Botulinum neurotoxin light chains inhibit both Ca<sup>2+</sup>‐induced and GTP analogue‐induced catecholamine release from permeabilised adrenal chromaffin cells

Glenn, Daphne E.; Burgoyne, Robert D.

doi:10.1016/0014-5793(96)00432-2

Cited by 32 publications

(14 citation statements)

References 51 publications

(65 reference statements)

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“…However, this complex is not essential per se in the triggered fusion steps of exocytosis because in our work higher [Ca 2+ ]free overcame the block to fusion (seen at 'physiological' [Ca 2+ ]free) caused by proteolysis. Such recovery has also been documented in earlier experiments with Clostridial toxins (Dreyer and Schmitt, 1983;Ahnert-Hilger and Weller, 1993;Bittner and Holz, 1993;Lawrence et al, 1994;Glenn and Burgoyne, 1996;Lawrence et al, 1996;Capogna et al, 1997;Land et al, 1997;Fassio et al, 1999), suggesting that the concept of a regulatory complex may extend beyond the present system. In general, the testing and rescue of exocytosis at only one [Ca 2+ ]free after complex disruption tells us a great deal about the modulatory role of the complex.…”

Section: Snares and The Process Of Exocytosissupporting

confidence: 75%

Regulated secretion: SNARE density, vesicle fusion and calcium dependence

Coorssen

Blank

Albertorio

et al. 2003

Journal of Cell Science

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SNAREs such as VAMP, SNAP-25 and syntaxin are essential for intracellular trafficking, but what are their exact molecular roles and how are their interactions with other proteins manifest? Capitalizing on the differential sensitivity of SNAREs to exogenous proteases, we quantified the selective removal of identified SNAREs from native secretory vesicles without loss of fusion competence. Using previously established fusion assays and a high sensitivity immunoblotting protocol, we analyzed the relationship between these SNARE proteins and Ca2+-triggered membrane fusion. Neither the extent of fusion nor the number of intermembrane fusion complexes per vesicle were correlated with the measured density of identified egg cortical vesicle (CV) SNAREs. Without syntaxin, CVs remained fusion competent. Surprisingly, for one (but not another) protease the Ca2+dependence of fusion was correlated with CV SNARE density, suggesting a native protein complex that associates with SNAREs, the architecture of which ensures high Ca2+ sensitivity. As SNAREs may function during CV docking in vivo, and as further proteolysis after SNARE removal eventually ablates fusion, we hypothesize that the triggered steps of regulated fusion(Ca2+ sensitivity and the catalysis and execution of fusion)require additional proteins that function downstream of SNAREs.

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Section: Snares and The Process Of Exocytosissupporting

confidence: 75%

Regulated secretion: SNARE density, vesicle fusion and calcium dependence

Coorssen

Blank

Albertorio

et al. 2003

Journal of Cell Science

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“…5, 6), which until now has been understood poorly. One fact that we and others knew is that these two forms of exocytosis are both SNAREdependent (Banerjee et al, 1996;Glenn and Burgoyne, 1996;Wang et al, 2004). Whereas Martin and coworkers found that cytosolic proteins including phosphatidylcholine transfer protein (Hay and Martin, 1993), phosphatidylinositol phosphate kinases (Hay and Martin, 1993;Aikawa and Martin, 2003; and CAPS (Ca 2ϩ -dependent activator protein for secretion) (Walent et al, 1992) are critical for Ca 2ϩ -dependent exocytosis, they also found that these proteins are dispensable for GTP-dependent exocytosis (Klenchin et al, 1998).…”

Section: Discussionmentioning

confidence: 99%

RalA and RalB Function as the Critical GTP Sensors for GTP-Dependent Exocytosis

Li¹,

Han²,

Chou³

et al. 2007

J. Neurosci.

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Although it has been established that the activation of GTPases by non-hydrolyzable GTP stimulates neurotransmitter release from many different secretory cell types, the underlying mechanisms remain unclear. In the present study we aimed to elucidate the functional role(s) for endogenous Ras-like protein A (RalA) and RalB GTPases in GTP-dependent exocytosis. For this purpose stable neuroendocrine pheochromocytoma 12 (PC12) cell lines were generated in which the expressions of both RalA and RalB were strongly downregulated. In these double knock-down cells GTP-dependent exocytosis was reduced severely and was restored after the expression of RalA or RalB was reintroduced by transfection. In contrast, Ca 2ϩ -dependent exocytosis and the docking of dense core vesicles analyzed by electron microscopy remained unchanged in the double knock-down cells. Furthermore, the transfected RalA and RalB appeared to be localized primarily on the dense core vesicles in undifferentiated and nerve growth factor-differentiated PC12 cells. Our results indicate that endogenous RalA and RalB function specifically as GTP sensors for the GTP-dependent exocytosis of dense core vesicles, but they are not required for the general secretory pathways, including tethering of vesicles to the plasma membrane and Ca 2ϩ -dependent exocytosis.

