Genetic dissection of the mechanisms and functions of post-transcriptional calcium channel modifications in Drosophila

• Applicant: Dr. Stefanie Ryglewski
• Subject Area: Cognitive, Systems and Behavioural Neurobiology
• Term: from 2015 to 2023
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 275581320

Voltage gated calcium channels (VGCCs) control multiple pivotal aspects of neuronal function, including synaptic vesicle release from presynaptic terminals, the amplification of postsynaptic potentials in dendrites, and activity dependent control of transcription in neuronal somata. Mammalian VGCC α1 subunits are encoded by 10 genes that are classified in 3 families, Cav1, Cav2, and Cav3. Each channel differs with respect to biophysical properties, localization, pharmacological profile, and thus function. Regardless, the essential functions of VGCC outnumber the available genes, but VGCC functional diversity can be dramatically increased by alternative splicing and co-assembly of the pore forming alpha1 subunit with accessory subunits. However, the resulting effects on VGCC biophysical properties and Sub cellular localization as well as the functional consequences are incompletely understood. We will combine electro- and optophysiological tools with molecular biology in the Drosophila genetic model system to address these questions. In Drosophila, each vertebrate VGCC family is represented by one gene: Dmca1D is homologous to Cav1, Dmca1A or cacophony to Cav2, andDmαG to Cav3. This results in less than 10 combinations for coassembly with accessory subunits, as opposed to more than 100 in mammals. During the last funding period, we have identifieddevelopmental and acute functions of specific VGCCs in different subneuronalcompartments of Drosophila motoneurons. We next started unraveling the functional code of Drosophila HVA channel interactions with accessory subunits by demonstrating that pairing of the Cav2 homolog cacophony with different accessory subunits affects distinctaspects of channel biophysical properties and sub-neuronal localization. Channel functional diversity is further increased by differential splicing. In fact Drosophila Cav2 channels produce currents with markedly different activation voltages and kinetics, andwe have developed the tools to test the underlying roles of alternativeisoform selection. Based on these findings we will now test how alternative splicing (aim 1) and pairing with accessory subunits (aim 2) tune the biophysical properties and the localization of DrosophilaCav2 channels. We will then test the resulting functional consequences on the cellular and behavioral levels by probing motoneuron excitability and input-output operations as well as firing patterns during behavior (aim 3). We expect novel insights into themechanisms that generate functional VGCC diversity that is relevant for specific neuronal operations and healthy brain function.

• DFG Programme: Research Grants