Cardiac repolarization, assessed in myocytes as action potential (AP) duration and on the electrocardiogram as the QT interval, requires the tightly regulated function of multiple ion channels and their accessory proteins. In the last decade, perturbations of repolarization have been directly implicated in the genesis of drug-induced arrhythmias and sudden cardiac death (SCD). While QT interval is a predictor of arrhythmias and sudden death, it is clear that a prolonged QT interval by itself is insufficient to cause such catastrophic events;additional environmental insults are usually required. There are several identified triggers including, myocardial ischemia, electrolyte imbalances (especially hypokalemia), and elevated catecholamine states, but perhaps the greatest environmental challenge to cardiac repolarization comes in the form of prescribed drugs. In the past decade, the single most common cause of the withdrawal or restriction of drugs that have already been marketed has been undesired QT prolongation. While the QT interval is a heritable quantitative trait, the genes that influence the QT interval as well as the response to QT prolonging drugs remain unknown. Common variants have long been thought to play a significant role in this complex trait and there are now several novel loci associated with QT interval through recent genome-wide association studies. Along with the power to identify novel loci, genome wide association studies do have limitations: they cannot distinguish which gene at a given locus is causal, nor do they reveal mechanistic insights. As we begin to unravel the discoveries of GWA studies, the first step will be to identify the functional gene(s) at each associated locus. We propose the use of a tractable zebrafish model that faithfully recapitulates key features of human myocardial repolarization. Using an approach that allows translation of human genetic discoveries into a tractable relevant model, we will test the hypothesis that gene knockdown in zebrafish will confirm candidate genes for each of five novel human repolarization loci. Once novel repolarization genes are identified, they will be tested for gene x drug interactions in our model. We propose the following specific aims:
Aim 1 : Validate candidate myocardial repolarization genes from five recently discovered novel genetic loci in a zebrafish model of cardiac repolarization. This will involve targeted knockdown of genes from these five loci and determination of effects on myocardial repolarization using optical voltage mapping.
Aim 2 : Quantitatively identify gene x drug interactions in myocardial repolarization. The most clinically important environmental exposures to myocardial repolarization are QT prolonging drugs. All 25 candidate repolarization genes will be tested for interactions in gene x drug experiments with QT prolonging drugs.
The QT interval on the electrocardiogram is a predictor of cardiac rhythm problems and sudden death, but the genes that govern this process remain largely unknown. Recently, large population-based studies have narrowed the search for these genes, identifying five genetic neighborhoods where these genes lie. We propose to use a zebrafish model to systematically knockdown the genes in these neighborhoods and measure the impact on the equivalent of the QT interval;we hope to conclusively identify the genes that modify the QT interval and achieve a better understanding of heart rhythm problems and sudden death.
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