Much of what we know about the cardiac Ryanodine receptor (RyR2) relates to its role in excitation-contraction (EC) coupling. During this process, cell membrane voltage-gated Ca²⁺channels are activated by depolarization and transport Ca²⁺ into the cytosol where it binds to the high-affinity Ca²⁺ activation sites of RyR2. When RyR2 channels open, a large amount of calcium travels from the sarcoplasmic reticulum (SR) lumen into the cytoplasm, leading to the activation of contractile proteins. However, RyR2 plays also a key role in regulating SR calcium levels. When the SR lumen levels of Ca²⁺ reach a certain threshold, RyR2 spontaneously releases it into the cytoplasm. This store overload-induced Ca²⁺ release (SOICR) can result in cytosolic Ca²⁺ waves and delayed afterpolarizations that may interfere with the normal EC coupling, leading to ventricular tachyarrhythmias and sudden death. In support of this notion, mutations on RyR2 that reduce the threshold for SOICR lead to congenital ventricular tachycardias (CPTV) under stress conditions.

In the 19 January edition of Nature Medicine a group lead by SR Wayne Chen at University of Calgary’s Libin Institute unveils the mechanism that operates SOICR. Using site-directed mutagenesis of negatively charged residues located in the proposed ion gate of the channel, the authors identified residue E4872 that, when mutated, renders the channel completely unresponsive to luminal Ca²⁺ (refractory to SOICR). This residue locates in a helix bundle crossing region (the proposed RyR2 gate) and establishes electrostatic intersubunit interactions with E4868 and R4874 to keep the gate closed. The authors propose a model in which at high lumen Ca²⁺ concentrations this cation binds to E4872 to disrupt the electrostatic interactions with D4868 and R4874 driving the transition of the channel from the closed to the open state.

The authors used several techniques (planar lipid bilayers, mutant RyR2 expressed in Hek293 cells and HL-1 cardiac cells, freshly isolated cardiomyocytes and intact hearts) to demonstrate that mutation on the residues involved in the electrostatic interactions of the helix bundle crossing region raise the threshold for SOICR. But would a mutation on this Ca²⁺ activation site be able to protect against stress-induced ventricular tachycardia? RyR2 R4496C is a mutation identified in CPTV patients that, when expressed in mice, leads to a similar cardiac phenotype. Chen’s group generated a R4496C/E4872Q compound heterozygous mice and found that they are completely resistant to CPTV induction. This result shows that RyR2 store Ca²⁺-sensing gate represents a potential therapeutic target for anti-arrhythmic therapies.

Furthermore, the level of conservation between the three RyR isoforms indicate that they share a similar SR Ca²⁺ sensing mechanism and that drugs directed against RyR2 Ca²⁺ sensing site can also be used to treat other SOICR related conditions, such as malignant hyperthermia that is caused by RyR1 mutations.

Chen’s team has shown how beneficial the silencing of SOICR would be in the disease scenario, but may it also lead to any undesired secondary effect?  Some of the published results point towards a complex scenario for SOICR:

• Mutation of E4872 makes the RyR2 channel less sensitive to activation through cytosolic Ca²⁺, demonstrating that both lumen and cytosol RyR2 Ca²⁺ sensing mechanisms are different but interactive.
• Mutation of E4872 affects EC coupling. The L-type Ca²⁺ channel current in E4872Q cardiomyocytes was significantly increased resulting in a reduced EC coupling gain, and the late decay times were faster.
• The fact that homozygous E4872Q mouse is embryonically lethal may indicate new roles of RyR2 in other scenarios (beyond development) and that total SOICR block may lead to complications.
• Drugs directed against the cardiac RyR2 may interfere with RyR1 and RyR3 and lead to undesired side effects on other tissues.

In summary, the recent results published by Chen et al depict a new mechanism of cardiac SOICR and should lead to the design of new therapies to treat stress induced arrhythmia and malignant hyperthermia. A better understanding of how other proteins involved in luminal Ca²⁺ sensitivity, such as calsequestrin (CASQ2), triadin and junctin integrate in this system would help to better understand this process and design new therapies.

Wenqian Chen et al (2014). The ryanodine receptor store-sensing gate controls Ca2+ waves and Ca2+-triggered arrhythmias. Nature Medicine 20, 184–192 https://www.nature.com/nm/journal/v20/n2/full/nm.3440.html