Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Dynamin-SNARE interactions control trans-SNARE formation in intracellular membrane fusion.

ABSTRACT: The fundamental processes of membrane fission and fusion determine size and copy numbers of intracellular organelles. Although SNARE proteins and tethering complexes mediate intracellular membrane fusion, fission requires the presence of dynamin or dynamin-related proteins. Here we study these reactions in native yeast vacuoles and find that the yeast dynamin homologue Vps1 is not only an essential part of the fission machinery, but also controls membrane fusion by generating an active Qa SNARE-tethering complex pool, which is essential for trans-SNARE formation. Our findings provide new insight into the role of dynamins in membrane fusion by directly acting on SNARE proteins.

SUBMITTER: Alpadi K

PROVIDER: S-EPMC3630463 | biostudies-other | 2013

REPOSITORIES: biostudies-other

ACCESS DATA

Json Xml

Publications

Dynamin-SNARE interactions control trans-SNARE formation in intracellular membrane fusion.

Alpadi Kannan K Kulkarni Aditya A Namjoshi Sarita S Srinivasan Sankaranarayanan S Sippel Katherine H KH Ayscough Kathryn K Zieger Martin M Schmidt Andrea A Mayer Andreas A Evangelista Michael M Quiocho Florante A FA Peters Christopher C

Nature communications 20130101

The fundamental processes of membrane fission and fusion determine size and copy numbers of intracellular organelles. Although SNARE proteins and tethering complexes mediate intracellular membrane fusion, fission requires the presence of dynamin or dynamin-related proteins. Here we study these reactions in native yeast vacuoles and find that the yeast dynamin homologue Vps1 is not only an essential part of the fission machinery, but also controls membrane fusion by generating an active Qa SNARE- ...[more]

PMID: 23591871

Similar Datasets

Sequential analysis of trans-SNARE formation in intracellular membrane fusion.

Project description:SNARE complexes are required for membrane fusion in the endomembrane system. They contain coiled-coil bundles of four helices, three (Q(a), Q(b), and Q(c)) from target (t)-SNAREs and one (R) from the vesicular (v)-SNARE. NSF/Sec18 disrupts these cis-SNARE complexes, allowing reassembly of their subunits into trans-SNARE complexes and subsequent fusion. Studying these reactions in native yeast vacuoles, we found that NSF/Sec18 activates the vacuolar cis-SNARE complex by selectively displacing the vacuolar Q(a) SNARE, leaving behind a Q(bc)R subcomplex. This subcomplex serves as an acceptor for a Q(a) SNARE from the opposite membrane, leading to Q(a)-Q(bc)R trans-complexes. Activity tests of vacuoles with diagnostic distributions of inactivating mutations over the two fusion partners confirm that this distribution accounts for a major share of the fusion activity. The persistence of the Q(bc)R cis-complex and the formation of the Q(a)-Q(bc)R trans-complex are both sensitive to the Rab-GTPase inhibitor, GDI, and to mutations in the vacuolar tether complex, HOPS (HOmotypic fusion and vacuolar Protein Sorting complex). This suggests that the vacuolar Rab-GTPase, Ypt7, and HOPS restrict cis-SNARE disassembly and thereby bias trans-SNARE assembly into a preferred topology.

| S-EPMC3260307 | biostudies-literature

CAPS drives trans-SNARE complex formation and membrane fusion through syntaxin interactions.

Project description:Ca(2+)-dependent activator protein for secretion (CAPS) is an essential factor for regulated vesicle exocytosis that functions in priming reactions before Ca(2+)-triggered fusion of vesicles with the plasma membrane. However, the precise events that CAPS regulates to promote vesicle fusion are unclear. In the current work, we reconstituted CAPS function in a SNARE-dependent liposome fusion assay using VAMP2-containing donor and syntaxin-1/SNAP-25-containing acceptor liposomes. The CAPS stimulation of fusion required PI(4,5)P(2) in acceptor liposomes and was independent of Ca(2+), but Ca(2+) dependence was restored by inclusion of synaptotagmin. CAPS stimulated trans-SNARE complex formation concomitant with the stimulation of full membrane fusion at physiological SNARE densities. CAPS bound syntaxin-1, and CAPS truncations that competitively inhibited syntaxin-1 binding also inhibited CAPS-dependent fusion. The results revealed an unexpected activity of a priming protein to accelerate fusion by efficiently promoting trans-SNARE complex formation. CAPS may function in priming by organizing SNARE complexes on the plasma membrane.

| S-EPMC2765074 | biostudies-literature

Phosphatidylinositol 3,5-bisphosphate regulates the transition between trans-SNARE complex formation and vacuole membrane fusion.

