Faculty Opinions recommendation of Photolysis of a caged peptide reveals rapid action of N-ethylmaleimide sensitive factor before neurotransmitter release.

Noncanonical secretagogues such as hypertonicity or α-latrotoxin have been extremely informative in studying neurotransmission. Lanthanum and lanthanides can also trigger neurotransmitter release through an unknown mechanism. Here, we studied the effect of lanthanides on neurotransmission in hippocampal cultures. Application of 2 mM La3+ caused rapid and robust neurotransmitter release within seconds. In addition, transient application of La3+ uncovered a sustained facilitation of miniature neurotransmission. The response to La3+ was detectable at 2 μM and increased in a concentration-dependent manner ≤2 mM. Rapid effect of La3+ was independent of extracellular and intracellular Ca2+ and did not require La3+ entry into cells or activation of phospholipaseCβ. Synapses deficient in synaptobrevin-2, the major synaptic vesicle soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) protein in the brain, did not display any rapid release in response to La3+, whereas the slow facilitation of release detected after La3+ removal remained intact. In contrast, preincubation with intracellular Ca2+ chelators selectively attenuated the delayed release triggered by La3+. Moreover, synapses deficient in synaptotagmin-1 maintained a rapid response to La3+, suggesting that La3+-triggered neurotransmitter release does not require synaptotagmin-1 as a sensor. Therefore La3+ has two separate effects on synaptic transmission. For its rapid action, La3+ interacts with a target on the surface membrane, and unlike other forms of release, it triggers strictly synaptobrevin-2–dependent fusion, implying that in central synapses synaptobrevin-2 function is secretagogue specific. For the delayed action, La3+ may act intracellularly after its entry or through intracellular Ca2+ via a mechanism that does not require synaptobrevin-2.

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Role of α-SNAP in Promoting Efficient Neurotransmission at the Crayfish Neuromuscular Junction

Journal of Neurophysiology ◽

10.1152/jn.1999.82.6.3406 ◽

1999 ◽

Vol 82 (6) ◽

pp. 3406-3416 ◽

Cited By ~ 34

Author(s):

Ping He ◽

R. Chase Southard ◽

Dong Chen ◽

S. W. Whiteheart ◽

R. L. Cooper

Keyword(s):

Neuromuscular Junction ◽

Neurotransmitter Release ◽

Synaptic Vesicles ◽

Synaptic Current ◽

Attachment Protein ◽

Quantal Analysis ◽

Sensitive Factor ◽

Electrophysiological Studies ◽

Crayfish Neuromuscular Junction

In this manuscript, we address the role of the soluble N-ethylmaleimide sensitive factor attachment protein (α-SNAP) in synaptic transmission at the neuromuscular junction of the crayfish opener muscle. Immunochemcial methods confirm the presence of α-SNAP in these preparations and show that it is concentrated in the synaptic areas. Microinjection and electrophysiological studies show that α-SNAP causes an increase in neurotransmitter release yet does not significantly affect the kinetics. More specific quantal analysis, using focal, macropatch, synaptic current recordings, shows that α-SNAP increases transmitter release by increasing the probability of exocytosis but not the number of potential release sites. These data demonstrate that the role of α-SNAP is to increase the efficiency of neurotransmission by increasing the probability that a stimulus will result in neurotransmitter release. What this suggests is that α-SNAP is critical for the formation and maintenance of a “ready release” pool of synaptic vesicles.

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Complexin splits the membrane-proximal region of a single SNAREpin

Biochemical Journal ◽

10.1042/bcj20160339 ◽

2016 ◽

Vol 473 (14) ◽

pp. 2219-2224 ◽

Cited By ~ 10

Author(s):

Linxiang Yin ◽

Jaewook Kim ◽

Yeon-Kyun Shin

Keyword(s):

Neurotransmitter Release ◽

Proximal Part ◽

Proximal Region ◽

Snare Complex ◽

Spontaneous Release ◽

Attachment Protein ◽

Protein Receptor ◽

Sensitive Factor ◽

Membrane Proximal Region ◽

Factor Attachment Protein Receptor

Tight regulation of neurotransmitter release by Ca2+ is critical in neurons, which requires suppression of spontaneous release. In the present study, we find that the complexin (Cpx) protein binds to the soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) complex to split the membrane-proximal part, whereby it inhibits spontaneous release.

