The SM protein Vps33 and the t-SNARE Habc domain promote fusion pore opening

Pieren, Michel; Schmidt, Andrea; Mayer, Andreas

doi:10.1038/nsmb.1809

Cited by 62 publications

(85 citation statements)

References 60 publications

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“…In contrast to its essential roles in metazoan membrane transport, the N-peptide motif of syntaxin seems to be dispensable for many yeast fusion pathways under normal growth conditions (60,61). Moreover, the N-peptide binding mode is entirely absent in the yeast SM proteins Sec1p and Vps33p (62)(63)(64). At first glance, these functional discrepancies conflict with the initiation factor model suggested here.…”

Section: Discussioncontrasting

confidence: 49%

Syntaxin N-terminal peptide motif is an initiation factor for the assembly of the SNARE–Sec1/Munc18 membrane fusion complex

Rathore

Bend

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

114

142

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Intracellular membrane fusion is mediated by the concerted action of N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs) and Sec1/Munc18 (SM) proteins. During fusion, SM proteins bind the N-terminal peptide (N-peptide) motif of the SNARE subunit syntaxin, but the function of this interaction is unknown. Here, using FRET-based biochemical reconstitution and Caenorhabditis elegans genetics, we show that the N-peptide of syntaxin-1 recruits the SM protein Munc18-1/nSec1 to the SNARE bundle, facilitating their assembly into a fusion-competent complex. The recruitment is achieved through physical tethering rather than allosteric activation of Munc18-1. Consistent with the recruitment role, the Npeptide is not spatially constrained along syntaxin-1, and it is functional when translocated to another SNARE subunit SNAP-25 or even when simply anchored in the target membrane. The N-peptide function is restricted to an early initiation stage of the fusion reaction. After association, Munc18-1 and the SNARE bundle together drive membrane merging without further involving the N-peptide. Thus, the syntaxin N-peptide is an initiation factor for the assembly of the SNARE-SM membrane fusion complex.I ntracellular membrane fusion is the basis of a wide range of fundamental biological processes, including organelle maintenance, hormone secretion, and inside-outside distribution of receptors and transporters. The merging of intracellular membrane bilayers is mediated by a fusion complex comprised of SNAREs and Sec1/Munc18 (SM) proteins (1). The core of the fusion machinery is the trans-SNARE complex (SNAREpin) formed by the pairing of the vesicle-rooted SNARE (v-SNARE) with the target membrane-associated SNAREs (t-SNAREs) (2-5). N-to C-terminal zippering of the trans-SNARE complex brings two membranes into close apposition and helps to overcome the energy barrier for fusion (6-10). SM proteins are soluble factors of 60-70 kDa that directly interact with their cognate trans-SNARE complexes to promote the speed and specificity of a fusion reaction (11)(12)(13)(14).Each fusion pathway in the cell requires a specific subset of SNAREs and SM proteins (15). The most intensely studied form of intracellular membrane fusion is calcium-triggered neurotransmitter release at the chemical synapse, which serves as the brain's major form of cell-cell communication (15)(16)(17)(18)(19). Neurotransmitter secretion is mediated by the fusion of exocytic vesicles with the plasma membrane and requires the v-SNARE vesicle-associated membrane protein 2 (VAMP2; also known as synaptobrevin-2), the t-SNAREs syntaxin-1 and soluble N-ethylmaleimide-sensitive factor attachment protein (SNAP)-25, and the SM protein Munc18-1/ nSec1 (UNC-18 in nematodes and ROP in flies) (20-28).The interaction between SNAREs and SM proteins involves multiple binding modes. The primary target of SM proteins is believed to be the four-helix SNARE bundle (29-31). Assembled from the SNARE motifs and the transmembrane domains of t-and vSNAREs (4, 5), the SNARE bun...

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Section: Discussioncontrasting

confidence: 49%

Syntaxin N-terminal peptide motif is an initiation factor for the assembly of the SNARE–Sec1/Munc18 membrane fusion complex

Rathore

Bend

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

114

142

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“…Monoclonal anti-hemagglutinin (anti-HA) antibodies (mouse ascites, Covance); poly-peptone and yeast extract (Pronadisa); dextrose and yeast nitrogen base without amino acids and ammonium sulfate (Difco). Polyclonal antibodies against Nyv1, Vam3, Sec17, Sec18, Vtc1 and Vtc4 were raised in rabbits or goats and were prepared as described previously (Müller et al, 2002;Pieren et al, 2010). The lytic enzyme (lyticase), recombinant Sec18, recombinant Ppx1 and recombinant Vam7 were expressed and purified as described previously (Gerasimaite et al, 2014;Müller et al, 2002;Stroupe et al, 2006).…”

Section: Methodsmentioning

confidence: 99%

“…Therefore, we tested whether their deficiency in SNARE activation might be due to altered interaction of Sec18/NSF with the SNARE complex. Vacuoles were isolated from cells that expressed the SNARE Vam3 with a Cterminal HA tag (Pieren et al, 2010). The membranes were treated with the cleavable crosslinker DTSSP, solubilized in detergent, and immunoprecipitated with anti-HA antibodies.…”

Section: Sec18 Is Recruited To Snare Complexes Only When Polyphosphatmentioning

confidence: 99%

Organelle size control – increasing vacuole content activates SNAREs to augment organelle volume through homotypic fusion

Desfougères

Neumann²,

Mayer

2016

Journal of Cell Science

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Cells control the size of their compartments relative to cell volume, but there is also size control within each organelle. Yeast vacuoles neither burst nor do they collapse into a ruffled morphology, indicating that the volume of the organellar envelope is adjusted to the amount of content. It is poorly understood how this adjustment is achieved. We show that the accumulating content of yeast vacuoles activates fusion of other vacuoles, thus increasing the volume-to-surface ratio. Synthesis of the dominant compound stored inside vacuoles, polyphosphate, stimulates binding of the chaperone Sec18/NSF to vacuolar SNAREs, which activates them and triggers fusion. SNAREs can only be activated by lumenal, not cytosolic, polyphosphate ( polyP). Control of lumenal polyP over SNARE activation in the cytosol requires the cytosolic cyclin-dependent kinase Pho80-Pho85 and the R-SNARE Nyv1. These results suggest that cells can adapt the volume of vacuoles to their content through feedback from the vacuole lumen to the SNAREs on the cytosolic surface of the organelle.

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“…Fig. 4A) (22,23). This fusion deficiency could be rescued by titrating HOPS into the fusion reaction (Fig.…”

Section: A Role Of the H Abc Domain In Hops Function Duringmentioning

confidence: 99%

“…domain in promoting efficient fusion (23). This nevertheless left the question open, how HOPS binding to the H abc domain of Vam3 and the SNARE complex might be coordinated.…”

mentioning

confidence: 99%

The Habc Domain of the SNARE Vam3 Interacts with the HOPS Tethering Complex to Facilitate Vacuole Fusion

Lürick¹,

Kuhlee

Bröcker³

et al. 2015

Journal of Biological Chemistry

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The SM protein Vps33 and the t-SNARE Habc domain promote fusion pore opening

Cited by 62 publications

References 60 publications

Syntaxin N-terminal peptide motif is an initiation factor for the assembly of the SNARE–Sec1/Munc18 membrane fusion complex

Syntaxin N-terminal peptide motif is an initiation factor for the assembly of the SNARE–Sec1/Munc18 membrane fusion complex

Organelle size control – increasing vacuole content activates SNAREs to augment organelle volume through homotypic fusion

The Habc Domain of the SNARE Vam3 Interacts with the HOPS Tethering Complex to Facilitate Vacuole Fusion

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