A Novel Ryanodine Receptor Mutation Linked to Sudden Death Increases Sensitivity to Cytosolic Calcium

Male 0301 basic medicine Ryanodine Receptor Calcium Release Channel Middle Aged Cyclic AMP-Dependent Protein Kinases Recombinant Proteins 3. Good health Tacrolimus Binding Proteins Electrocardiography 03 medical and health sciences Catecholamines Cytosol Death, Sudden, Cardiac HEK293 Cells Phenotype Diastole Mutagenesis, Site-Directed Tachycardia, Ventricular Humans Point Mutation Calcium Ion Channel Gating
DOI: 10.1161/circresaha.111.244970 Publication Date: 2011-06-10T09:07:57Z
ABSTRACT
Rationale: Mutations in the cardiac type 2 ryanodine receptor (RyR2) have been linked to catecholaminergic polymorphic ventricular tachycardia (CPVT). CPVT-associated RyR2 mutations cause fatal ventricular arrhythmias in young individuals during β-adrenergic stimulation. Objective: This study sought to determine the effects of a novel RyR2-G230C mutation and whether this mutation and RyR2-P2328S alter the sensitivity of the channel to luminal calcium (Ca 2+ ). Methods and Results: Functional characterizations of recombinant human RyR2-G230C channels were performed under conditions mimicking stress. Human RyR2 mutant channels were generated by site-directed mutagenesis and heterologously expressed in HEK293 cells together with calstabin2. RyR2 channels were measured to examine the regulation of the channels by cytosolic versus luminal sarcoplasmic reticulum Ca 2+ . A 50-year-old white man with repeated syncopal episodes after exercise had a cardiac arrest and harbored the mutation RyR2-G230C. cAMP-dependent protein kinase–phosphorylated RyR2-G230C channels exhibited a significantly higher open probability at diastolic Ca 2+ concentrations, associated with a depletion of calstabin2. The luminal Ca 2+ sensitivities of RyR2-G230C and RyR2-P2328S channels were WT-like. Conclusions: The RyR2-G230C mutant exhibits similar biophysical defects compared with previously characterized CPVT mutations: decreased binding of the stabilizing subunit calstabin2 and a leftward shift in the Ca 2+ dependence for activation under conditions that simulate exercise, consistent with a “leaky” channel. Both RyR2-G230C and RyR2-P2328S channels exhibit normal luminal Ca 2+ activation. Thus, diastolic sarcoplasmic reticulum Ca 2+ leak caused by reduced calstabin2 binding and a leftward shift in the Ca 2+ dependence for activation by diastolic levels of cytosolic Ca 2+ is a common mechanism underlying CPVT.
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