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Biochemical and Functional Studies of Cortical Vesicle Fusion: The SNARE Complex and Ca(2+) Sensitivity

Cortical vesicles (CV) possess components critical to the mechanism of exocytosis. The homotypic fusion of CV centrifuged or settled into contact has a sigmoidal Ca(2+) activity curve comparable to exocytosis (CV–PM fusion). Here we show that Sr(2+) and Ba(2+) also trigger CV–CV fusion, and agents a...

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Detalles Bibliográficos
Autores principales: Coorssen, Jens R., Blank, Paul S., Tahara, Masahiro, Zimmerberg, Joshua
Formato: Texto
Lenguaje:English
Publicado: The Rockefeller University Press 1998
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2175215/
https://www.ncbi.nlm.nih.gov/pubmed/9864359
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author Coorssen, Jens R.
Blank, Paul S.
Tahara, Masahiro
Zimmerberg, Joshua
author_facet Coorssen, Jens R.
Blank, Paul S.
Tahara, Masahiro
Zimmerberg, Joshua
author_sort Coorssen, Jens R.
collection PubMed
description Cortical vesicles (CV) possess components critical to the mechanism of exocytosis. The homotypic fusion of CV centrifuged or settled into contact has a sigmoidal Ca(2+) activity curve comparable to exocytosis (CV–PM fusion). Here we show that Sr(2+) and Ba(2+) also trigger CV–CV fusion, and agents affecting different steps of exocytotic fusion block Ca(2+), Sr(2+), and Ba(2+)-triggered CV–CV fusion. The maximal number of active fusion complexes per vesicle, <n\>(Max), was quantified by NEM inhibition of fusion, showing that CV–CV fusion satisfies many criteria of a mathematical analysis developed for exocytosis. Both <n\>(Max) and the Ca(2+) sensitivity of fusion complex activation were comparable to that determined for CV–PM fusion. Using Ca(2+)-induced SNARE complex disruption, we have analyzed the relationship between membrane fusion (CV–CV and CV–PM) and the SNARE complex. Fusion and complex disruption have different sensitivities to Ca(2+), Sr(2+), and Ba(2+), the complex remains Ca(2+)- sensitive on fusion-incompetent CV, and disruption does not correlate with the quantified activation of fusion complexes. Under conditions which disrupt the SNARE complex, CV on the PM remain docked and fusion competent, and isolated CV still dock and fuse, but with a markedly reduced Ca(2+) sensitivity. Thus, in this system, neither the formation, presence, nor disruption of the SNARE complex is essential to the Ca(2+)-triggered fusion of exocytotic membranes. Therefore the SNARE complex alone cannot be the universal minimal fusion machine for intracellular fusion. We suggest that this complex modulates the Ca(2+) sensitivity of fusion.
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spelling pubmed-21752152008-05-01 Biochemical and Functional Studies of Cortical Vesicle Fusion: The SNARE Complex and Ca(2+) Sensitivity Coorssen, Jens R. Blank, Paul S. Tahara, Masahiro Zimmerberg, Joshua J Cell Biol Regular Articles Cortical vesicles (CV) possess components critical to the mechanism of exocytosis. The homotypic fusion of CV centrifuged or settled into contact has a sigmoidal Ca(2+) activity curve comparable to exocytosis (CV–PM fusion). Here we show that Sr(2+) and Ba(2+) also trigger CV–CV fusion, and agents affecting different steps of exocytotic fusion block Ca(2+), Sr(2+), and Ba(2+)-triggered CV–CV fusion. The maximal number of active fusion complexes per vesicle, <n\>(Max), was quantified by NEM inhibition of fusion, showing that CV–CV fusion satisfies many criteria of a mathematical analysis developed for exocytosis. Both <n\>(Max) and the Ca(2+) sensitivity of fusion complex activation were comparable to that determined for CV–PM fusion. Using Ca(2+)-induced SNARE complex disruption, we have analyzed the relationship between membrane fusion (CV–CV and CV–PM) and the SNARE complex. Fusion and complex disruption have different sensitivities to Ca(2+), Sr(2+), and Ba(2+), the complex remains Ca(2+)- sensitive on fusion-incompetent CV, and disruption does not correlate with the quantified activation of fusion complexes. Under conditions which disrupt the SNARE complex, CV on the PM remain docked and fusion competent, and isolated CV still dock and fuse, but with a markedly reduced Ca(2+) sensitivity. Thus, in this system, neither the formation, presence, nor disruption of the SNARE complex is essential to the Ca(2+)-triggered fusion of exocytotic membranes. Therefore the SNARE complex alone cannot be the universal minimal fusion machine for intracellular fusion. We suggest that this complex modulates the Ca(2+) sensitivity of fusion. The Rockefeller University Press 1998-12-28 /pmc/articles/PMC2175215/ /pubmed/9864359 Text en This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 4.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/4.0/).
spellingShingle Regular Articles
Coorssen, Jens R.
Blank, Paul S.
Tahara, Masahiro
Zimmerberg, Joshua
Biochemical and Functional Studies of Cortical Vesicle Fusion: The SNARE Complex and Ca(2+) Sensitivity
title Biochemical and Functional Studies of Cortical Vesicle Fusion: The SNARE Complex and Ca(2+) Sensitivity
title_full Biochemical and Functional Studies of Cortical Vesicle Fusion: The SNARE Complex and Ca(2+) Sensitivity
title_fullStr Biochemical and Functional Studies of Cortical Vesicle Fusion: The SNARE Complex and Ca(2+) Sensitivity
title_full_unstemmed Biochemical and Functional Studies of Cortical Vesicle Fusion: The SNARE Complex and Ca(2+) Sensitivity
title_short Biochemical and Functional Studies of Cortical Vesicle Fusion: The SNARE Complex and Ca(2+) Sensitivity
title_sort biochemical and functional studies of cortical vesicle fusion: the snare complex and ca(2+) sensitivity
topic Regular Articles
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2175215/
https://www.ncbi.nlm.nih.gov/pubmed/9864359
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