scholarly journals Differential regulation of synchronous versus asynchronous neurotransmitter release by the C2 domains of synaptotagmin 1

2010 ◽  
Vol 107 (33) ◽  
pp. 14869-14874 ◽  
Author(s):  
M. Yoshihara ◽  
Z. Guan ◽  
J. T. Littleton
Keyword(s):  
Neurotransmitter Release ◽  
Differential Regulation ◽  
C2 Domains ◽  
Synaptotagmin 1

scholarly journals Synaptotagmin-1 C2B domain interacts simultaneously with SNAREs and membranes to promote membrane fusion

eLife ◽  
2016 ◽  
Vol 5 ◽  
Author(s):  
Shen Wang ◽  
Yun Li ◽  
Cong Ma
Keyword(s):  
Membrane Fusion ◽  
Neurotransmitter Release ◽  
Functional Significance ◽  
Underlying Mechanism ◽  
C2 Domains ◽  
Membrane Deformation ◽  
Synaptotagmin 1 ◽  
Bottom Face ◽  
Liposome Fusion

Synaptotagmin-1 (Syt1) acts as a Ca2+ sensor for neurotransmitter release through its C2 domains. It has been proposed that Syt1 promotes SNARE-dependent fusion mainly through its C2B domain, but the underlying mechanism is poorly understood. In this study, we show that the C2B domain interacts simultaneously with acidic membranes and SNARE complexes via the top Ca2+-binding loops, the side polybasic patch, and the bottom face in response to Ca2+. Disruption of the simultaneous interactions completely abrogates the triggering activity of the C2B domain in liposome fusion. We hypothesize that the simultaneous interactions endow the C2B domain with an ability to deform local membranes, and this membrane-deformation activity might underlie the functional significance of the Syt1 C2B domain in vivo.

scholarly journals Polybasic patches in both C2 domains of Synaptotagmin-1 are required for evoked neurotransmitter release

2021 ◽  
Author(s):  
Zhenyong Wu ◽  
Lu Ma ◽  
Nicholas A Courtney ◽  
Jie Zhu ◽  
Yongli Zhang ◽  
...  
Keyword(s):  
Optical Tweezers ◽  
Single Molecule ◽  
Calcium Binding ◽  
Neurotransmitter Release ◽  
Transmembrane Domain ◽  
Snare Complex ◽  
Critical Feature ◽  
C2 Domains ◽  
Electrophysiological Recordings ◽  
Synaptotagmin 1

Synaptotagmin-1 (Syt1) is a vesicular calcium sensor required for synchronous neurotransmitter release. It is composed of a single-pass transmembrane domain linked to two tandem C2 domains (C2A and C2B) that bind calcium, acidic lipids, and SNARE proteins that drive fusion of the synaptic vesicle with the plasma membrane. Despite its essential role, how Syt1 couples calcium entry to synchronous release is not well understood. Calcium binding to C2B, but not to C2A, is critical for synchronous release and C2B additionally binds the SNARE complex. The C2A domain is also required for Syt1 function, but it is not clear why. Here we asked what critical feature of C2A may be responsible for its functional role, and compared this to the analogous feature in C2B. We focused on highly conserved poly-lysine patches located on the sides of C2A (K189-192) and C2B (K324-327). We tested effects of charge-neutralization mutations in either region (Syt1K189-192A and Syt1K326-327A) side-by-side to determine their relative contributions to Syt1 function in cultured cortical mouse neurons and in single-molecule experiments. Combining electrophysiological recordings and optical tweezers measurements to probe dynamic single C2 domain-membrane interactions, we show that both C2A and C2B polybasic patches contribute to membrane binding, and both are required for evoked release. The readily releasable vesicle pool or spontaneous release were not affected, so both patches are specifically required for synchronization of release. We suggest these patches contribute to cooperative binding to membranes, increasing the overall affinity of Syt1 for negatively charged membranes and facilitating evoked release.

scholarly journals Phosphatidylinositol 4,5 bisphosphate controls the cis and trans interactions of synaptotagmin 1

2019 ◽  
Author(s):  
S.B. Nyenhuis ◽  
A. Thapa ◽  
D. S. Cafiso
Keyword(s):  
Neurotransmitter Release ◽  
Plasma Membranes ◽  
Full Length ◽  
Membrane Insertion ◽  
Membrane Interface ◽  
Vesicle Membrane ◽  
C2 Domains ◽  
Synaptotagmin 1 ◽  
Site Directed Spin Labeling ◽  
Cis And Trans

