| Voltage-dependent calcium channels regulate many aspects of neuronal biology, including synaptic transmission. In addition to their alpha 1 subunit, which encodes the essential voltage gate and selective pore, calcium channels also contain auxiliary alpha2delta, beta, and gamma subunits. Despite progress in understanding the biophysical properties of calcium channels, the in vivo functions of these auxiliary subunits remain unclear. We have isolated mutations in the gene encoding alpha2delta-3 using a forward genetic screen in Drosophila. Null mutations in this gene are embryonic lethal and can be rescued by expression in the nervous system, demonstrating that the essential function of this subunit is neuronal. Electrophysiological recordings from genotypes that survive to the third-instar stage demonstrate that synaptic transmission is severely impaired in these mutants, suggesting shared functions with the calcium channel alpha1 mutant cacophony (cac). We conclude that alpha2delta-3 is critical for calcium-dependent synaptic function and that this Drosophila isoform is the likely partner to the N-type calcium channel alpha1 subunit encoded by the cac locus. Consistent with this hypothesis, cac-GFP fluorescence at the neuromuscular junction is reduced in alpha2delta-3 mutants. This is the first characterization of an alpha2delta-3 mutant in any organism and indicates a necessary role for alpha2delta-3 in presynaptic vesicle release and calcium channel expression at active zones.;Synaptogenesis involves the transformation of a growth cone into synaptic boutons specialized for transmitter release. In Drosophila alpha 2delta-3 null embryos, we find that motor neuron terminals fail to develop synaptic boutons and cytoskeletal abnormalities arise, including the loss of ankyrin2. Nevertheless, functional presynaptic specializations are present and apposed to clusters of postsynaptic glutamate receptors. The alpha 2delta-3 protein has been thought to function strictly as an auxiliary subunit of the Ca2+ channel, but the phenotype of alpha2delta-3 mutations cannot be explained by a channel defect: embryos lacking the pore-forming alpha1 subunit cacophony form boutons. The synaptogenic function of alpha 2delta-3 requires only the alpha2 peptide, whose expression suffices to rescue bouton formation. Our results indicate that alpha 2delta proteins have functions independent of their roles in the biophysics and localization of Ca2+ channels, and synaptic architecture depends on these novel functions. |
