Syntaxin 1A drives fusion of large dense-core neurosecretory granules into a planar lipid bilayer

The SNARE complex, involved in vesicular trafficking and exocytosis, is composed of proteins in the vesicular membrane (v-SNAREs) that intertwine with proteins of the target membrane (t-SNAREs). Our results show that modified large dense-core neurosecretory granules (NSGs), isolated from the bovine...

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Published inCell biochemistry and biophysics Vol. 41; no. 1; pp. 11 - 24
Main Authors McNally, James M, Woodbury, Dixon J, Lemos, José R
Format Journal Article
LanguageEnglish
Published United States Springer Nature B.V 01.01.2004
Subjects
Amino Acid Motifs
Animals
Binding sites
Biophysics - methods
Calcium - metabolism
Cattle
Edetic Acid - chemistry
Exocytosis
Gene Deletion
Hydrogen-Ion Concentration
Lipid Bilayers - chemistry
Membranes
Mutation
Protein Structure, Tertiary
Proteins
Secretory Vesicles - metabolism
Solubility
Syntaxin 1 - metabolism
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Summary:The SNARE complex, involved in vesicular trafficking and exocytosis, is composed of proteins in the vesicular membrane (v-SNAREs) that intertwine with proteins of the target membrane (t-SNAREs). Our results show that modified large dense-core neurosecretory granules (NSGs), isolated from the bovine neurohypophysis, spontaneously fuse with a planar lipid membrane containing only the t-SNARE syntaxin 1A. This provides evidence that syntaxin alone is able to form a functional fusion complex with native v-SNAREs of the NSG. The fusion was similar to constitutive, not regulated, exocytosis because changes in free [Ca2+] had no effect on the syntaxin-mediated fusion. Several deletion mutants of syntaxin 1A were also tested. The removal of the regulatory domain did not significantly reduce spontaneous fusion. However, a syntaxin deletion mutant consisting of only the transmembrane domain was incapable of eliciting spontaneous fusion. Finally, a soluble form of syntaxin 1A (lacking its transmembrane domain) was used to saturate the free syntaxin-binding sites of modified NSGs. This treatment blocks spontaneous fusion of these granules to a bilayer containing full-length syntaxin 1A. This method provides an effective model system to study possible regulatory components affecting vesicle fusion.
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ISSN:1085-9195
1559-0283
1085-9195
DOI:10.1385/cbb:41:1:011