AbstractSynaptotagmin 1 acts as the Ca2+-sensor for synchronous neurotransmitter release; however, the mechanism by which it functions is not understood and is presently a topic of considerable interest. Here we describe measurements on full-length membrane reconstituted synaptotagmin 1 using site-directed spin labeling where we characterize the linker region as well as the cis (vesicle membrane) and trans (cytoplasmic membrane) binding of its two C2 domains. In the full-length protein, the C2A domain does not undergo membrane insertion in the absence of Ca2+; however, the C2B domain will bind to and penetrate in trans to a membrane containing phosphatidylinositol 4,5 bisphosphate (PIP2), even if phosphatidylserine (PS) is present in the cis membrane. In the presence of Ca2+, the Ca2+-binding loops of C2A and C2B both insert into the membrane interface; moreover, C2A preferentially inserts into PS containing bilayers and will bind in a cis configuration to membranes containing PS even if a PIP2 membrane is presented in trans. The data are consistent with a bridging activity for Syt1 where the two domains bind to opposing vesicle and plasma membranes. The failure of C2A to bind membranes in the absence of Ca2+ and the long unstructured segment linking C2A to the vesicle membrane indicates that synaptotagmin 1 could act to significantly shorten the vesicle-plasma membrane distance with increasing levels of Ca2+.

scholarly journals A novel dual Ca2+ sensor system regulates Ca2+-dependent neurotransmitter release

2021 ◽  
Vol 220 (4) ◽  
Author(s):  
Lei Li ◽  
Haowen Liu ◽  
Mia Krout ◽  
Janet E. Richmond ◽  
Yu Wang ◽  
...  
Keyword(s):  
Neurotransmitter Release ◽  
Release Kinetics ◽  
Sensor System ◽  
C2 Domains ◽  
Loosely Coupled ◽  
C Elegans ◽  
Membrane Tethering ◽  
Synaptotagmin 1 ◽  
Tm Domain ◽  
Fast Release

Ca2+-dependent neurotransmitter release requires synaptotagmins as Ca2+ sensors to trigger synaptic vesicle (SV) exocytosis via binding of their tandem C2 domains—C2A and C2B—to Ca2+. We have previously demonstrated that SNT-1, a mouse synaptotagmin-1 (Syt1) homologue, functions as the fast Ca2+ sensor in Caenorhabditis elegans. Here, we report a new Ca2+ sensor, SNT-3, which triggers delayed Ca2+-dependent neurotransmitter release. snt-1;snt-3 double mutants abolish evoked synaptic transmission, demonstrating that C. elegans NMJs use a dual Ca2+ sensor system. SNT-3 possesses canonical aspartate residues in both C2 domains, but lacks an N-terminal transmembrane (TM) domain. Biochemical evidence demonstrates that SNT-3 binds both Ca2+ and the plasma membrane. Functional analysis shows that SNT-3 is activated when SNT-1 function is impaired, triggering SV release that is loosely coupled to Ca2+ entry. Compared with SNT-1, which is tethered to SVs, SNT-3 is not associated with SV. Eliminating the SV tethering of SNT-1 by removing the TM domain or the whole N terminus rescues fast release kinetics, demonstrating that cytoplasmic SNT-1 is still functional and triggers fast neurotransmitter release, but also exhibits decreased evoked amplitude and release probability. These results suggest that the fast and slow properties of SV release are determined by the intrinsically different C2 domains in SNT-1 and SNT-3, rather than their N-termini–mediated membrane tethering. Our findings therefore reveal a novel dual Ca2+ sensor system in C. elegans and provide significant insights into Ca2+-regulated exocytosis.

scholarly journals Functional synergy between the Munc13 C-terminal C1 and C2 domains

eLife ◽  
2016 ◽  
Vol 5 ◽  
Author(s):  
Xiaoxia Liu ◽  
Alpay Burak Seven ◽  
Marcial Camacho ◽  
Victoria Esser ◽  
Junjie Xu ◽  
...  
Keyword(s):  
Neurotransmitter Release ◽  
Plasma Membranes ◽  
Snare Complex ◽  
Complex Assembly ◽  
C2 Domains ◽  
Vesicle Docking ◽  
Synaptotagmin 1 ◽  
Primed State ◽  
Multiple Domains ◽  
Stimulation Of

