Presynaptic events involved in neurotransmission

Nelson, Nathan

doi:10.1016/0928-4257(93)90028-r

Cited by 32 publications

(24 citation statements)

References 42 publications

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“…The Ac39 and Ac116 subunits of V-ATPase had already been shown to be highly enriched in mammalian SVs (33,34,39) where the proton pump plays a crucial role in the uptake of neurotransmitters. However, the presence of the V-ATPase c subunit on mammalian SVs had never been demonstrated.…”

Section: Discussionmentioning

confidence: 99%

“…Freeze-thawing, in contrast, promotes a massive association of synaptobrevin 2 with t-SNAREs to form SNARE complexes. Freeze-thawing may induce a conformational change in synaptobrevin 2 which mimics a physiological change occurring in vivo and which is part of the events leading to exocytosis.The Ac39 and Ac116 subunits of V-ATPase had already been shown to be highly enriched in mammalian SVs (33,34,39) where the proton pump plays a crucial role in the uptake of neurotransmitters. However, the presence of the V-ATPase c subunit on mammalian SVs had never been demonstrated.…”

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The V Sector of the V-ATPase, Synaptobrevin, and Synaptophysin Are Associated on Synaptic Vesicles in a Triton X-100-resistant, Freeze-thawing Sensitive, Complex

Galli¹,

McPherson

Camilli

1996

Journal of Biological Chemistry

130

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Anti-synaptobrevin 2 immunoprecipitates obtained from freshly prepared Triton X-100 extracts of rat synaptosomes contained, in addition to synaptophysin, a 10-kDa band, which we identified by peptide sequencing and Western blotting as the c subunit of the vacuolar proton pump (V-ATPase) also called ductin or mediatophore. Ac39 and Ac116, two other transmembrane subunits of the V 0 sector of the V-ATPase, were also found by Western blotting to be enriched in the immunoprecipitates. None of these V-ATPase subunits, or synaptophysin, was present in anti-synaptobrevin 2 immunoprecipitates obtained from frozen-thawed Triton X-100 extracts, which were greatly enriched, instead, in SNAP-25 and syntaxin 1. Accordingly, V-ATPase subunit c was found in anti-synaptophysin immunoprecipitates generated from fresh, but not frozen-thawed extracts, and was not found in anti-syntaxin 1 immunoprecipitates. Thus, the two complexes appear to be mutually exclusive. Subcellular fractionation of rat brain demonstrated that V-ATPase subunit c is localized with synaptobrevin 2 and synaptophysin in synaptic vesicles. The coprecipitation of V-ATPase subunit c with the synaptobrevin-synaptophysin complex suggests that this interaction may play a role in recruiting the proton pump into synaptic vesicles. Freeze-thawing, which involves a mild denaturing step, may produce a conformational change which dissociates the complex and mimics a change which occurs in vivo as a prerequisite to SNARE complex formation. Synaptic vesicles (SV),1 the vesicular carriers responsible for secretion of non-peptide neurotransmitters at synapses, are a very useful experimental model for investigating fundamental mechanisms in vesicular traffic (1-4). Studies carried out on SVs have converged with studies on neurotoxins, genetic studies in yeast, and cell-free studies of vesicular transport, and allowed to formulate a fist hypothetical model of docking and fusion (5). The model predicts that small proteins located at the cytosolic surface of the vesicular carrier, v-SNAREs, interact with small proteins located at the cytosolic surface of the target membrane, t-SNAREs, to a form the so-called SNARE complex, and that this interaction is one of the key steps leading to fusion (5,6). In the case of SVs, v-SNAREs are the synaptobrevins and t-SNAREs are syntaxin and SNAP-25 (5).Formation of SNARE complexes, and in particular of the synaptic SNARE complex, must be highly regulated. The presence of the same v-SNAREs on different types of secretory organelles, and of t-SNAREs all along the plasmalemma of neurons, clearly indicates that SNAREs are not themselves responsible for the spatiotemporal regulation of exocytosis (7-9). A variety of other factors which control formation and dissociation of the SNARE complex have already been identified (for review, see Ref. 10).Studies on purified SVs have demonstrated that at least a significant fraction of synaptobrevin is in a complex with synaptophysin, and that this complex is mutually exclusive with the interaction of synapt...

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Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

The V Sector of the V-ATPase, Synaptobrevin, and Synaptophysin Are Associated on Synaptic Vesicles in a Triton X-100-resistant, Freeze-thawing Sensitive, Complex

Galli¹,

McPherson

Camilli

1996

Journal of Biological Chemistry

130

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“…This occurs in the vacuoles of S. cerevisiae in response to glucose deprivation (64), and in goblet cell apical membranes of M. sexta during moulting or starving of the larvae (65,66). Whether reversible dissociation of V 1 V 0 might also have a regulatory role in chromaffin cells, or whether V 0 itself might have an independent function, for example in exocytosis, is still a matter for speculation (67,68). It is noteworthy that in synaptic vesicles V 0 appears to exist in a complex with the vesicle membrane proteins synaptobrevin and synaptophysin (69).…”

Section: 2-kda Protein Of V-atpase From Chromaffin Granulesmentioning

confidence: 99%

Identification and Characterization of a Novel 9.2-kDa Membrane Sector-associated Protein of Vacuolar Proton-ATPase from Chromaffin Granules

