Unexpected Structural and Functional Consequences of the R33Q Homozygous Mutation in Cardiac Calsequestrin: A Complex Arrhythmogenic Cascade in a Knock In Mouse Model

Catecholaminergic polymorphic ventricular tachycardia (CPVT) is an inherited arrhythmogenic disorder characterized by life threatening arrhythmias elicited by physical and emotional stress in young individuals. The recessive form of CPVT is associated with mutation in the cardiac calsequestrin gene (CASQ2). We engineered and characterized a homozygous CASQ2R33Q/R33Q mouse model that closely mimics the clinical phenotype of CPVT patients. CASQ2R33Q/R33Q mice develop bidirectional VT on exposure to environmental stress whereas CASQ2R33Q/R33Q myocytes show reduction of the sarcoplasmic reticulum (SR) calcium content, adrenergically mediated delayed (DADs) and early (EADs) afterdepolarizations leading to triggered activity. Furthermore triadin, junctin, and CASQ2-R33Q proteins are significantly decreased in knock-in mice despite normal levels of mRNA, whereas the ryanodine receptor (RyR2), calreticulin, phospholamban, and SERCA2a-ATPase are not changed. Trypsin digestion studies show increased susceptibility to proteolysis of mutant CASQ2. Despite normal histology, CASQ2R33Q/R33Q hearts display ultrastructural changes such as disarray of junctional electron-dense material, referable to CASQ2 polymers, dilatation of junctional SR, yet normal total SR volume. Based on the foregoings, we propose that the phenotype of the CASQ2R33Q/R33Q CPVT mouse model is portrayed by an unexpected set of abnormalities including (1) reduced CASQ2 content, possibly attributable to increased degradation of CASQ2-R33Q, (2) reduction of SR calcium content, (3) dilatation of junctional SR, and (4) impaired clustering of mutant CASQ2.

[1]  S. Priori,et al.  Catecholaminergic polymorphic ventricular tachycardia-related mutations R33Q and L167H alter calcium sensitivity of human cardiac calsequestrin. , 2008, The Biochemical journal.

[2]  S. Priori,et al.  Luminal Ca2+ Regulation of Single Cardiac Ryanodine Receptors: Insights Provided by Calsequestrin and its Mutants , 2008, The Journal of general physiology.

[3]  José Jalife,et al.  Arrhythmogenic Mechanisms in a Mouse Model of Catecholaminergic Polymorphic Ventricular Tachycardia , 2007, Circulation research.

[4]  M. Varsányi,et al.  Characterization of human cardiac calsequestrin and its deleterious mutants. , 2007, Journal of molecular biology.

[5]  S. Priori,et al.  Abstract 910: Catecholaminergic Polymorphic Ventricular Tachycardia: Genetics, Natural History and Response to Therapy , 2007 .

[6]  Tao Yang,et al.  Modest Reductions of Cardiac Calsequestrin Increase Sarcoplasmic Reticulum Ca2+ Leak Independent of Luminal Ca2+ and Trigger Ventricular Arrhythmias in Mice , 2007, Circulation research.

[7]  J. Seidman,et al.  Calsequestrin 2 (CASQ2) mutations increase expression of calreticulin and ryanodine receptors, causing catecholaminergic polymorphic ventricular tachycardia. , 2007, The Journal of clinical investigation.

[8]  S. Cala,et al.  Different endoplasmic reticulum trafficking and processing pathways for calsequestrin (CSQ) and epitope-tagged CSQ. , 2006, Experimental cell research.

[9]  S. Priori,et al.  Clinical Phenotype and Functional Characterization of CASQ2 Mutations Associated With Catecholaminergic Polymorphic Ventricular Tachycardia , 2006, Circulation.

[10]  D. Roden,et al.  Casq2 deletion causes sarcoplasmic reticulum volume increase, premature Ca2+ release, and catecholaminergic polymorphic ventricular tachycardia. , 2006, The Journal of clinical investigation.

[11]  S. Priori,et al.  Arrhythmogenesis in Catecholaminergic Polymorphic Ventricular Tachycardia: Insights From a RyR2 R4496C Knock-In Mouse Model , 2006 .

[12]  S. Priori,et al.  Abnormal Interactions of Calsequestrin With the Ryanodine Receptor Calcium Release Channel Complex Linked to Exercise-Induced Sudden Cardiac Death , 2006, Circulation research.

[13]  Sara Negri,et al.  Arrhythmogenesis in catecholaminergic polymorphic ventricular tachycardia: insights from a RyR2 R4496C knock-in mouse model. , 2006, Circulation research.

[14]  S. Priori,et al.  Bidirectional Ventricular Tachycardia and Fibrillation Elicited in a Knock-In Mouse Model Carrier of a Mutation in the Cardiac Ryanodine Receptor , 2005, Circulation research.

[15]  I. Fraser,et al.  Navigating the signalling network in mouse cardiac myocytes , 2002, Nature.

[16]  A. Wilde,et al.  Absence of Calsequestrin 2 Causes Severe Forms of Catecholaminergic Polymorphic Ventricular Tachycardia , 2002, Circulation research.

[17]  Carlo Napolitano,et al.  Clinical and Molecular Characterization of Patients With Catecholaminergic Polymorphic Ventricular Tachycardia , 2002, Circulation.

[18]  D. Lancet,et al.  A missense mutation in a highly conserved region of CASQ2 is associated with autosomal recessive catecholamine-induced polymorphic ventricular tachycardia in Bedouin families from Israel. , 2001, Cold Spring Harbor symposia on quantitative biology.

[19]  L. Gepstein,et al.  Electroanatomic mapping of arrhythmogenic right ventricular dysplasia. , 2001, Journal of the American College of Cardiology.

[20]  R. Tibshirani,et al.  Significance analysis of microarrays applied to the ionizing radiation response , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[21]  S. Priori,et al.  Mutations in the Cardiac Ryanodine Receptor Gene (hRyR2) Underlie Catecholaminergic Polymorphic Ventricular Tachycardia , 2001, Circulation.

[22]  F. Protasi,et al.  Formation and maturation of the calcium release apparatus in developing and adult avian myocardium. , 1996, Developmental biology.

[23]  A. Varró,et al.  An estimate of the calcium content of the sarcoplasmic reticulum in rat ventricular myocytes , 1993, Pflügers Archiv.

[24]  P B Corr,et al.  Mechanisms underlying early and delayed afterdepolarizations induced by catecholamines. , 1990, The American journal of physiology.

[25]  Craig T. January,et al.  Early Afterdepolarizations: Mechanism of Induction and Block A Role for L‐Type Ca2+ Current , 1989, Circulation research.

[26]  B A Mobley,et al.  Sizes of components in frog skeletal muscle measured by methods of stereology , 1975, The Journal of general physiology.

[27]  U. K. Laemmli,et al.  Cleavage of Structural Proteins during the Assembly of the Head of Bacteriophage T4 , 1970, Nature.

[28]  D. Terentyev,et al.  Modulation of sarcoplasmic reticulum calcium release by calsequestrin in cardiac myocytes. , 2004, Biological research.