Distinctive malfunctions of calmodulin mutations associated with heart RyR2-mediated arrhythmic disease

Vassilakopoulou, Vyronia; Calver, Brian L.; Thanassoulas, Angelos; Beck, Konrad; Hu, Handan; Buntwal, Luke; Smith, Adrian; Theodoridou, Maria; Kashir, Junaid; Blayney, Lynda Mary; Livaniou, Evangelia; Nounesis, George; Lai, F. Anthony; Nomikos, Michail

doi:10.1016/j.bbagen.2015.07.001

Cited by 34 publications

(83 citation statements)

References 45 publications

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“…The N54I variant is unique, as it is the only mutant located in the N‐terminal lobe of CaM and it is not part of the Ca 2+ coordination or the hydrophobic target binding patches. In line with this, with this, it has previously been found that the Ca 2+ ‐binding affinity of CaM in neither the C‐ nor N‐domain is affected (Vassilakopoulou ). Most studies point toward a defective RyR2 interaction and regulation by CPVT‐associated CaM variants as being the underlying mechanism (Hwang ).…”

Section: Discussionsupporting

confidence: 81%

Impact of arrhythmogenic calmodulin variants on small conductance Ca ²⁺ ‐activated K ⁺ (SK3) channels

et al. 2019

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Calmodulin (CaM) is a ubiquitous Ca2+‐sensing protein regulating many important cellular processes. Several CaM‐associated variants have been identified in a small group of patients with cardiac arrhythmias. The mechanism remains largely unknown, even though a number of ion channels, including the ryanodine receptors and the L‐type calcium channels have been shown to be functionally affected by the presence of mutant CaM. CaM is constitutively bound to the SK channel, which underlies the calcium‐gated ISK contributing to cardiac repolarization. The CaM binding to SK channels is essential for gating, correct assembly, and membrane expression. To elucidate the effect of nine different arrhythmogenic CaM variants on SK3 channel function, HEK293 cells stably expressing SK3 were transiently co‐transfected with CaMWT or variant and whole‐cell patch‐clamp recordings were performed with a calculated free Ca2+ concentration of 400 nmol/L. MDCK cells were transiently transfected with SK3 and/or CaMWT or variant to address SK3 and CaM localization by immunocytochemistry. The LQTS‐associated variants CaMD96V, CaMD130G, and CaMF142L reduced ISK,Ca compared with CaMWT (P < 0.01, P < 0.001, and P < 0.05, respectively). The CPVT associated variant CaMN54I also reduced the ISK,Ca (P < 0.05), which was linked to an accumulation of SK3/CaMN54I channel complexes in intracellular compartments (P < 0.05). The CPVT associated variants, CaMA103V and CaMD132E only revealed a tendency toward reduced current, while the variants CaMF90L and CaMN98S, causing LQTS syndrome, did not have any impact on ISK,Ca. In conclusion, we found that the arrhythmogenic CaM variants CaMN54I, CaMD96V, CaMD130G, and CaMF142L significantly down‐regulate the SK3 channel current, but with distinct mechanism.

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Section: Discussionsupporting

confidence: 81%

Impact of arrhythmogenic calmodulin variants on small conductance Ca ²⁺ ‐activated K ⁺ (SK3) channels

et al. 2019

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“…and Vassilakopoulou et al . found that CaM‐N54I, ‐D96V, ‐F90L and ‐D130G increased [ 3 H]‐ryanodine binding (a measure of RyR2 open propensity) to porcine cardiac SR vesicles . We previously found that CaM‐N54I, ‐D96V, ‐N98S and ‐D130G markedly diminished CaM‐dependent inhibition of RyR2‐mediated SOICR in HEK293 cells and, to a noticeably smaller degree, so did CaM‐F142L.…”

Section: Discussionmentioning

confidence: 91%

Diminished inhibition and facilitated activation of RyR2‐mediated Ca²⁺ release is a common defect of arrhythmogenic calmodulin mutations

et al. 2019

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A number of calmodulin (CaM) mutations cause severe cardiac arrhythmias, but their arrhythmogenic mechanisms are unclear. While some of the arrhythmogenic CaM mutations have been shown to impair CaM‐dependent inhibition of intracellular Ca2+ release through the ryanodine receptor type 2 (RyR2), the impact of a majority of these mutations on RyR2 function is unknown. Here, we investigated the effect of 14 arrhythmogenic CaM mutations on the CaM‐dependent RyR2 inhibition. We found that all the arrhythmogenic CaM mutations tested diminished CaM‐dependent inhibition of RyR2‐mediated Ca2+ release and increased store‐overload induced Ca2+ release (SOICR) in HEK293 cells. Moreover, all the arrhythmogenic CaM mutations tested either failed to inhibit or even promoted RyR2‐mediated Ca2+ release in permeabilized HEK293 cells with elevated cytosolic Ca2+, which was markedly different from the inhibitory action of CaM wild‐type. The CaM mutations also altered the Ca2+‐dependency of CaM binding to the RyR2 CaM‐binding domain. These results demonstrate that diminished inhibition, and even facilitated activation, of RyR2–mediated Ca2+ release is a common defect of arrhythmogenic CaM mutations.