Ludwig¹,

Kerscher²,

Brandt³

et al. 1998

Journal of Biological Chemistry

285

232

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Vacuolar proton-translocating ATPase (holoATPase and free membrane sector) was isolated from bovine chromaffin granules by blue native polyacrylamide gel electrophoresis. A 5-fold excess of membrane sector over holoenzyme was determined in isolated chromaffin granule membranes. M9.2, a novel extremely hydrophobic 9.2-kDa protein comprising 80 amino acids, was detected in the membrane sector. It shows sequence and structural similarity to Vma21p, a yeast protein required for assembly of vacuolar ATPase. A second membrane sector-associated protein (M8-9) was identified and characterized by aminoterminal protein sequencing.Proton-translocating adenosine triphosphatases have fundamental roles in energy conservation, secondary active transport, the acidification of intracellular compartments, and cellular pH homeostasis. They fall into three broad classes, called F, P, and V (1), of which the vacuolar type (V-ATPases) 1 is both the most recently recognized and the least well characterized. ATPases of this class occur in endomembranes bounding the acidic compartments of animal, plant, and fungal cells (2) and also in the plasma membranes of some specialized cell types. They have been purified from several mammalian sources, including adrenal secretory vesicles (3, 4), brain clathrincoated vesicles, (5, 6), and kidney medulla microsomes (7), as well as from the vacuoles of fungi and higher plants. Most V-ATPases contain some 6 -10 different subunits (2), but subunit composition depends on the source of the enzyme, and tissue-specific isoforms exist (8). The V-type ATPases are structurally similar to those of the F-type, having a transmembrane proton-conducting sector and an extramembrane catalytic sector. By analogy with the two sectors of F-ATPases (9 -12), these are termed V 0 and V 1 , respectively. For a recent review, see Ref. 13.In this work, the recently developed technique of blue native polyacrylamide gel electrophoresis (BN-PAGE; Refs. 14 -17) was employed to purify vacuolar ATPase holoenzyme (V 1 V 0 ) and free membrane sector (V 0 ) simultaneously from adrenal secretory vesicle membranes. Combined with high resolution Tricine-SDS-PAGE in the second dimension, the subunit composition, particularly with respect to small polypeptides, was determined. Two novel proteins, 8 -9 and 9.2 kDa in size, were found in the membrane sector. Here we report the detailed analysis of the larger of these two polypeptides. EXPERIMENTAL PROCEDURESMaterials-Restriction enzymes and T4-DNA ligase were obtained from New England Biolabs. Taq DNA polymerase was from Stratagene, and TA Cloning Kit ® was from Invitrogen. Sequenase version 2.0 sequencing kit, [␣-35 S]dATP and Hybond N ϩ membranes were obtained from Amersham Pharmacia Biotech. ABI Prism™ dye terminator cycle sequencing kit was purchased from Perkin Elmer. The cDNA library from bovine adrenal medulla was a kind gift from Leonora Ciufo (University of Edinburgh, Edinburgh, Scotland, United Kingdom). Human EST clone ID 143553 (GenBank™accession number R75754) was obtained fr...

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“…The secretory granules have a luminal pH of 5.0-5.5 (Arvan and Castle, 1986;Orci et al, 1986;Urbe et al, 1997), and their acidification has important roles in the condensation and maturation of their contents (Hutton, 1994). The electrochemical proton gradient across the membrane provides the energy for accumulation of neurotransmitters into synaptic vesicles (Moriyama and Futai, 1990;Nelson, 1993). Many growth factors are produced as latent inactive forms within the endoplasmic reticulum and undergo several modifications during travel through the Golgi apparatus, before being converted to mature active forms within the secretory vesicles.…”

Section: Introductionmentioning

confidence: 99%

Thea3 isoform of V-ATPase regulates insulin secretion from pancreatic β-cells

Sun‐Wada

Toyomura

Murata

et al. 2006

Journal of Cell Science

200

175

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Vacuolar-type H+-ATPase (V-ATPase) is a multi-subunit enzyme that has important roles in the acidification of a variety of intracellular compartments and some extracellular milieus. Four isoforms for the membrane-intrinsic subunit (subunit a) of the V-ATPase have been identified in mammals, and they confer distinct cellular localizations and activities on the proton pump. We found that V-ATPase with the a3 isoform is highly expressed in pancreatic islets, and is localized to membranes of insulin-containing secretory granules in β-cells. oc/oc mice, which have a null mutation at the a3 locus, exhibited a reduced level of insulin in the blood, even with high glucose administration. However, islet lysates contained mature insulin, and the ratio of the amount of insulin to proinsulin in oc/oc islets was similar to that of wild-type islets, indicating that processing of insulin was normal even in the absence of the a3 function. The insulin contents of oc/oc islets were reduced slightly, but this was not significant enough to explain the reduced levels of the blood insulin. The secretion of insulin from isolated islets in response to glucose or depolarizing stimulation was impaired. These results suggest that the a3 isoform of V-ATPase has a regulatory function in the exocytosis of insulin secretion.

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Presynaptic events involved in neurotransmission

Cited by 32 publications

References 42 publications

The V Sector of the V-ATPase, Synaptobrevin, and Synaptophysin Are Associated on Synaptic Vesicles in a Triton X-100-resistant, Freeze-thawing Sensitive, Complex

The V Sector of the V-ATPase, Synaptobrevin, and Synaptophysin Are Associated on Synaptic Vesicles in a Triton X-100-resistant, Freeze-thawing Sensitive, Complex

Identification and Characterization of a Novel 9.2-kDa Membrane Sector-associated Protein of Vacuolar Proton-ATPase from Chromaffin Granules

Thea3 isoform of V-ATPase regulates insulin secretion from pancreatic β-cells

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