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(Circulation Research. 2006;98:88.)
© 2006 American Heart Association, Inc.

Cellular Biology

Arrhythmogenic Mutation-Linked Defects in Ryanodine Receptor Autoregulation Reveal a Novel Mechanism of Ca²⁺ Release Channel Dysfunction

Christopher H. George, Hala Jundi, Nicola Walters, N. Lowri Thomas, Robert R. West, F. Anthony Lai

From the Department of Cardiology, Wales Heart Research Institute, Cardiff University School of Medicine, Cardiff, UK.

Correspondence to Dr Christopher H. George, Department of Cardiology, Wales Heart Research Institute, Cardiff University, Heath Park, Cardiff, UK CF14 4XN. E-mail georgech{at}cf.ac.uk

Arrhythmogenic cardiac ryanodine receptor (RyR2) mutations are associated with stress-induced malignant tachycardia, frequently leading to sudden cardiac death (SCD). The causative mechanisms of RyR2 Ca²⁺ release dysregulation are complex and remain controversial. We investigated the functional impact of clinically-severe RyR2 mutations occurring in the central domain, and the C-terminal I domain, a key locus of RyR2 autoregulation, on interdomain interactions and Ca²⁺ release in living cells. Using high-resolution confocal microscopy and fluorescence resonance energy transfer (FRET) analysis of interaction between fusion proteins corresponding to amino- (N-) and carboxyl- (C-) terminal RyR2 domains, we determined that in resting cells, RyR2 interdomain interaction remained unaltered after introduction of SCD-linked mutations and normal Ca²⁺ regulation was maintained. In contrast, after channel activation, the abnormal Ca²⁺ release via mutant RyR2 was intrinsically linked to altered interdomain interaction that was equivalent with all mutations and exhibited threshold characteristics (caffeine >2.5 mmol/L; Ca²⁺ >150 nmol/L). Noise analysis revealed that I domain mutations introduced a distinct pattern of conformational instability in Ca²⁺ handling and interdomain interaction after channel activation that was absent in signals obtained from the central domain mutation. I domain–linked channel instability also occurred in intact RyR2 expressed in CHO cells and in HL-1 cardiomyocytes. These new insights highlight a critical role for mutation-linked defects in channel autoregulation, and may contribute to a molecular explanation for the augmented Ca²⁺ release following RyR2 channel activation. Our findings also suggest that the mutational locus may be an important mechanistic determinant of Ca²⁺ release channel dysfunction in arrhythmia and SCD.

Key Words: ryanodine receptor • mutations • interdomain interaction • arrhythmia

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