scholarly journals Studying calcium-triggered vesicle fusion in a single vesicle-vesicle content and lipid-mixing system

2012 ◽  
Vol 8 (1) ◽  
pp. 1-16 ◽  
Author(s):  
Minjoung Kyoung ◽  
Yunxiang Zhang ◽  
Jiajie Diao ◽  
Steven Chu ◽  
Axel T Brunger
Keyword(s):  
Vesicle Fusion ◽  
Vesicle Content ◽  
Single Vesicle

scholarly journals A fast, single-vesicle fusion assay mimics physiological SNARE requirements

2010 ◽  
Vol 107 (8) ◽  
pp. 3517-3521 ◽  
Author(s):  
Erdem Karatekin ◽  
Jérôme Di Giovanni ◽  
Cécile Iborra ◽  
Jeff Coleman ◽  
Ben O'Shaughnessy ◽  
...  
Keyword(s):  
Vesicle Fusion ◽  
Single Vesicle

scholarly journals SNARE-Mediated Single-Vesicle Fusion Events with Supported and Freestanding Lipid Membranes

2017 ◽  
Vol 113 (11) ◽  
pp. 2573-2574 ◽  
Author(s):  
Jan W. Kuhlmann ◽  
Meike Junius ◽  
Ulf Diederichsen ◽  
Claudia Steinem
Keyword(s):  
Lipid Membranes ◽  
Vesicle Fusion ◽  
Single Vesicle

scholarly journals Single-Vesicle Fusion Assay Reveals Munc18-1 Binding to the SNARE Core Is Sufficient for Stimulating Membrane Fusion

2010 ◽  
Vol 1 (3) ◽  
pp. 168-174 ◽  
Author(s):  
Jiajie Diao ◽  
Zengliu Su ◽  
Xiaobing Lu ◽  
Tae-Young Yoon ◽  
Yeon-Kyun Shin ◽  
...  
Keyword(s):  
Membrane Fusion ◽  
Vesicle Fusion ◽  
Single Vesicle

scholarly journals High-Throughput Monitoring of Single Vesicle Fusion Using Freestanding Membranes and Automated Analysis

Langmuir ◽  
2018 ◽  
Vol 34 (20) ◽  
pp. 5849-5859 ◽  
Author(s):  
Sathish Ramakrishnan ◽  
Andrea Gohlke ◽  
Feng Li ◽  
Jeff Coleman ◽  
Weiming Xu ◽  
...  
Keyword(s):  
High Throughput ◽  
Automated Analysis ◽  
Vesicle Fusion ◽  
Single Vesicle

High Affinity Host–Guest FRET Pair for Single-Vesicle Content-Mixing Assay: Observation of Flickering Fusion Events

2015 ◽  
Vol 137 (28) ◽  
pp. 8908-8911 ◽  
Author(s):  
Bokyoung Gong ◽  
Bong-Kyu Choi ◽  
Jae-Yeol Kim ◽  
Dinesh Shetty ◽  
Young Ho Ko ◽  
...  
Keyword(s):  
High Affinity ◽  
Vesicle Content ◽  
Single Vesicle ◽  
Fret Pair

scholarly journals A single vesicle-vesicle fusion assay for in vitro studies of SNAREs and accessory proteins

2012 ◽  
Vol 7 (5) ◽  
pp. 921-934 ◽  
Author(s):  
Jiajie Diao ◽  
Yuji Ishitsuka ◽  
Hanki Lee ◽  
Chirlmin Joo ◽  
Zengliu Su ◽  
...  
Keyword(s):  
Vesicle Fusion ◽  
In Vitro Studies ◽  
Accessory Proteins ◽  
Single Vesicle

scholarly journals Complexin inhibits spontaneous release and synchronizes Ca2+-triggered synaptic vesicle fusion by distinct mechanisms

eLife ◽  
2014 ◽  
Vol 3 ◽  
Author(s):  
Ying Lai ◽  
Jiajie Diao ◽  
Daniel J Cipriano ◽  
Yunxiang Zhang ◽  
Richard A Pfuetzner ◽  
...  
Keyword(s):  
Lipid Content ◽  
Point Contact ◽  
Vesicle Fusion ◽  
Neuronal Cultures ◽  
Spontaneous Release ◽  
Terminal Domain ◽  
Synaptotagmin 1 ◽  
Free Membrane ◽  
Synaptic Vesicle Fusion ◽  
Single Vesicle

