Key result
Expression of arrhythmogenic CaM mutations in zebrafish embryos triggered altered heart rates and irregular rhythms, alongside decreased structural stability and reduced Ca2+-binding affinities.
Why the study?
Arrhythmogenic calmodulin mutations are linked to life-threatening syndromes such as LQTS and CPVT, but their in vivo functional effects on cardiac function require investigation.
Arrhythmogenic CaM mutations trigger different arrhythmogenic phenotypes, such as bradycardia or tachycardia, through divergent biochemical properties and reduced Ca2+-binding affinities.
May link CaM mutations to bradycardia or tachycardia; leaves open clinical relevance pending human validation.
Calmodulin (CaM) is a ubiquitous, small cytosolic calcium (Ca 2+ )‐binding sensor that plays a vital role in many cellular processes by binding and regulating the activity of over 300 protein targets. In cardiac muscle, CaM modulates directly or indirectly the activity of several proteins that play a key role in excitation‐contraction coupling (ECC), such as ryanodine receptor type 2 (RyR2), l ‐type Ca 2+ (Ca v 1.2), sodium (NaV1.5) and potassium (KV7.1) channels. Many recent clinical and genetic studies have reported a series of CaM mutations in patients with life‐threatening arrhythmogenic syndromes, such as long QT syndrome (LQTS) and catecholaminergic polymorphic ventricular tachycardia (CPVT). We recently showed that four arrhythmogenic CaM mutations (N98I, D132E, D134H, and Q136P) significantly reduce the binding of CaM to RyR2. Herein, we investigate in vivo functional effects of these CaM mutations on the normal zebrafish embryonic heart function by microinjecting complementary RNA corresponding to CaM N98I , CaM D132E , CaM D134H , and CaM Q136P mutants. Expression of CaM D132E and CaM D134H mutants results in significant reduction of the zebrafish heart rate, mimicking a severe form of human bradycardia, whereas expression of CaM Q136P results in an increased heart rate mimicking human ventricular tachycardia. Moreover, analysis of cardiac ventricular rhythm revealed that the CaM D132E and CaM N98I zebrafish groups display an irregular pattern of heart beating and increased amplitude in comparison to the control groups. Furthermore, circular dichroism spectroscopy experiments using recombinant CaM proteins reveals a decreased structural stability of the four mutants compared to the wild‐type CaM protein in the presence of Ca 2+ . Finally, Ca 2+ ‐binding studies indicates that all CaM mutations display reduced CaM Ca 2+ ‐binding affinities, with CaM D132E exhibiting the most prominent change. Our data suggest that CaM mutations can trigger different arrhythmogenic phenotypes through multiple and complex molecular mechanisms.
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Da’as et al. (2024) studied Arrhythmogenic syndromes (LQTS, CPVT). Microinjection of arrhythmogenic CaM mutations (N98I, D132E, D134H, Q136P) vs. Control groups / wild-type CaM was evaluated on Heart rate, rhythm, structural stability, and Ca2+-binding affinity. Expression of arrhythmogenic CaM mutations in zebrafish embryos triggered altered heart rates and irregular rhythms, alongside decreased structural stability and reduced Ca2+-binding affinities.
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