Calsequestrin 2 (CASQ2) mutations increase expression of calreticulin and ryanodine receptors, causing catecholaminergic polymorphic ventricular tachycardia

Abstract

Catecholamine-induced polymorphic ventricular tachycardia (CPVT) is a familial disorder caused by cardiac ryanodine receptor type 2 (RyR2) or calsequestrin 2 (CASQ2) gene mutations. To define how CASQ2 mutations cause CPVT, we produced and studied mice carrying a human D307H missense mutation (CASQ 307/307 ) or a CASQ2-null mutation (CASQ ΔE9/ΔE9 ). Both CASQ2 mutations caused identical consequences. Young mutant mice had structurally normal hearts but stress-induced ventricular arrhythmias; aging produced cardiac hypertrophy and reduced contractile function. Mutant myocytes had reduced CASQ2 and increased calreticulin and RyR2 (with normal phosphorylated proportions) but unchanged calstabin levels, as well as reduced total sarcoplasmic reticulum (SR) Ca 2+ , prolonged Ca 2+ release, and delayed Ca 2+ reuptake. Stress further diminished Ca 2+ transients, elevated cytosolic Ca 2+ , and triggered frequent, spontaneous SR Ca 2+ release. Treatment with Mg 2+ , a RyR2 inhibitor, normalized myocyte Ca 2+ cycling and decreased CPVT in mutant mice, indicating RyR2 dysfunction was critical to mutant CASQ2 pathophysiology. We conclude that CPVT-causing CASQ2 missense mutations function as null alleles. In the absence of CASQ2, calreticulin, a fetal Ca 2+ -binding protein normally downregulated at birth, remains a prominent SR component. Adaptive changes to CASQ2 deficiency (increased posttranscriptional expression of calreticulin and RyR2) maintained electrical-mechanical coupling, but increased RyR2 leakiness, a paradoxical response further exacerbated by stress. The central role of RyR2 dysfunction in CASQ2 deficiency unifies the pathophysiologic mechanism underlying CPVT due to RyR2 or CASQ2 mutations and suggests a therapeutic approach for these inherited cardiac arrhythmias. IntroductionCatecholamine-induced polymorphic ventricular tachycardia (CPVT) is a familial arrhythmogenic disorder characterized by adrenergic-stimulated VT that can deteriorate into ventricular fibrillation to cause sudden cardiac death (1, 2). Recurrent syncope, seizures, and cardiovascular collapse are triggered by exercise or emotional stress (2, 3). Mutations in the cardiac sarcoplasmic reticulum (SR) Ca 2+ release channel, ryanodine receptor type 2 (RyR2), or in cardiac calsequestrin 2 (CASQ2), a SR Ca 2+ -storage protein, cause CPVT (2, 4). Despite considerable knowledge about the coupling of Ca 2+ release from the SR, activation of sarcomere contraction, and Ca 2+ reuptake into the SR during relaxation (reviewed in refs. 5, 6), the mechanisms by which CASQ2 mutations alter Ca 2+ handling or cause CPVT are incompletely understood. CASQ, the most abundant Ca 2+ -buffering protein in the lumen of striated muscle SR (7,8), is encoded by the skeletal (CASQ1) and cardiac (CASQ2) calsequestrin genes (9, 10). Each molecule of CASQ2, a 415-amino acid polypeptide, binds 40-50 Ca 2+ with low affinity (K d = 1 mM), allowing cardiac SR Ca 2+ concentrations to approach 20 mM while free Ca 2+ concentrations are only 1 mM (7,(11)(12)(13). CASQ2 also part...

Facilitated maturation of Ca2+ handling properties of human embryonic stem cell-derived cardiomyocytes by calsequestrin expression

“…Furthermore, neither cellular hypertrophy nor t-tubule biogenesis was observed after Ad-CSQ transduction. Developmentally, immature CMs are known to express significant levels of calreticulin; calreticulin decreases after birth due to posttranscriptional modification and is subsequently replaced by CSQ during SR maturation (21,22,31). Our results show that Ad-CSQ transduction also did not affect calreticulin.…”

“…By contrast, CSQ2 deletion causes a striking increase in the SR volume as a compensatory response to maintain an operable SR Ca 2ϩ storage (15). As a result, premature spontaneous SR Ca 2ϩ release and subsequently, arrhythmias occur due to the relieved inhibition of RyRs by CSQ (15,31). Therefore, whereas the experimental approaches are complementary in their overall goal to obtain a better understanding of CSQ in cardiac physiology, ours differs by design.…”

Altered Ca2+ sparks in aging skeletal and cardiac muscle

“…One such protein is RyR2, where specific mutations result in a leaky RyR Ca 2+ release channel that can result in arrhythmiagenesis under stress conditions (Jiang et al, 2002;George et al, 2003;Lehnart et al, 2004;Kannankeril et al, 2006;Liu et al, 2006). Another series of mutations in calsequestrin (CSQ2) have been shown to induce leaky Ca 2+ release from the SR that is mediated at least in part by induction of increased RyR2 expression (Terentyev et al, 2003;di Barletta et al, 2006;Terentyev et al, 2006;Dirksen et al, 2007;Song et al, 2007). The role of CSQ in control of SR Ca 2+ release is emphasized by findings that overexpression of CSQ results in reduced Ca 2+ spark frequency and co-ordination, mirroring the effect of CSQ mutations on SR Ca 2+ release .…”

Chain-reaction Ca2+ signaling in the heart

“…Ordered Ca 2+ cycling is essential to normal rhythmic activity of the heart, and disturbances in Ca 2+ handling have previously been shown to underlie diverse Ca 2+ -dependent cardiac arrhythmias (7)(8)(9). In this issue of the JCI, catecholamine-induced polymorphic ventricular tachycardia (CPVT) caused by mutations in calsequestrin 2 (CASQ2) is the subject of the study by Song et al (10); Ca 2+ signaling plays a central role in the dysfunction.…”