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“…The same SNAREs are involved in the exocytosis of chromaffin granules (Glenn and Burgoyne, 1996) and secretory granules in PC12 cells (Banerjee et al, 1996). Furthermore, it is known that ϳ 20% of both syntaxin 1 and SNAP-25 are present on vesicles, although the majority are localized to the plasma membrane (Tagaya et al, 1995;Walch-Solinema et al, 1995;Gaisano et al, 1996).…”

Section: Discussionmentioning

confidence: 99%

“…The same SNAREs also have been found to be involved in exocytosis of chromaffin granules (Glenn and Burgoyne, 1996) and secretory granules in PC12 cells (Banerjee et al, 1996). Because secretory granules derive from ISGs, we asked whether any of the SNAREs involved in exocytosis are also involved in ISG-ISG fusion.…”

Section: Neurotoxin Cleavage Of Syntaxin 1 and Vamp2 Does Not Inhibitmentioning

confidence: 99%

Homotypic Fusion of Immature Secretory Granules During Maturation Requires Syntaxin 6

Wendler

Page

Urbé

et al. 2001

MBoC

122

121

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Homotypic fusion of immature secretory granules (ISGs) gives rise to mature secretory granules (MSGs), the storage compartment in endocrine and neuroendocrine cells for hormones and neuropeptides. With the use of a cell-free fusion assay, we investigated which soluble Nethylmaleimide-sensitive fusion protein attachment receptor (SNARE) molecules are involved in the homotypic fusion of ISGs. Interestingly, the SNARE molecules mediating the exocytosis of MSGs in neuroendocrine cells, syntaxin 1, SNAP-25, and VAMP2, were not involved in homotypic ISG fusion. Instead, we have identified syntaxin 6 as a component of the core machinery responsible for homotypic ISG fusion. Subcellular fractionation studies and indirect immunofluorescence microscopy show that syntaxin 6 is sorted away during the maturation of ISGs to MSGs. Although, syntaxin 6 on ISG membranes is associated with SNAP-25 and SNAP-29/GS32, we could not find evidence that these target (t)-SNARE molecules are involved in homotypic ISG fusion. Nor could we find any involvement for the vesicle (v)-SNARE VAMP4, which is known to be associated with syntaxin 6. Importantly, we have shown that homotypic fusion requires the function of syntaxin 6 on both donor as well as acceptor membranes, which suggests that t-t-SNARE interactions, either direct or indirect, may be required during fusion of ISG membranes. INTRODUCTIONCellular organization requires accurate protein transport throughout the entire secretory pathway. Key requirements for protein transport are vesicular carriers with a full complement of machinery to enable them to find and fuse with the correct downstream compartment. This machinery includes the soluble N-ethylmaleimide-sensitive fusion protein (NSF) attachment protein (SNAP) receptors (SNAREs). The SNAREs and SNAPs together with NSF are the core components of the highly conserved machinery involved in all docking and fusion steps in membrane traffic pathways so far described (Robinson and Martin, 1998;Jahn and Sudhof, 1999;Mayer, 1999). There are two classes of SNAREs, vesicle (v)-SNAREs and target (t)-SNAREs, which are defined according to their localization on vesicles or target membranes, respectively, although t-SNAREs have also been detected on vesicles (Tagaya et al., 1995;Walch-Solinema et al., 1995;Gaisano et al., 1996). Typically two tSNAREs and one v-SNARE build a 7S complex composed of a bundle of 4 ␣-helices (Sutton et al., 1998). SNAREs have been shown to be the minimal machinery needed to drive the fusion of lipid bilayers (Weber et al., 1998) and to provide an inherent level of specificity (McNew et al., 2000, summarized in Clague andHerrmann, 2000).Although the majority of membrane fusion events are heterotypic, i.e., between membranes from, or derived from, different intracellular compartments, there are several membrane fusion events, which are homotypic. Two well-described homotypic fusion events occur after cell division when cells exit mitosis, whereupon both the Golgi complex and the endoplasmic reticulum reassemble in the d...

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Botulinum neurotoxin light chains inhibit both Ca²⁺‐induced and GTP analogue‐induced catecholamine release from permeabilised adrenal chromaffin cells

Cited by 32 publications

References 51 publications

Regulated secretion: SNARE density, vesicle fusion and calcium dependence

Regulated secretion: SNARE density, vesicle fusion and calcium dependence

RalA and RalB Function as the Critical GTP Sensors for GTP-Dependent Exocytosis

Homotypic Fusion of Immature Secretory Granules During Maturation Requires Syntaxin 6

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Botulinum neurotoxin light chains inhibit both Ca2+‐induced and GTP analogue‐induced catecholamine release from permeabilised adrenal chromaffin cells

Cited by 32 publications

References 51 publications

Regulated secretion: SNARE density, vesicle fusion and calcium dependence

Regulated secretion: SNARE density, vesicle fusion and calcium dependence

RalA and RalB Function as the Critical GTP Sensors for GTP-Dependent Exocytosis

Homotypic Fusion of Immature Secretory Granules During Maturation Requires Syntaxin 6

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Botulinum neurotoxin light chains inhibit both Ca²⁺‐induced and GTP analogue‐induced catecholamine release from permeabilised adrenal chromaffin cells