Catecholaminergic polymorphic ventricular tachycardia (CPVT) is a familial 
arrhythmogenic disorder associated with mutations in the cardiac ryanodine 
receptor (RyR2) and cardiac calsequestrin (CASQ2) genes. Previous in vitro 
studies suggested that RyR2 and CASQ2 interact as parts of a multimolecular 
Ca(2+)-signaling complex; however, direct evidence for such interactions and 
their potential significance to myocardial function remain to be determined. We 
identified a novel CASQ2 mutation in a young female with a structurally normal 
heart and unexplained syncopal episodes. This mutation results in the 
nonconservative substitution of glutamine for arginine at amino acid 33 of CASQ2 
(R33Q). Adenoviral-mediated expression of CASQ2(R33Q) in adult rat myocytes led 
to an increase in excitation-contraction coupling gain and to more frequent 
occurrences of spontaneous propagating (Ca2+ waves) and local Ca2+ signals 
(sparks) with respect to control cells expressing wild-type CASQ2 (CASQ2WT). As 
revealed by a Ca2+ indicator entrapped inside the sarcoplasmic reticulum (SR) of 
permeabilized myocytes, the increased occurrence of spontaneous Ca2+ sparks and 
waves was associated with a dramatic decrease in intra-SR [Ca2+]. Recombinant 
CASQ2WT and CASQ2R33Q exhibited similar Ca(2+)-binding capacities in vitro; 
however, the mutant protein lacked the ability of its WT counterpart to inhibit 
RyR2 activity at low luminal [Ca2+] in planar lipid bilayers. We conclude that 
the R33Q mutation disrupts interactions of CASQ2 with the RyR2 channel complex 
and impairs regulation of RyR2 by luminal Ca2+. These results show that 
intracellular Ca2+ cycling in normal heart relies on an intricate interplay of 
CASQ2 with the proteins of the RyR2 channel complex and that disruption of these 
interactions can lead to cardiac arrhythmia.