Botulinum toxin dysregulates calcium activity and prevents Dpp/BMP secretion from epithelial cells in the developing...

It has long been thought that neurons are the only cells to utilize ion flux to temporally regulate secretion for cell-to-cell signaling, but we discovered that ectoderm and mesenchymal cells are excitable, undergo voltage-dependent calcium waves, and harness calcium influx to control secretion of essential morphogens, bone morphogenetic proteins (BMPs). Furthermore, altering ion flux during morphogenesis results in craniofacial and limb abnormalities in organisms as diverse as insects and mammals, suggesting that the role for electrical signaling is conserved. In neurons, calcium activates the SNARE complex to drive fusion of secretory vesicles to temporally regulate secretion. Our discoveries inspired the hypothesis that calcium induces the SNARE complex to control BMP secretion. Here, we use pharmacological inhibition of the SNARE complex (Botulinum toxin, BoNT-C) and genetic tools in Drosophila to test this hypothesis in the wing primordia (wing disc). To test whether the SNARE complex is important in epithelial cells for BMP secretion, we apply BoNT-C to the wing disc and measure BMP secretion and calcium activity. We found that BoNT-C inhibits BMP secretion and increases endogenous calcium activity in the epithelial cells of the wing disc, as it does in neurons. Furthermore, expression of BoNT-C in the BMP-producing cells significantly reduces measures of downstream signaling. These data open the possibility that the SNARE complex may control the secretion of other developmental morphogens. Furthermore, our data suggest that pharmacological inhibition of the SNARE complex may not be specific to neuronal targets.