Characterization of catecholaminergic polymorphic ventricular tachycardia (CPVT) mutations and specific drug screen in the pharynx of C. elegans as an optogenetic arrhythmia model

• Applicant: Dr. Christina Schüler
• Subject Area: Cell Biology ; Biochemistry ; Biophysics
• Term: since 2016
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 321173572

Catecholaminergic polymorphic ventricular tachycardia (CPVT) is a condition characterized by physical activity or stress induced abnormal heart rhythm (arrhythmia). The disease derives from an alteration of intracellular calcium handling involving the calcium-induced calcium release mechanism of myocytes. Most mutations that have been linked to CPVT are found in two genes, the Ryanodine receptor 2 (RyR2), a calcium release channel located in the sarcoplasmic reticulum (SR), and Calsequestrin 2 (CASQ2), the major calcium binding protein in the SR. Many different CPVT mutations are known (>180). Thus, to enable a specific drug development, but also to study distinct arrhythmias, simple model systems are required, that allow to implement and to characterize these mutations. We assessed the pharynx of Caenorhabditis elegans, which is a rhythmically active muscular pump, as a heart model. The pharynx utilizes homologs of most of the ion channels, pumps and transporters defining human cardiac physiology. To yield precise rhythmicity, we optically pace the pharynx using Channelrhodopsin-2. We will genomically insert, using the CRISPR-Cas9 approach, different mutations of RyR2 (unc-68 in C. elegans), known to induce CPVT in humans, but are also located in a proposed calstabin 2 (FKBP12.6) binding site or close to areas known to disturb calstabin binding. Furthermore, we will investigate the stabilization of the closed-state of RyR via calstabin binding in C. elegans in more detail by testing further calstabin isoforms, or rather their knock-outs. By electrophysiological recordings and calcium and voltage imaging, we will characterize the CPVT mutations. Using a microfluidic chip based electrophysiological method for higher throughput screening, in combination with the optogenetic stimulation, we will identify mutation-specific drugs. In addition, we will extend our optogenetic pharynx model for the analysis of RyR1-mutation that evoke neuromuscular diseases.

• DFG Programme: Research Grants