Synaptic Vesicle Protein 2 Binds Adenine Nucleotides

Jia Yao,Sandra M Bajjalieh

doi:10.1074/jbc.m800738200

Abstract

Synaptic vesicle protein 2 (SV2) is required for normal calcium-regulated secretion of hormones and neurotransmitters. Neurons lacking the two most widely expressed isoforms, SV2A and SV2B, have a reduced readily releasable pool of synaptic vesicles, indicating that SV2 contributes to vesicle priming. The presence of putative ATP-binding sites in SV2 suggested that SV2 might be an ATP-binding protein. To explore this, we examined the binding of the photoaffinity reagent 8-azido-ATP[gamma] biotin to purified, recombinant SV2 in the presence and absence of other nucleotides. Our results indicate that SV2A and SV2B bind nucleotides, with the highest affinity for adenine-containing nucleotides. SV2A contains two binding sites located in the cytoplasmic domains preceding the first and seventh transmembrane domains. These results suggest that SV2-mediated vesicle priming could be regulated by adenine nucleotides, which might provide a link between cellular energy levels and regulated secretion.

Highlights

When Synaptic vesicle protein 2 (SV2) expression is knocked out or reduced in neurons, chromaffin cells, or pancreatic islet cells, regulated secretion is decreased because of a reduction in the number of vesicles primed for release (10 –12)
To explore the possibility that SV2 action requires or is regulated by nucleotide binding, we examined the ability of the photoaffinity reagent 8-azido-ATP[␥] biotin to bind to recombinant SV2 and the ability of other nucleotides to compete for binding
SV2 is a synaptic vesicle protein that contributes to vesicle priming

Summary

Introduction

When SV2 expression is knocked out or reduced in neurons, chromaffin cells, or pancreatic islet cells, regulated secretion is decreased because of a reduction in the number of vesicles primed for release (10 –12). Synapses from SV2 knock-out mice have equal numbers of vesicles attached to the plasma membrane [10] and fewer SNARE complexes [12]. This suggests that SV2 acts after vesicle docking and before SNARE complex. Loss of SV2 does not affect vesicular uptake of calcium [11]3 or neurotransmitters.4 It is unclear whether SV2 acts as a transporter or, like adenylate cyclase, has a transporter-like structure but performs a nontransport function [15]. Several major facilitator transporters contain nucleotidebinding sites and in some cases ATP binding regulates transporter activity. To explore the possibility that SV2 action requires or is regulated by nucleotide binding, we examined the ability of the photoaffinity reagent 8-azido-ATP[␥] biotin to bind to recombinant SV2 and the ability of other nucleotides to compete for binding

Methods

Results

Conclusion