Synaptobrevin binding to synaptophysin: a potential mechanism for controlling the exocytotic fusion machine

EMBO J. 1995 Jan 16;14(2):224-31.

Abstract

The synaptic vesicle protein synaptobrevin (VAMP) has recently been implicated as one of the key proteins involved in exocytotic membrane fusion. It interacts with the synaptic membrane proteins syntaxin I and synaptosome-associated protein (SNAP)-25 to form a complex which precedes exocytosis [Söllner et al. (1993b) Cell, 75, 409-418]. Here we demonstrate that the majority of synaptobrevin is bound to the vesicle protein synaptophysin in detergent extracts. No syntaxin I was found in this complex when synaptophysin-specific antibodies were used for immunoprecipitation. Conversely, no synaptophysin was associated with the synaptobrevin-syntaxin I complex when syntaxin-specific antibodies were used for immunoprecipitation. Thus, the synaptobrevin pool bound to synaptophysin is not available for binding to syntaxin I and SNAP-25, and vice versa. Synaptobrevin-synaptophysin binding was also demonstrated by chemical cross-linking in isolated nerve terminals. Furthermore, recombinant synaptobrevin II efficiently bound synaptophysin and its isoform synaptoporin, but not the more distantly related synaptic vesicle protein p29. Recombinant synaptobrevin I bound with similar efficiency, whereas the non-neuronal isoform cellubrevin displayed a lower affinity towards synaptophysin. Treatment with high NaCl concentrations resulted in a dissociation of the synaptobrevin-synaptophysin complex. In addition, the interaction of synaptobrevin with synaptophysin was irreversibly abolished by low amounts of SDS, while the interaction with syntaxin I was enhanced. We conclude that synaptophysin selectively interacts with synaptobrevin in a complex which excludes the t-SNAP receptors syntaxin I and SNAP-25, suggesting a role for synaptophysin in the control of exocytosis.

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Cell Fusion*
  • Cross-Linking Reagents
  • Exocytosis*
  • Membrane Proteins / isolation & purification
  • Membrane Proteins / metabolism*
  • Molecular Sequence Data
  • Nerve Tissue Proteins / isolation & purification
  • Nerve Tissue Proteins / metabolism*
  • Precipitin Tests
  • Protein Binding
  • R-SNARE Proteins
  • Rats
  • Synaptophysin / isolation & purification
  • Synaptophysin / metabolism*

Substances

  • Cross-Linking Reagents
  • Membrane Proteins
  • Nerve Tissue Proteins
  • R-SNARE Proteins
  • Synaptophysin