Project description:Phosphoinositides (PIs) regulate a myriad of cellular functions including membrane fusion, as exemplified by the yeast vacuole, which uses various PIs at different stages of fusion. In light of this, the effect of phosphatidylinositol 3,5-bisphosphate (PI(3,5)P2) on vacuole fusion remains unknown. PI(3,5)P2 is made by the PI3P 5-kinase Fab1 and has been characterized as a regulator of vacuole fission during hyperosmotic shock, where it interacts with the TRP Ca2+ channel Yvc1. Here we demonstrate that exogenously added dioctanoyl (C8) PI(3,5)P2 abolishes homotypic vacuole fusion. This effect was not linked to Yvc1, as fusion was equally affected using yvc1Δ vacuoles. Thus, the effects of C8-PI(3,5)P2 on fusion and fission operate through distinct mechanisms. Further testing showed that C8-PI(3,5)P2 inhibited vacuole fusion after trans-SNARE pairing. Although SNARE complex formation was unaffected, we found that C8-PI(3,5)P2 blocked outer leaflet lipid mixing. Overproduction of endogenous PI(3,5)P2 by the fab1T2250A hyperactive kinase mutant also inhibited the lipid mixing stage, bolstering the model in which PI(3,5)P2 inhibits fusion when present at elevated levels. Taken together, this work identifies a novel function for PI(3,5)P2 as a regulator of vacuolar fusion. Moreover, it suggests that this lipid acts as a molecular switch between fission and fusion.

| S-EPMC6589561 | biostudies-literature

Arrest of trans-SNARE zippering uncovers loosely and tightly docked intermediates in membrane fusion.

Project description:Soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins mediate intracellular membrane fusion in the secretory pathway. They contain conserved regions, termed SNARE motifs, that assemble between opposing membranes directionally from their N termini to their membrane-proximal C termini in a highly exergonic reaction. However, how this energy is utilized to overcome the energy barriers along the fusion pathway is still under debate. Here, we have used mutants of the SNARE synaptobrevin to arrest trans-SNARE zippering at defined stages. We have uncovered two distinct vesicle docking intermediates where the membranes are loosely and tightly connected, respectively. The tightly connected state is irreversible and independent of maintaining assembled SNARE complexes. Together, our results shed new light on the intermediate stages along the pathway of membrane fusion.

| S-EPMC5986196 | biostudies-literature

HOPS prevents the disassembly of trans-SNARE complexes by Sec17p/Sec18p during membrane fusion.

Project description:SNARE-dependent membrane fusion requires the disassembly of cis-SNARE complexes (formed by SNAREs anchored to one membrane) followed by the assembly of trans-SNARE complexes (SNAREs anchored to two apposed membranes). Although SNARE complex disassembly and assembly might be thought to be opposing reactions, the proteins promoting disassembly (Sec17p/Sec18p) and assembly (the HOPS complex) work synergistically to support fusion. We now report that trans-SNARE complexes formed during vacuole fusion are largely associated with Sec17p. Using a reconstituted proteoliposome fusion system, we show that trans-SNARE complex, like cis-SNARE complex, is sensitive to Sec17p/Sec18p mediated disassembly. Strikingly, HOPS inhibits the disassembly of SNARE complexes in the trans-, but not in the cis-, configuration. This selective HOPS preservation of trans-SNARE complexes requires HOPS:SNARE recognition and is lost when the apposed bilayers are dissolved in Triton X-100; it is also observed during fusion of isolated vacuoles. HOPS thus directs the Sec17p/Sec18p chaperone system to maximize functional trans-SNARE complex for membrane fusion, a new role of tethering factors during membrane traffic.

| S-EPMC2892374 | biostudies-literature

A dynamin homolog promotes the transition from hemifusion to content mixing in intracellular membrane fusion.

Project description:The convergence of the antagonistic reactions of membrane fusion and fission at the hemifusion/hemifission intermediate has generated a captivating enigma of whether Soluble N-ethylmaleimide sensitive factor Attachment Protein Receptor (SNAREs) and dynamin have unusual counter-functions in fission and fusion, respectively. SNARE-mediated fusion and dynamin-driven fission are fundamental membrane flux reactions known to occur during ubiquitous cellular communication events such as exocytosis, endocytosis and vesicle transport. Here we demonstrate the influence of the dynamin homolog Vps1 (Vacuolar protein sorting 1) on lipid mixing and content mixing properties of yeast vacuoles, and on the incorporation of SNAREs into fusogenic complexes. We propose a novel concept that Vps1, through its oligomerization and SNARE domain binding, promotes the hemifusion-content mixing transition in yeast vacuole fusion by increasing the number of trans-SNAREs.

| S-EPMC3976715 | biostudies-literature

Supramolecular SNARE assembly precedes hemifusion in SNARE-mediated membrane fusion.

