Control of Cardiac Growth by Ca2+Dependent Phosphorylation of Histones

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DESCRIPTION (provided by applicant): Calcium is central in the regulation of many physiological processes. In the heart, calcium signals are decoded by calcium/calmodulin-dependent protein kinases (CaMKs) that regulate muscle contraction, gene expression, cell growth and apoptosis. Understanding how calcium signals are transmitted to the nucleus to modulate various cellular activities remains an important question in human biology. We have recently made a novel discovery showing that CaMKII couples calcium signals to chromatin remodeling in cardiac cells. Preliminary results show that a specific nuclear isoform of CaMKII selectively phosphorylates histone H3 in vitro and in primary cardiomyocytes. Activation of CaMK activity by agonist treatment, which induces hypertrophic growth, increases histone H3 phosphorylation. Conversely specific elimination of nuclear CaMKII in cardiac cells significantly reduces histone H3 phosphorylation and inhibits cardiac growth. These exciting new results reveal for the first time that CaMK enzymes are linked to chromatin and lead to our current hypothesis that CaMK- dependent phosphorylation of histones controls cardiac growth and may play an important role in heart pathogenesis. The goal of this application is to demonstrate this novel function of CaMK in the heart. For this, we propose to test the following hypotheses derived from our preliminary results: 1) To demonstrate the functional role of CaMK-mediated chromatin remodeling in cardiac diseases. For this we will test whether changes in histone H3 phosphorylation is detected at fetal cardiac genes and whether in vitro transcription is altered from chromatin reconstituted with mutant histones. 2) To test whether histone H3 phosphorylation is implicated in other cardiac disorders, and to begin to elucidate the biochemical mechanisms of CaMK- mediated chromatin remodeling by investigating the various amino acids that are phosphorylated by nuclear CaMKII in histone H3. The comprehensive experimental approaches will include chromatin immunoprecipitation, manipulation of gene expression using recombinant adenovirus and RNA interference in primary cardiac cells, confocal microscopy, mass spectrometry and in vitro transcription assays. This study may lead to a new area of investigation on CaMKs in the cardiac field and possibly in other fields where CaMKs exert their functional role. Most importantly, it will lead to potential new therapautic strategies to treate cardiac diseases such as heart failure. PUBLIC HEALTH RELEVANCE: Calcium/calmodulin-dependent protein kinases (CaMKs) are enzymes that are present in all major organs and are involved in a variety of cardiac disorders. We made a new discovery showing that a specific isoform of CaMK enriched in cardiac nuclei, couples calcium signals to chromatin remodeling. The goal of this application is to demonstrate that this new function of CaMK is central to pathological events leading to cardiac hypertrophy and to other cardiac diseases. Thus, the proposed study has potential to open new avenues in the CaMK field and may bring novel therapeutic intervention for the treatment of heart disease.