coated Membranes ◽  
Factor Attachment Protein Receptor

ER-to-Golgi transport, and perhaps intraGolgi transport involves a set of interacting soluble N-ethylmaleimide–sensitive factor attachment protein receptor (SNARE) proteins including syntaxin 5, GOS-28, membrin, rsec22b, and rbet1. By immunoelectron microscopy we find that rsec22b and rbet1 are enriched in COPII-coated vesicles that bud from the ER and presumably fuse with nearby vesicular tubular clusters (VTCs). However, all of the SNAREs were found on both COPII- and COPI-coated membranes, indicating that similar SNARE machinery directs both vesicle pathways. rsec22b and rbet1 do not appear beyond the first Golgi cisterna, whereas syntaxin 5 and membrin penetrate deeply into the Golgi stacks. Temperature shifts reveal that membrin, rsec22b, rbet1, and syntaxin 5 are present together on membranes that rapidly recycle between peripheral and Golgi-centric locations. GOS-28, on the other hand, maintains a fixed localization in the Golgi. By immunoprecipitation analysis, syntaxin 5 exists in at least two major subcomplexes: one containing syntaxin 5 (34-kD isoform) and GOS-28, and another containing syntaxin 5 (41- and 34-kD isoforms), membrin, rsec22b, and rbet1. Both subcomplexes appear to involve direct interactions of each SNARE with syntaxin 5. Our results indicate a central role for complexes among rbet1, rsec22b, membrin, and syntaxin 5 (34 and 41 kD) at two membrane fusion interfaces: the fusion of ER-derived vesicles with VTCs, and the assembly of VTCs to form cis-Golgi elements. The 34-kD syntaxin 5 isoform, membrin, and GOS-28 may function in intraGolgi transport.

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