PtdInsP2 and PtdSer cooperate to trap synaptotagmin-1 to the plasma membrane in the presence of calcium

Pérez-Lara, Ángel; Thapa, Anusa; Nyenhuis, Sarah B.; Nyenhuis, David A.; Halder, Partho; Tietzel, Michael; Tittmann, Kai; Cafiso, David S.; Jahn, Reinhard

doi:10.7554/elife.15886

Cited by 102 publications

(186 citation statements)

References 71 publications

Supporting

Mentioning

159

Contrasting

Order By: Relevance

“…This promotes specific PI(4,5)P 2 interactions and penetration of adjacent hydrophobic residues, some of which are exposed as a result of Ca 2+ activation, into the lipid bilayer 1,58 . The MD simulation suggests that charge neutralization in the Ca 2+ -binding site of the C2B domain by Ca 2+ reduces the electrostatic repulsion of the domain from the PM and promotes association with PI(4,5)P 2 , as recently described for syt1 27 and the constitutive PM localization of DOC2B D218,220N . We thus conclude that both Ca 2+ and PI(4,5)P 2 are required for DOC2B translocation in living cells.…”

Section: Discussionsupporting

confidence: 61%

“…Indeed, PI(4,5)P 2 was found to be crucial for Ca 2+ -dependent translocation of the PKC C2 domain 23,24 . In addition, specific interactions of PI(4,5)P 2 with C2 domains of synaptotagmin (syt) 1 and rabphilin 3A, which present the highest homology to DOC2B C2 domains, have been suggested by nuclear magnetic resonance and in vitro assays 25-27 . Other biochemical studies have established that PI(4,5)P 2 enhances the Ca 2+ -dependent and independent interactions of syt1 and DOC2B C2 domains with liposomes 1,25 .…”

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Phosphatidylinositol (4, 5)‐bisphosphate targets double C2 domain protein B to the plasma membrane

Michaeli

Gottfried

Bykhovskaia

et al. 2017

Traffic

View full text Add to dashboard Cite

DOC2B is a high-affinity Ca2+ sensor that translocates from the cytosol to the plasma membrane (PM) and promotes vesicle priming and fusion. However, the molecular mechanism underlying its translocation and targeting to the PM in living cells is not completely understood. DOC2B interacts in vitro with the PM components phosphatidylserine, phosphatidylinositol (4,5)-bisphosphate [PI(4,5)P2] and target SNAREs (t-SNAREs). Here we show that PI(4,5)P2 hydrolysis at the PM of living cells abolishes DOC2B translocation, whereas manipulations of t-SNAREs and other phosphoinositides have no effect. Moreover, we were able to redirect DOC2B to intracellular membranes by synthesizing PI(4,5)P2 in those membranes. Molecular dynamics simulations and mutagenesis in the calcium and PI(4,5)P2-binding sites strengthened our findings, demonstrating that both calcium and PI(4,5)P2 are required for the DOC2B–PM association and revealing multiple PI(4,5)P2–C2B interactions. In addition, we show that DOC2B translocation to the PM is ATP-independent, and occurs in a diffusion-like manner. Our data suggest that the Ca2+-triggered translocation of DOC2B is diffusion-driven and aimed at PI(4,5)P2-containing membranes.

show abstract

Section: Discussionsupporting

confidence: 61%

Section: Introductionmentioning

confidence: 99%

Phosphatidylinositol (4, 5)‐bisphosphate targets double C2 domain protein B to the plasma membrane

Michaeli

Gottfried

Bykhovskaia

et al. 2017

Traffic

View full text Add to dashboard Cite

show abstract

“…The Rph3A C2B-PIP 2 -Ca 2+ structures obtained here confirmed the prediction, providing solid evidences that support the existence of a polybasic conserved region located in the β3-β4 strands of some C2 domains (12,(24)(25)(26) (Fig. S5).…”

Section: Resultssupporting

confidence: 71%

“…This membrane-bending mechanism driven by Ca 2+ has been described for other PIP 2 -interacting C2 domain-bearing proteins that also participate in vesicle fusion, like synaptotagmins (28) and DOC2B (20). In fact, the preferred use of the Syt1 C2B polybasic region for PIP 2 binding has been recently demonstrated by different biophysical techniques (25,26).…”