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Complexin-1 regulated transition in the assembly of single neuronal SNARE complex

10.1101/2021.06.07.447331 ◽

2021 ◽

Author(s):

Hao Tongrui ◽

Feng Nan ◽

Gong Fan ◽

Liu Jiaquan ◽

Lu Ma ◽

...

Keyword(s):

Synaptic Vesicle ◽

Molecular Mechanism ◽

Optical Tweezers ◽

Neurotransmitter Release ◽

Snare Complex ◽

Snare Proteins ◽

Synaptic Vesicle Exocytosis ◽

Sensitive Factor ◽

Vesicle Exocytosis ◽

Vesicle Pool

Neurotransmitter release is mediated by the synaptic vesicle exocytosis. Important proteins in this process have been identified including the molecular machine Synaptic-soluble N-ethylmaleimide-sensitive factor attachment receptor (SNARE) proteins, and other regulators. Complexin (Cpx) is one of the vital regulators in this process. The functions of Cpx are proposed to maintain a proper primed vesicle pool by preventing its premature depletion, which facilitates the vesicle fusion in the presence of Ca2+. However, the molecular mechanism remains unclear. Using dual-trap optical tweezers, we detected the interaction of complexin-1 (CpxI) with SNARE. We found that the CpxI stabilizes partially folded SNARE complexes by competing with C-terminal of Vamp protein and interacting with the C-terminal of t-SNARE complex.

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Characterization of Ginkgo biloba Leaf Flavonoids as Neuroexocytosis Regulators

Molecules ◽

10.3390/molecules25081829 ◽

2020 ◽

Vol 25 (8) ◽

pp. 1829

Author(s):

Choongjin Ban ◽

Joon-Bum Park ◽

Sora Cho ◽

Hye Rin Kim ◽

Yong Joon Kim ◽

...

Keyword(s):

Ginkgo Biloba ◽

Neurotransmitter Release ◽

Attachment Protein ◽

Potential Source ◽

Protein Receptor ◽

Physicochemical Stability ◽

Sensitive Factor ◽

Low Toxicity ◽

Ginkgo Biloba Leaf

Ginkgo biloba leaf (GBL) is known as a potential source of bioactive flavonoids, such as quercetin, arresting the neuronal soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE)-zippering. Here, the GBL flavonoids were isolated in two different manners and then examined for their bioactivity, physicochemical stability, and biocompatibility. The majority of flavonoids in the non-hydrolyzed and acidolyzed isolates, termed non-hydrolyzed isolate (NI) and acidolyzed isolate (AI) hereafter, were rich in flavonol glycosides and aglycones, respectively. Glycosidic/aglyconic quercetin and kaempferol were abundant in both NI and AI, whereas a little of apigenin, luteolin, and isorhamnetin were found in AI. NI was more thermostable in all pH ranges than quercetin, kaempferol, and AI. NI and AI both inhibited neurotransmitter release from differentiated neuronal PC-12 cells. NI and AI showed 1/2–1/3 lower EC50/CC50 values than quercetin and kaempferol. The NI and AI exhibited no toxicity assessed by the tests on chorioallantoic membranes of hen’s eggs, removing toxicological concerns of irritation potential. Moreover, GBL isolates, particularly AI, showed antioxidant and anti-inflammatory activities in the use below the CC50 levels. Taken together, these results suggest that GBL isolates that are rich in antioxidant flavonoids are effective anti-neuroexocytotic agents with high stability and low toxicity.

