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Trans-SNARE complex dynamics and number determine nascent fusion pore properties
The fusion pore is the first crucial intermediate formed during exocytosis, yet little is known regarding the mechanisms that determine the size and kinetic properties of these transient structures(1). Here, we reduced the number of available SNAREs in neurons and observed changes in transmitter rel...
Autores principales: | , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
2018
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5808578/ https://www.ncbi.nlm.nih.gov/pubmed/29420480 http://dx.doi.org/10.1038/nature25481 |
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author | Bao, Huan Das, Debasis Courtney, Nicholas A. Jiang, Yihao Briguglio, Joseph S. Lou, Xiaochu Roston, Daniel Cui, Qiang Chanda, Baron Chapman, Edwin R. |
author_facet | Bao, Huan Das, Debasis Courtney, Nicholas A. Jiang, Yihao Briguglio, Joseph S. Lou, Xiaochu Roston, Daniel Cui, Qiang Chanda, Baron Chapman, Edwin R. |
author_sort | Bao, Huan |
collection | PubMed |
description | The fusion pore is the first crucial intermediate formed during exocytosis, yet little is known regarding the mechanisms that determine the size and kinetic properties of these transient structures(1). Here, we reduced the number of available SNAREs in neurons and observed changes in transmitter release suggestive of alterations in fusion pores. To address this, we employed reconstituted fusion assays using nanodiscs to trap pores in their initial open state. Optical measurements revealed that increasing the number of SNARE complexes enhanced the rate of release from single pores, and enabled the escape of larger cargos. To determine whether this was due to changes in nascent pore size versus stability, we developed a novel approach, based on nanodiscs and planar lipid bilayer electrophysiology, that affords μsec time resolution at the single event level. Remarkably, both parameters were affected by SNARE copy number. Increasing the number of v-SNAREs per nanodisc from three to five caused a two-fold increase in pore size and decreased the rate of pore closure by more than three orders of magnitude. Moreover, trans-SNARE pairing was highly dynamic: flickering nascent pores closed upon addition of a v-SNARE fragment, revealing that the fully assembled, stable, SNARE complex does not form at this stage of exocytosis. Finally, a deletion at the base of the SNARE complex, that mimics the action of botulinum neurotoxin A, dramatically reduced fusion pore stability. In summary, trans-SNARE complexes are dynamic, and the number of SNAREs recruited to drive fusion determine fundamental properties of individual pores. |
format | Online Article Text |
id | pubmed-5808578 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2018 |
record_format | MEDLINE/PubMed |
spelling | pubmed-58085782018-07-31 Trans-SNARE complex dynamics and number determine nascent fusion pore properties Bao, Huan Das, Debasis Courtney, Nicholas A. Jiang, Yihao Briguglio, Joseph S. Lou, Xiaochu Roston, Daniel Cui, Qiang Chanda, Baron Chapman, Edwin R. Nature Article The fusion pore is the first crucial intermediate formed during exocytosis, yet little is known regarding the mechanisms that determine the size and kinetic properties of these transient structures(1). Here, we reduced the number of available SNAREs in neurons and observed changes in transmitter release suggestive of alterations in fusion pores. To address this, we employed reconstituted fusion assays using nanodiscs to trap pores in their initial open state. Optical measurements revealed that increasing the number of SNARE complexes enhanced the rate of release from single pores, and enabled the escape of larger cargos. To determine whether this was due to changes in nascent pore size versus stability, we developed a novel approach, based on nanodiscs and planar lipid bilayer electrophysiology, that affords μsec time resolution at the single event level. Remarkably, both parameters were affected by SNARE copy number. Increasing the number of v-SNAREs per nanodisc from three to five caused a two-fold increase in pore size and decreased the rate of pore closure by more than three orders of magnitude. Moreover, trans-SNARE pairing was highly dynamic: flickering nascent pores closed upon addition of a v-SNARE fragment, revealing that the fully assembled, stable, SNARE complex does not form at this stage of exocytosis. Finally, a deletion at the base of the SNARE complex, that mimics the action of botulinum neurotoxin A, dramatically reduced fusion pore stability. In summary, trans-SNARE complexes are dynamic, and the number of SNAREs recruited to drive fusion determine fundamental properties of individual pores. 2018-01-31 2018-02-08 /pmc/articles/PMC5808578/ /pubmed/29420480 http://dx.doi.org/10.1038/nature25481 Text en Users may view, print, copy, and download text and data-mine the content in such documents, for the purposes of academic research, subject always to the full Conditions of use: http://www.nature.com/authors/editorial_policies/license.html#terms |
spellingShingle | Article Bao, Huan Das, Debasis Courtney, Nicholas A. Jiang, Yihao Briguglio, Joseph S. Lou, Xiaochu Roston, Daniel Cui, Qiang Chanda, Baron Chapman, Edwin R. Trans-SNARE complex dynamics and number determine nascent fusion pore properties |
title | Trans-SNARE complex dynamics and number determine nascent fusion pore properties |
title_full | Trans-SNARE complex dynamics and number determine nascent fusion pore properties |
title_fullStr | Trans-SNARE complex dynamics and number determine nascent fusion pore properties |
title_full_unstemmed | Trans-SNARE complex dynamics and number determine nascent fusion pore properties |
title_short | Trans-SNARE complex dynamics and number determine nascent fusion pore properties |
title_sort | trans-snare complex dynamics and number determine nascent fusion pore properties |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5808578/ https://www.ncbi.nlm.nih.gov/pubmed/29420480 http://dx.doi.org/10.1038/nature25481 |
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