Conserved prefusion protein assembly in regulated exocytosis.

The regulated release of hormones and neurotransmitters is a fundamental process throughout the animal kingdom. The short time scale for the calcium triggering of vesicle fusion in regulated secretion suggests that the calcium sensor synaptotagmin and the soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) membrane fusion machinery are well ordered before the calcium signal. To gain insight into the organization of the prefusion protein assembly in regulated exocytosis, we undertook a structural/functional study of the vesicular synaptotagmin1 and the plasma membrane SNARE proteins, which copurify from the brain in the absence of calcium.

Synaptotagmin interaction with the syntaxin/SNAP-25 dimer is mediated by an evolutionarily conserved motif and is sensitive to inositol hexakisphosphate.

Synaptotagmins are membrane proteins that possess tandem C2 domains and play an important role in regulated membrane fusion in metazoan organisms. Here we show that both synaptotagmins I and II, the two major neuronal isoforms, can interact with the syntaxin/synaptosomal-associated protein of 25 kDa (SNAP-25) dimer, the immediate precursor of the soluble NSF attachment protein receptor (SNARE) fusion complex. A stretch of basic amino acids highly conserved throughout the animal kingdom is responsible for this calcium-independent interaction.

Complexin regulates the closure of the fusion pore during regulated vesicle exocytosis.

Membrane fusion during exocytosis and throughout the cell is believed to involve members of the SNARE (soluble N-ethylmaleimide-sensitive fusion protein attachment protein receptors) family of proteins. The assembly of these proteins into a four-helix bundle may be part of the driving force for bilayer fusion. Regulated exocytosis in neurons and related cell types is specialized to be fast and Ca(2+)-dependent suggesting the involvement of other regulatory proteins specific for regulated exocytosis.