Mutations in aDrosophilaα₂δ Voltage-Gated Calcium Channel Subunit Reveal a Crucial Synaptic Function

Abstract

Voltage-dependent calcium channels regulate many aspects of neuronal biology, including synaptic transmission. In addition to their ␣ 1 subunit, which encodes the essential voltage gate and selective pore, calcium channels also contain auxiliary ␣ 2 ␦, ␤, and ␥ 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 an ␣ 2 ␦ calcium channel subunit (d␣ 2 ␦-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. The photoreceptor phenotype of d␣ 2 ␦-3 mutants resembles that of the calcium channel ␣ 1 mutant cacophony (cac), suggesting shared functions. We have examined in detail genotypes that survive to the third-instar stage. Electrophysiological recordings demonstrate that synaptic transmission is severely impaired in these mutants. Thus the ␣ 2 ␦ calcium channel subunit is critical for calcium-dependent synaptic function. As such, this Drosophila isoform is the likely partner to the presynaptic calcium channel ␣ 1 subunit encoded by the cac locus. Consistent with this hypothesis, cacGFP fluorescence at the neuromuscular junction is reduced in d␣ 2 ␦-3 mutants. This is the first characterization of an ␣ 2 ␦-3 mutant in any organism and indicates a necessary role for ␣ 2 ␦-3 in presynaptic vesicle release and calcium channel expression at active zones.

The Physiology, Pathology, and Pharmacology of Voltage-Gated Calcium Channels and Their Future Therapeutic Potential

“…A gene incorrectly termed CACNA2D5 in an article by Whittaker and Hynes (2002) has the sequence of a 2 d-4 (CACNA2D4), and the more remotely related gene, erroneously termed CACNA2D4 in that review corresponds to the more distantly related CACHD1, and has not yet been characterized. Homologous genes are also found in Drosophila melanogaster and Caenorhabditis elegans (Dickman et al, 2008;Saheki and Bargmann, 2009) and have been characterized to affect both calcium channel and presynaptic functions. Furthermore, proteins of the CLCA gene family, which have a related domain structure, have been identified to be auxiliary subunits of calcium-activated chloride channels (Yurtsever et al, 2012).…”

“…The a 2 d subunits have a similar domain structure to many proteins involved in extracellular matrix and extracellular protein-protein interactions (Whittaker and Hynes, 2002). Indeed, a 2 d-1 has been reported to interact with thrombospondins and to have a structural role in synapse formation (Eroglu et al, 2009), and the a 2 d proteins in Drosophila have been shown to have an effect on presynaptic morphology (Dickman et al, 2008;Kurshan et al, 2009), as is also the case for a 2 d-3 in an auditory system synapse (Pirone et al, 2014). Since a 2 d subunits are involved in calcium channel trafficking as well as function, it may be difficult to tease apart their roles independent of calcium channels.…”

Identification of ion channel genes in the Acyrthosiphon pisum genome

“…These proteins, ACYPI005770 and ACYPI008838, show 71% and 47% similarity to the D. melanogaster orthologue, respectively. D. melanogaster also has three a2d subunits, which are post-translationally cleaved into two peptides (a2 and d) (Dickman et al, 2008), and the analysis of the A. pisum genome reveals four orthologues of these genes with the a2d 1 subunit being duplicated (Supplementary Table S2). …”

Frequenin/NCS-1 and the Ca2+-channel α1-subunit co-regulate synaptic transmission and nerve-terminal growth

“…Knockdown of Ca 2+ channels by RNA interference yields reduced nerve-terminal growth (Rieckhof et al, 2003). Conversely, loss-of-function mutations in the  2  voltage-gated Ca 2+ -channel subunit cause enhanced nerve-terminal growth (Dickman et al, 2008;Ly et al, 2008). However, nerve-terminal growth was not altered in fuseless mutants, which probably have defects in intraterminal Ca 2+ levels, because defects in  1 -subunit trafficking were reported (Long et al, 2008).…”