Neurotransmitter release requires SNARE complexes to bring membranes together, NSF-SNAPs to recycle the SNAREs, Munc18-1 and Munc13s to orchestrate SNARE complex assembly, and Synaptotagmin-1 to trigger fast Ca2+-dependent membrane fusion. However, it is unclear whether Munc13s function upstream and/or downstream of SNARE complex assembly, and how the actions of their multiple domains are integrated. Reconstitution, liposome-clustering and electrophysiological experiments now reveal a functional synergy between the C1, C2B and C2C domains of Munc13-1, indicating that these domains help bridging the vesicle and plasma membranes to facilitate stimulation of SNARE complex assembly by the Munc13-1 MUN domain. Our reconstitution data also suggest that Munc18-1, Munc13-1, NSF, αSNAP and the SNAREs are critical to form a ‘primed’ state that does not fuse but is ready for fast fusion upon Ca2+ influx. Overall, our results support a model whereby the multiple domains of Munc13s cooperate to coordinate synaptic vesicle docking, priming and fusion.

scholarly journals Synaptotagmin-1 membrane binding is driven by the C2B domain and assisted cooperatively by the C2A domain

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Clémence Gruget ◽  
Oscar Bello ◽  
Jeff Coleman ◽  
Shyam S. Krishnakumar ◽  
Eric Perez ◽  
...  
Keyword(s):  
Neurotransmitter Release ◽  
Mutational Analysis ◽  
Membrane Binding ◽  
Surface Force ◽  
Membrane Interaction ◽  
C2 Domains ◽  
Binding Energetics ◽  
Synaptotagmin 1 ◽  
Site Binding ◽  
C2a Domain

Abstract Synaptotagmin interaction with anionic lipid (phosphatidylserine/phosphatidylinositol) containing membranes, both in the absence and presence of calcium ions (Ca2+), is critical to its central role in orchestrating neurotransmitter release. The molecular surfaces involved, namely the conserved polylysine motif in the C2B domain and Ca2+-binding aliphatic loops on both C2A and C2B domains, are known. Here we use surface force apparatus combined with systematic mutational analysis of the functional surfaces to directly measure Syt1-membrane interaction and fully map the site-binding energetics of Syt1 both in the absence and presence of Ca2+. By correlating energetics data with the molecular rearrangements measured during confinement, we find that both C2 domains cooperate in membrane binding, with the C2B domain functioning as the main energetic driver, and the C2A domain acting as a facilitator.

scholarly journals Phosphatidylinositol Phosphates as Co-activators of Ca2+ Binding to C2 Domains of Synaptotagmin 1

2006 ◽  
Vol 281 (23) ◽  
pp. 15845-15852 ◽  
Author(s):  
LiYi Li ◽  
Ok-Ho Shin ◽  
Jeong-Seop Rhee ◽  
Demet Araç ◽  
Jong-Cheol Rah ◽  
...  
Keyword(s):  
C2 Domains ◽  
Synaptotagmin 1 ◽  
Phosphatidylinositol Phosphates

scholarly journals The structure and flexibility analysis of the Arabidopsis Synaptotagmin 1 reveal the basis of its regulation at membrane contact sites

2021 ◽  
Author(s):  
Juan Luis Benavente ◽  
Dritan Siliqi ◽  
Lourdes Infantes ◽  
Laura Lagartera ◽  
Alberto Mills ◽  
...  
Keyword(s):  
Cell Function ◽  
Lipid Extraction ◽  
Membrane Lipid ◽  
Lipid Binding ◽  
Lipid Transfer ◽  
C2 Domains ◽  
Membrane Contact Sites ◽  
Contact Sites ◽  
Cellular Environment ◽  
Synaptotagmin 1

Cell function requires the maintenance of membrane lipid homeostasis as changes in cellular environment unbalance this equilibrium. The non-vesicular lipid transfer at endoplasmic reticulum (ER) and plasma membrane (PM) contact sites (CS) is central to restore it. Extended synaptotagmins (E-Syts) are ER proteins that play a central role in this process as they act as molecular tethers with PM and as lipid transfer proteins between these organelles. E-Syts are constitutively anchored to the ER through an N-terminal hydrophobic segment and bind to the PM via C-terminal C2 domains. In plants, synaptotagmins (SYTs) are orthologous of E-Syts and regulate the ER-PM communication by the activity of their two C2 domains in response to abiotic stresses. We have combined macromolecular crystallography, small-angle X-ray scattering, structural bioinformatics and biochemical data to analyze the regulation of plant synaptotagmin 1 (SYT1). Our data show that the binding of SYT1 to the PM is regulated by the interaction of the first C2 domain through a Ca2+-dependent lipid binding site and by a site for phosphorylated forms of phosphatidylinositol in such a way that two different molecular signals are integrated in response to stress. In addition, our data show that SYT1 is highly flexible by virtue of up to three hinge points, including one that connects the two C2 domains. This feature provides conformational freedom to SYT1 to define a large and complementary interaction surface with the PM. This structural plasticity, in turn, may facilitate lipid extraction, protein loading and subsequent transfer between PM and ER.

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