Background Immunoreactive signal for the desmosomal protein plakoglobin (γ-catenin) is reduced at cardiac intercalated disks in patients with arrhythmogenic right ventricular cardiomyopathy (ARVC), a highly arrhythmogenic condition caused by mutations in genes encoding desmosomal proteins. Previously, we observed a “false positive” case in which plakoglobin signal was reduced in a patient initially thought to have ARVC but who actually had cardiac sarcoidosis. Sarcoidosis can masquerade clinically as ARVC, but has not previously been associated with altered desmosomal proteins. Methods and Results We observed marked reduction in immunoreactive signal for plakoglobin at cardiac myocyte junctions in patients with sarcoidosis and giant cell myocarditis, both highly arrhythmogenic forms of myocarditis associated with granulomatous inflammation. In contrast, plakoglobin signal was not depressed in lymphocytic (non-granulomatous) myocarditis. To determine whether cytokines might promote dislocation of plakoglobin from desmosomes, we incubated cultures of neonatal rat ventricular myocytes with selected inflammatory mediators. Brief exposure to low concentrations of IL-17, TNFα and IL-6, cytokines implicated in granulomatous myocarditis, caused translocation of plakoglobin from cell-cell junctions to intracellular sites, whereas other potent cytokines implicated in non-granulomatous myocarditis had no effect, even at much high concentrations. We also observed myocardial expression of IL-17 and TNFα, and elevated serum levels of inflammatory mediators including IL-6R, IL-8, MCP1 and MIP1β in ARVC patients (all p<0.0001 compared with controls). Conclusions These results suggest novel disease mechanisms involving desmosomal proteins in granulomatous myocarditis and implicate cytokines, perhaps derived in part from the myocardium, in disruption of desmosomal proteins and arrhythmogenesis in ARVC.

Background-Cardiac repolarization, the process by which cardiomyocytes return to their resting potential after each beat, is a highly regulated process that is critical for heart rhythm stability. Perturbations of cardiac repolarization increase the risk for life-threatening arrhythmias and sudden cardiac death. Although genetic studies of familial long-QT syndromes have uncovered several key genes in cardiac repolarization, the major heritable contribution to this trait remains unexplained. Identification of additional genes may lead to a better understanding of the underlying biology, aid in identification of patients at risk for sudden death, and potentially enable new treatments for susceptible individuals. Methods and Results-We extended and refined a zebrafish model of cardiac repolarization by using fluorescent reporters of transmembrane potential. We then conducted a drug-sensitized genetic screen in zebrafish, identifying 15 genes, including GINS3, that affect cardiac repolarization. Testing these genes for human relevance in 2 concurrently completed genome-wide association studies revealed that the human GINS3 ortholog is located in the 16q21 locus, which is strongly associated with QT interval. Conclusions-This sensitized zebrafish screen identified 15 novel myocardial repolarization genes. Among these genes isGINS3, the human ortholog of which is a major locus in 2 concurrent human genome-wide association studies of QT interval. These results reveal a novel network of genes that regulate cardiac repolarization. (Circulation. 2009;120:553-559.) Key Words: electrophysiology Ⅲ genes Ⅲ ion channels Ⅲ myocardial repolarization T he ECG QT interval duration is a predictor of mortality in familial long QT (LQT) syndromes 1 and in a wide range of acquired heart diseases. 2,3 QT prolongation by drugs can lead to fatal arrhythmias and has been a major cause of the withdrawal of medications from the market in the last decade. 4 Virtually all drugs that cause this adverse effect inhibit the rapid component of the delayed rectifier current, I Kr . 5 Clinical Perspective on p 559The QT interval, a summation of individual cellular action potential (AP) durations (APDs), is known to depend on the function of multiple ion channels and their accessory proteins. Much of our current understanding of cardiac repolarization comes from study of LQT syndromes. 6,7 These disorders are marked by abnormal cardiac repolarization and a high incidence of sudden death. 8 Although the majority of LQT disorders result from mutation of ion channel genes, there is increasing recognition of the role of non-ion channel mechanisms such as ankyrin B 9 and the caveolar scaffolding protein caveolin 3. 10 The zebrafish has been shown to recapitulate much of the complexity of higher vertebrates 11 yet retains the potential for exploring integrative physiology on a genomic scale. 12,13 Here, we studied cardiac repolarization as a complex trait using a functional screen to identify novel genetic determinants. Received September 11, 2008; accept...

The gating modifier toxins are a large family of protein toxins that modify either activation or inactivation of voltage-gated ion channels. ω-Aga-IVA is a gating modifier toxin from spider venom that inhibits voltage-gated Ca2+ channels by shifting activation to more depolarized voltages. We identified two Glu residues near the COOH-terminal edge of S3 in the α1A Ca2+ channel (one in repeat I and the other in repeat IV) that align with Glu residues previously implicated in forming the binding sites for gating modifier toxins on K+ and Na+ channels. We found that mutation of the Glu residue in repeat I of the Ca2+ channel had no significant effect on inhibition by ω-Aga-IVA, whereas the equivalent mutation of the Glu in repeat IV disrupted inhibition by the toxin. These results suggest that the COOH-terminal end of S3 within repeat IV contributes to forming a receptor for ω-Aga-IVA. The strong predictive value of previous mapping studies for K+ and Na+ channel toxins argues for a conserved binding motif for gating modifier toxins within the voltage-sensing domains of voltage-gated ion channels.