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“…In cardiac cells, CaM is known to interact and regulate multiple key proteins involved in excitation–contraction coupling (ECC) and Ca 2+ homeostasis. These proteins include the cardiac ryanodine receptor type 2 (RyR2), voltage‐operated potassium (K + ), sodium (Na + ), and L‐type Ca 2+ channels . The RyR2 is a large transmembrane high‐conductance Ca 2+ release channel that mediates Ca 2+ release from the sarcoplasmic reticulum (SR) to activate cardiac muscle contraction .…”

Section: Introductionmentioning

confidence: 99%

“…These proteins include the cardiac ryanodine receptor type 2 (RyR2), voltage‐operated potassium (K + ), sodium (Na + ), and L‐type Ca 2+ channels . The RyR2 is a large transmembrane high‐conductance Ca 2+ release channel that mediates Ca 2+ release from the sarcoplasmic reticulum (SR) to activate cardiac muscle contraction . RyR2 gating is crucial for the receptor function and is mediated by a number of modulators (including ions, small molecules, and proteins), which regulate the exact timing of channel opening or closing .…”

Section: Introductionmentioning

confidence: 99%

“…In humans, there are three CaM genes ( CALM1, CALM2 , and CALM3 ) that encode an identical protein and are all expressed in cardiac tissue . A number of recent genetic studies have identified several missense mutations in all three CaM genes, in individuals with a family history of severe cardiac disorders and early onset sudden cardiac death …”

Section: Introductionmentioning

confidence: 99%

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Arrhythmogenic calmodulin E105A mutation alters cardiac RyR2 regulation leading to cardiac dysfunction in zebrafish

Da’as

Thanassoulas

Calver

et al. 2019

Annals of the New York Academy of Sciences

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Calmodulin (CaM) is a universal calcium (Ca2+)‐binding messenger that regulates many vital cellular events. In cardiac muscle, CaM associates with ryanodine receptor 2 (RyR2) and regulates excitation–contraction coupling. Mutations in human genes CALM1, CALM2, and CALM3 have been associated with life‐threatening heart disorders, such as long QT syndrome (LQTS) and catecholaminergic polymorphic ventricular tachycardia. A novel de novo LQTS‐associated missense CaM mutation (E105A) was recently identified in a 6‐year‐old boy, who experienced an aborted first episode of cardiac arrest. Herein, we report the first molecular characterization of the CaM E105A mutation. Expression of the CaM E105A mutant in zebrafish embryos resulted in cardiac arrhythmia and increased heart rate, suggestive of ventricular tachycardia. In vitro biophysical and biochemical analysis revealed that E105A confers a deleterious effect on protein stability and a reduced Ca2+‐binding affinity due to loss of cooperativity. Finally, the CaM E105A mutation resulted in reduced CaM–RyR2 interaction and defective modulation of ryanodine binding. Our findings suggest that the CaM E105A mutation dysregulates normal cardiac function by a complex mechanism involving alterations in both CaM–Ca2+ and CaM–RyR2 interactions.

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Distinctive malfunctions of calmodulin mutations associated with heart RyR2-mediated arrhythmic disease

Cited by 34 publications

References 45 publications

Impact of arrhythmogenic calmodulin variants on small conductance Ca ²⁺ ‐activated K ⁺ (SK3) channels

Impact of arrhythmogenic calmodulin variants on small conductance Ca ²⁺ ‐activated K ⁺ (SK3) channels

Diminished inhibition and facilitated activation of RyR2‐mediated Ca²⁺ release is a common defect of arrhythmogenic calmodulin mutations

Arrhythmogenic calmodulin E105A mutation alters cardiac RyR2 regulation leading to cardiac dysfunction in zebrafish

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Distinctive malfunctions of calmodulin mutations associated with heart RyR2-mediated arrhythmic disease

Cited by 34 publications

References 45 publications

Impact of arrhythmogenic calmodulin variants on small conductance Ca 2+ ‐activated K + (SK3) channels

Impact of arrhythmogenic calmodulin variants on small conductance Ca 2+ ‐activated K + (SK3) channels

Diminished inhibition and facilitated activation of RyR2‐mediated Ca2+ release is a common defect of arrhythmogenic calmodulin mutations

Arrhythmogenic calmodulin E105A mutation alters cardiac RyR2 regulation leading to cardiac dysfunction in zebrafish

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Impact of arrhythmogenic calmodulin variants on small conductance Ca ²⁺ ‐activated K ⁺ (SK3) channels

Impact of arrhythmogenic calmodulin variants on small conductance Ca ²⁺ ‐activated K ⁺ (SK3) channels

Diminished inhibition and facilitated activation of RyR2‐mediated Ca²⁺ release is a common defect of arrhythmogenic calmodulin mutations