Abstract
The regulation of SNARE complex assembly likely plays an important role in governing the specificity as well as the timing of membrane fusion. Here we identify a novel brain-enriched protein, amisyn, with a tomosyn- and VAMP-like coiled-coil-forming domain that binds specifically to syntaxin 1a and syntaxin 4 both in vitro and in vivo, as assessed by co-immunoprecipitation from rat brain. Amisyn is mostly cytosolic, but a fraction co-sediments with membranes. The amisyn coil domain can form SNARE complexes of greater thermostability than can VAMP2 with syntaxin 1a and SNAP-25 in vitro, but it lacks a transmembrane anchor and so cannot act as a v-SNARE in this complex. The amisyn coil domain prevents the SNAP-25 C-terminally mediated rescue of botulinum neurotoxin E inhibition of norepinephrine exocytosis in permeabilized PC12 cells to a greater extent than it prevents the regular exocytosis of these vesicles. We propose that amisyn forms nonfusogenic complexes with syntaxin 1a and SNAP-25, holding them in a conformation ready for VAMP2 to replace it to mediate the membrane fusion event, thereby contributing to the regulation of SNARE complex formation.
Highlights
The exocytosis of synaptic and dense core vesicles with the plasma membrane in neurons and neuroendocrine cells requires proteins of the SNARE1 families
We propose that amisyn forms nonfusogenic complexes with syntaxin 1a and SNAP-25, holding them in a conformation ready for VAMP2 to replace it to mediate the membrane fusion event, thereby contributing to the regulation of SNARE complex formation
In addition to mediating the fusion event, the SNARE proteins have been implicated in the specificity of membrane fusion, the SNARE hypothesis stating that a particular vSNARE on a transport vesicle should only form a specific complex with its cognate t-SNARE on the correct target membrane, thereby ensuring that the vesicles only fuse with the right compartment [14]
Summary
The exocytosis of synaptic and dense core vesicles with the plasma membrane in neurons and neuroendocrine cells requires proteins of the SNARE1 families. A brain-specific protein named m-tomosyn [27, 28], which binds to syntaxin via a VAMP-like coil domain in its C terminus [29], has recently been proposed to trigger the removal of n-sec1 from syntaxin, thereby regulating formation of the synaptic SNARE complex [27].
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