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Phosphorylation of theN-Ethylmaleimide-sensitive Factor Is Associated with Depolarization-dependent Neurotransmitter Release from Synaptosomes

Journal of Biological Chemistry ◽

10.1074/jbc.m007394200 ◽

2001 ◽

Vol 276 (15) ◽

pp. 12174-12181 ◽

Cited By ~ 46

Author(s):

Elena A. Matveeva ◽

Sidney W. Whiteheart ◽

Thomas C. Vanaman ◽

John T. Slevin

Keyword(s):

Protein Function ◽

Neurotransmitter Release ◽

Phorbol Esters ◽

Form Complex ◽

Calcium Dependent ◽

Subunit Interface ◽

Calmodulin Kinase ◽

Sensitive Factor ◽

Molecular Modeling Studies

Critical to SNARE protein function in neurotransmission are the accessory proteins, solubleN-ethylmaleimide-sensitive factor (NSF) attachment protein (SNAP), and NSF, that play a role in activation of the SNAREs for membrane fusion. In this report, we demonstrate the depolarization-induced, calcium-dependent phosphorylation of NSF in rat synaptosomes. Phosphorylation of NSF is coincident with neurotransmitter release and requires an influx of external calcium. Phosphoamino acid analysis of the radiolabeled NSF indicates a role for a serine/threonine-specific kinase. Synaptosomal phosphorylation of NSF is stimulated by phorbol esters and is inhibited by staurosporine, chelerythrine, bisindolylmaleimide I, calphostin C, and Ro31-8220 but not the calmodulin kinase II inhibitor, Kn-93, suggesting a role for protein kinase C (PKC). Indeed, NSF is phosphorylated by PKCin vitroat Ser-237 of the catalytic D1 domain. Mutation of this residue to glutamic acid or to alanine eliminatesin vitrophosphorylation. Molecular modeling studies suggest that Ser-237 is adjacent to an inter-subunit interface at a position where its phosphorylation could affect NSF activity. Consistently, mutation of Ser-237 to Glu, to mimic phosphorylation, results in a hexameric form of NSF that does not bind to SNAP-SNARE complexes, whereas the S237A mutant does form complex. These data suggest a negative regulatory role for PKC phosphorylation of NSF.

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The ubiquitin ligase Ariadne-1 regulates NSF for neurotransmitter release

10.1101/2020.01.23.916619 ◽

2020 ◽

Author(s):

Juanma Ramírez ◽

Miguel Morales ◽

Nerea Osinalde ◽

Imanol Martínez-Padrón ◽

Ugo Mayor ◽

...

Keyword(s):

Ubiquitin Ligase ◽

Neurotransmitter Release ◽

Quantitative Proteomics ◽

Dependent Manner ◽

Loss Of Function ◽

Sensitive Factor ◽

Active Zones ◽

Protein Ensembles ◽

Mutant Phenotypes

ABSTRACTAriadne-1 (Ari-1) is an essential E3 ubiquitin-ligase whose neuronal substrates are yet to be identified. We have used an in vivo ubiquitin biotinylation strategy coupled to quantitative proteomics to identify putative Ari-1 substrates in Drosophila heads. Sixteen candidates met the established criteria. Amongst those, we identified Comatose (Comt), the homologue of the N-ethylmaleimide sensitive factor (NSF). Using an in vivo GFP pulldown approach, we validate Comt/NSF to be an ubiquitination substrate of Ari-1 in fly neurons. The interaction results in the monoubiquitination of Comt/NSF. We also report that Ari-1 loss of function mutants display a lower rate of spontaneous neurotransmitter release due to failures at the pre-synaptic side. By contrast, evoked release in Ari-1 mutants is enhanced in a Ca2+ dependent manner without modifications in the number of active zones, indicating that the probability of release per synapse is increased in these mutants. The distinct Ari-1 mutant phenotypes in spontaneous versus evoked release indicate that NSF activity discriminates the two corresponding protein ensembles that mediate each mode of release. Our results, thus, provide a mechanism to regulate NSF activity in the synapse through Ari-1-dependent ubiquitination.

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