Exocytotic fusion pores are composed of both lipids and proteins

Bao, Huan; Goldschen-Ohm, Marcel P.; Jeggle, Pia; Chanda, Baron; Edwardson, J. Michael; Chapman, Edwin R.

doi:10.1038/nsmb.3141

Cited by 77 publications

(113 citation statements)

References 56 publications

(83 reference statements)

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“…The fusion pore may be composed by both proteins and lipids (Bao et al . ; Sharma and Lindau ). However, since it is suggested any changes in foot amplitude of amperometrical spikes may be related to fusion pore composition (Wang et al .…”

Section: Discussionmentioning

confidence: 96%

Extracellular and intracellular sphingosine‐1‐phosphate distinctly regulates exocytosis in chromaffin cells

Jiang

Delaney

Zanin

et al. 2019

Journal of Neurochemistry

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Sphingosine‐1‐phosphate (S1P) is an essential bioactive sphingosine lipid involved in many neurological disorders. Sphingosine kinase 1 (SphK1), a key enzyme for S1P production, is concentrated in presynaptic terminals. However, the role of S1P/SphK1 signaling in exocytosis remains elusive. By detecting catecholamine release from single vesicles in chromaffin cells, we show that a dominant negative SphK1 (SphK1DN) reduces the number of amperometric spikes and increases the duration of foot, which reflects release through a fusion pore, implying critical roles for S1P in regulating the rate of exocytosis and fusion pore expansion. Similar phenotypes were observed in chromaffin cells obtained from SphK1 knockout mice compared to those from wild‐type mice. In addition, extracellular S1P treatment increased the number of amperometric spikes, and this increase, in turn, was inhibited by a selective S1P3 receptor blocker, suggesting extracellular S1P may regulate the rate of exocytosis via activation of S1P3. Furthermore, intracellular S1P application induced a decrease in foot duration of amperometric spikes in control cells, indicating intracellular S1P may regulate fusion pore expansion during exocytosis. Taken together, our study represents the first demonstration that S1P regulates exocytosis through distinct mechanisms: extracellular S1P may modulate the rate of exocytosis via activation of S1P receptors while intracellular S1P may directly control fusion pore expansion during exocytosis. Open science badges This article has received a badge for *Open Materials* because it provided all relevant information to reproduce the study in the manuscript. The complete Open Science Disclosure form for this article can be found at the end of the article. More information about the Open Practices badges can be found at https://cos.io/our-services/open-science-badges/.

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Section: Discussionmentioning

confidence: 96%

Extracellular and intracellular sphingosine‐1‐phosphate distinctly regulates exocytosis in chromaffin cells

Jiang

Delaney

Zanin

et al. 2019

Journal of Neurochemistry

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“…Most studies concluded that only a few copies of neuronal SNAREs are sufficient for calcium-triggered exocytosis and fusion of small liposomes (Bao et al, 2016; Shi et al, 2012; Sinha et al, 2011; Mohrmann et al, 2010; van den Bogaart et al, 2010; Karatekin et al, 2010; Domanska et al, 2010). Despite this, the average synaptic vesicle carries 70 v-SNARE copies (Takamori et al, 2006) and at least as many t-SNAREs are clustered at plasma membrane docking and fusion sites in neuroendocrine cells (Knowles et al, 2010).…”

Section: Discussionmentioning

confidence: 99%

Dilation of fusion pores by crowding of SNARE proteins

Bello

Thiyagarajan

et al. 2017

eLife

112

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Hormones and neurotransmitters are released through fluctuating exocytotic fusion pores that can flicker open and shut multiple times. Cargo release and vesicle recycling depend on the fate of the pore, which may reseal or dilate irreversibly. Pore nucleation requires zippering between vesicle-associated v-SNAREs and target membrane t-SNAREs, but the mechanisms governing the subsequent pore dilation are not understood. Here, we probed the dilation of single fusion pores using v-SNARE-reconstituted ~23-nm-diameter discoidal nanolipoprotein particles (vNLPs) as fusion partners with cells ectopically expressing cognate, 'flipped' t-SNAREs. Pore nucleation required a minimum of two v-SNAREs per NLP face, and further increases in v-SNARE copy numbers did not affect nucleation rate. By contrast, the probability of pore dilation increased with increasing v-SNARE copies and was far from saturating at 15 v-SNARE copies per face, the NLP capacity. Our experimental and computational results suggest that SNARE availability may be pivotal in determining whether neurotransmitters or hormones are released through a transient ('kiss and run') or an irreversibly dilating pore (full fusion).DOI: http://dx.doi.org/10.7554/eLife.22964.001

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“…(B) NDs reconstituted with neuronal v‐ SNARE s are mixed with small liposomes reconstituted with complementary neuronal t‐ SNARE s. Bulk cargo release is monitored by an increase in the fluorescence of a cargo‐sensitive dye present in the bath. Liposomes were loaded with calcium or glutamate and release was monitored using Mag‐Fluo‐4 or iG luSn FR , respectively. Release of sulforhodamine B from single t‐ SNARE liposomes upon fusion with v‐ SNARE ND s has also been monitored in a dye efflux assay using surface‐tethered liposomes .…”

Section: Nanodisc‐based Fusion Assays: a New Look At Fusion Poresmentioning

confidence: 99%

Toward a unified picture of the exocytotic fusion pore

Karatekin

2018

FEBS Letters

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Neurotransmitter and hormone release involve calcium-triggered fusion of a cargo-loaded vesicle with the plasma membrane. The initial connection between the fusing membranes, called the fusion pore, can evolve in various ways, including rapid dilation to allow full cargo release, slow expansion, repeated opening-closing, and resealing. Pore dynamics determine the kinetics of cargo release and the mode of vesicle recycling, but how these processes are controlled are poorly understood. Previous reconstitutions could not monitor single pores, limiting mechanistic insight they could provide. Recently developed nanodisc-based fusion assays allow reconstitution and monitoring of single pores with unprecedented detail and hold great promise for future discoveries. They recapitulate various aspects of exocytotic fusion pores, but comparison is difficult because different approaches suggested very different exocytotic fusion pore properties, even for the same cell type. In this Review, I discuss how most of the data can be reconciled, by recognizing how different methods probe different aspects of the same fusion process. The resulting picture is that fusion pores have broadly distributed properties arising from stochastic processes which can be modulated by physical constraints imposed by proteins, lipids and membranes.

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Exocytotic fusion pores are composed of both lipids and proteins

Cited by 77 publications

References 56 publications

Extracellular and intracellular sphingosine‐1‐phosphate distinctly regulates exocytosis in chromaffin cells

Extracellular and intracellular sphingosine‐1‐phosphate distinctly regulates exocytosis in chromaffin cells

Dilation of fusion pores by crowding of SNARE proteins

Toward a unified picture of the exocytotic fusion pore

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