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

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Abstract

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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Authors and Affiliations

  1. Department of Physiology, University of Massachusetts Medical School, Worcester, MA

    James M. McNally & José R. Lemos

  2. Department of Physiology and Developmental Biology, Brigham Young University, Provo, UT

    Dixon J. Woodbury

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  1. James M. McNally
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  2. Dixon J. Woodbury
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  3. José R. Lemos
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Correspondence to José R. Lemos.

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McNally, J.M., Woodbury, D.J. & Lemos, J.R. Syntaxin 1A drives fusion of large dense-core neurosecretory granules into a planar lipid bilayer. Cell Biochem Biophys 41, 11–23 (2004). https://doi.org/10.1385/CBB:41:1:011

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