Section: Resultsmentioning

confidence: 99%

Structural characterization of the Rabphilin-3A–SNAP25 interaction

Ferrer-Orta

Perez-Sanchez

Coronado-Parra

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

Membrane fusion is essential in a myriad of eukaryotic cell biological processes, including the synaptic transmission. Rabphilin-3A is a membrane trafficking protein involved in the calcium-dependent regulation of secretory vesicle exocytosis in neurons and neuroendocrine cells, but the underlying mechanism remains poorly understood. Here, we report the crystal structures and biochemical analyses of Rabphilin-3A C2B-SNAP25 and C2B-phosphatidylinositol 4,5-bisphosphate (PIP 2 ) complexes, revealing how Rabphilin-3A C2 domains operate in cooperation with PIP 2 /Ca 2+ and SNAP25 to bind the plasma membrane, adopting a conformation compatible to interact with the complete SNARE complex. Comparisons with the synaptotagmin1-SNARE show that both proteins contact the same SNAP25 surface, but Rabphilin-3A uses a unique structural element. Data obtained here suggest a model to explain the Ca 2+ -dependent fusion process by membrane bending with a myriad of variations depending on the properties of the C2 domain-bearing protein, shedding light to understand the fine-tuning control of the different vesicle fusion events.C2 domains | membrane fusion | Rabphilin-3A | SNAP-25 | X-ray crystallography N eurons are able to communicate with others through the liberation of neurotransmitters during synapses. Numerous proteins are recruited to the presynaptic space to execute a highly controlled and precise mechanism, resulting in the liberation of neurotransmitters to the synaptic cleft. Central components of the process are the soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins: syntaxin-1A (STX1A), VAMP2 (synaptobrevin-2), and synaptosome-associated protein of 25 kDa (SNAP25), which fold into a stable four-helix bundle that brings together vesicle and plasma membranes, driving membrane fusion (1-4). Other important players are complexin, Munc18-1, Munc13, and a collection of proteins that share a common structural motif: the C2 domain (5, 6). These domains are regulated by their ability to bind Ca 2+ , phospholipids, and protein interactions, endowing them with properties to fine-tune the wide variety of vesicle release modes (7). Nowadays, we have detailed information on many of the steps contributing to docking, priming, and fusion of these vesicles, but a clear picture on how these regulators contribute in each particular state still remains under debate, mainly due to the lack of high-resolution structural data.Rabphilin-3A (Rph3A) is a membrane trafficking protein involved in the Ca 2+ -dependent regulation of secretory vesicle exocytosis in neurons and neuroendocrine cells; however, its exact role in the process still remains under debate. The protein is targeted to synaptic or secretory vesicles through the interaction with the small G-proteins Rab3 or Rab27, via its N-terminal Rabbinding domain (8-10). Its C-terminal domain consists of tandem C2 domains that are responsible for the Ca 2+ -and phospholipidsdependent membrane specificity of the protein (11, 12) and also participate in oth...

show abstract

“…2a–d, Supplementary Video 2). We also tested potential interactions that involve the polybasic region of Syt1 C2B by mutating two Lys residues (K326A/K327A, referred to as KA mutant) in order to disrupt any dynamic binding modes involving the highly charged polybasic region 20,22 . All mutants of the Syt1 C2B domain are properly folded (Fig.…”

Section: Specific Mutations Disrupt the Tripartite Interfacementioning

confidence: 99%

The primed SNARE–complexin–synaptotagmin complex for neuronal exocytosis

Zhou

Wang

et al. 2017

Nature

270

501

View full text Add to dashboard Cite

Summary Synaptotagmin, complexin and neuronal SNARE proteins mediate evoked synchronous neurotransmitter release, but the molecular mechanisms mediating the cooperation between these molecules remain unclear. Here, we determined crystal structures of the primed pre-fusion SNARE-complexin-synaptotagmin-1 complex. These structures reveal an unexpected tripartite interface between synaptotagmin-1 and both the SNARE complex and complexin. Simultaneously, a second synaptotagmin-1 molecule interacted with the other side of the SNARE complex via the previously identified primary interface. Mutations that disrupt either interface in solution also severely impaired evoked synchronous release in neurons, suggesting that both interfaces are essential for the primed pre-fusion state. Ca2+ binding to the synaptotagmin-1 molecules unlocks the complex, allows full zippering of the SNARE complex, and triggers membrane fusion. The tripartite SNARE-complexin-synaptotagmin-1 complex at a synaptic vesicle docking site has to be unlocked for triggered fusion to commence, explaining the cooperation between complexin and synaptotagmin-1 in synchronizing evoked release on the sub-millisecond timescale.

show abstract

PtdInsP2 and PtdSer cooperate to trap synaptotagmin-1 to the plasma membrane in the presence of calcium

Cited by 102 publications

References 71 publications

Phosphatidylinositol (4, 5)‐bisphosphate targets double C2 domain protein B to the plasma membrane

Phosphatidylinositol (4, 5)‐bisphosphate targets double C2 domain protein B to the plasma membrane

Structural characterization of the Rabphilin-3A–SNAP25 interaction

The primed SNARE–complexin–synaptotagmin complex for neuronal exocytosis

Contact Info

Product

Resources

About