Previously we showed that fast Ca2+-triggered vesicle fusion with reconstituted neuronal SNAREs and synaptotagmin-1 begins from an initial hemifusion-free membrane point contact, rather than a hemifusion diaphragm, using a single vesicle–vesicle lipid/content mixing assay (<xref ref-type="bibr" rid="bib5">Diao et al., 2012</xref>). When complexin-1 was included, a more pronounced Ca2+-triggered fusion burst was observed, effectively synchronizing the process. Here we show that complexin-1 also reduces spontaneous fusion in the same assay. Moreover, distinct effects of several complexin-1 truncation mutants on spontaneous and Ca2+-triggered fusion closely mimic those observed in neuronal cultures. The very N-terminal domain is essential for synchronization of Ca2+-triggered fusion, but not for suppression of spontaneous fusion, whereas the opposite is true for the C-terminal domain. By systematically varying the complexin-1 concentration, we observed differences in titration behavior for spontaneous and Ca2+-triggered fusion. Taken together, complexin-1 utilizes distinct mechanisms for synchronization of Ca2+-triggered fusion and inhibition of spontaneous fusion.

scholarly journals A single-vesicle content mixing assay for SNARE-mediated membrane fusion

2010 ◽  
Vol 1 (1) ◽  
Author(s):  
Jiajie Diao ◽  
Zengliu Su ◽  
Yuji Ishitsuka ◽  
Bin Lu ◽  
Kyung Suk Lee ◽  
...  
Keyword(s):  
Membrane Fusion ◽  
Vesicle Content ◽  
Single Vesicle

scholarly journals Molecular Determinants of Complexin Clamping in Reconstituted Single-Vesicle Fusion

2021 ◽  
Author(s):  
Manindra Bera ◽  
Sathish Ramakrishnan ◽  
Jeff Coleman ◽  
Shyam S Krishnakumar ◽  
James E Rothman
Keyword(s):  
Specific Interaction ◽  
Vesicle Fusion ◽  
Assembly Process ◽  
Inhibitory Function ◽  
Physiological Relevance ◽  
Vesicular Release ◽  
Molecular Determinants ◽  
Synaptotagmin 1 ◽  
Single Vesicle

Previously we reported that Synaptotagmin-1 and Complexin synergistically clamp the SNARE assembly process to generate and maintain a pool of docked vesicles that fuse rapidly and synchronously upon Ca2+ influx (Ramakrishnan et al. 2020). Here using the same in vitro single-vesicle fusion assay, we establish the molecular details of the Complexin clamp and its physiological relevance. We find that a delay in fusion kinetics, likely imparted by Synaptotagmin-1, is needed for Complexin to block fusion. Systematic truncation/mutational analyses reveal that continuous alpha-helical accessory-central domains of Complexin are essential for its inhibitory function and specific interaction of the accessory helix with the SNAREpins, analogous to the trans clamping model, enhances this functionality. The c-terminal domain promotes clamping by locally elevating Complexin concentration through interactions with the membrane. Further, we find that Complexin likely contributes to rapid Ca2+-synchronized vesicular release by preventing un-initiated fusion rather than by directly facilitating vesicle fusion.

scholarly journals Molecular mechanisms of atlastin-mediated ER membrane fusion revealed by a FRET-based single-vesicle fusion assay

2017 ◽  
Vol 7 (1) ◽  
Author(s):  
Kyung Tae Kim ◽  
Yeojin Moon ◽  
Yunsu Jang ◽  
Kang Taek Lee ◽  
Changwook Lee ◽  
...  
Keyword(s):  
Membrane Fusion ◽  
Molecular Mechanisms ◽  
Vesicle Fusion ◽  
Single Vesicle ◽  
Er Membrane
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