Project description:Formation of the soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) complex facilitates intracellular membrane fusion. A single SNARE complex is thought to be insufficient; multiple copies of SNARE complexes must work cooperatively. However, the mechanism by which such a higher-order SNARE protein structure is assembled is unknown. EPR and fluorescence analyses show that at least three copies of target-membrane SNARE proteins self-assemble through the interaction between the transmembrane domains (TMDs), and this multimeric structure serves as scaffolding for trans-SNARE assembly. SNARE core formation in solution induces oligomerization of the TMDs of vesicle-associated SNAREs in the apposing membrane, transiently forming a supramolecular protein structure spanning two membranes. This higher-order protein intermediate evolves, by involving lipid molecules, to the hemifusion state. Hemifusion is subsequently followed by distal leaflet mixing and formation of the cis-SNARE complex.

| S-EPMC2575085 | biostudies-literature

Complex lipid requirements for SNARE- and SNARE chaperone-dependent membrane fusion.

Project description:Membrane fusion without lysis has been reconstituted with purified yeast vacuolar SNAREs (soluble N-ethylmaleimide-sensitive factor attachment protein receptors), the SNARE chaperones Sec17p/Sec18p and the multifunctional HOPS complex, which includes a subunit of the SNARE-interactive Sec1-Munc18 family, and vacuolar lipids: phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylinositol (PI), phosphatidylserine (PS), phosphatidic acid (PA), cardiolipin (CL), ergosterol (ERG), diacylglycerol (DAG), and phosphatidylinositol 3-phosphate (PI3P). We now report that many of these lipids are required for rapid and efficient fusion of the reconstituted SNARE proteoliposomes in the presence of SNARE chaperones. Omission of either PE, PA, or PI3P from the complete set of lipids strongly reduces fusion, and PC, PE, PA, and PI3P constitute a minimal set of lipids for fusion. PA could neither be replaced by other lipids with small headgroups such as DAG or ERG nor by the acidic lipids PS or PI. PA is needed for full association of HOPS and Sec18p with proteoliposomes having a minimal set of lipids. Strikingly, PA and PE are as essential for SNARE complex assembly as for fusion, suggesting that these lipids facilitate functional interactions among SNAREs and SNARE chaperones.

| S-EPMC2785640 | biostudies-literature

A lipid-anchored SNARE supports membrane fusion.

Project description:Intracellular membrane fusion requires R-SNAREs and Q-SNAREs to assemble into a four-helical parallel coiled-coil, with their hydrophobic anchors spanning the two apposed membranes. Based on the fusion properties of chemically defined SNARE- proteoliposomes, it has been proposed that the assembly of this helical bundle transduces force through the entire bilayer via the transmembrane SNARE anchor domains to drive fusion. However, an R-SNARE, Nyv1p, with a genetically engineered lipid anchor that spans half of the bilayer suffices for the fusion of isolated vacuoles, although this organelle has other R-SNAREs. To demonstrate unequivocally the fusion activity of lipid-anchored Nyv1p, we reconstituted proteoliposomes with purified lipid-anchored Nyv1p as the only protein. When these proteoliposomes were incubated with those bearing cognate Q-SNAREs, there was trans-SNARE complex assembly but, in accord with prior studies of the neuronal SNAREs, little lipid mixing. However, the addition of physiological fusion accessory proteins (HOPS, Sec17p, and Sec18p) allows lipid-anchored Nyv1p to support fusion, suggesting that trans-SNARE complex function is not limited to force transduction across the bilayers through the transmembrane domains.

| S-EPMC3198343 | biostudies-literature

Variable cooperativity in SNARE-mediated membrane fusion.

Project description:The soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) complex drives the majority of intracellular and exocytic membrane fusion events. Whether and how SNAREs cooperate to mediate fusion has been a subject of intense study, with estimates ranging from a single SNARE complex to 15. Here we show that there is no universally conserved number of SNARE complexes involved as revealed by our observation that this varies greatly depending on membrane curvature. When docking rates of small (∼40 nm) and large (∼100 nm) liposomes reconstituted with different synaptobrevin (the SNARE present in synaptic vesicles) densities are taken into account, the lipid mixing efficiency was maximal with small liposomes with only one synaptobrevin, whereas 23-30 synaptobrevins were necessary for efficient lipid mixing in large liposomes. Our results can be rationalized in terms of strong and weak cooperative coupling of SNARE complex assembly where each mode implicates different intermediate states of fusion that have been recently identified by electron microscopy. We predict that even higher variability in cooperativity is present in different physiological scenarios of fusion, and we further hypothesize that plasticity of SNAREs to engage in different coupling modes is an important feature of the biologically ubiquitous SNARE-mediated fusion reactions.

| S-EPMC4143004 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data