Community Research and Development Information Service - CORDIS



Project ID: 623739
Funded under: FP7-PEOPLE
Country: Italy

Periodic Report Summary 1 - THE CARDIAC CODE (The role of DOT1L in regulating the epigenetic signature of cardiomyocytes during heart development)

Cardiovascular diseases are an extremely relevant socio-economical burden: congenital heart disease (CHD) is the most common form of inborn malformation, and adult heart disease is a major cause of morbidity and mortality in developed countries. Congenital and adult heart disease share a common link: regulation of cardiomyocyte (CM) proliferation. Impaired CM proliferation during embryogenesis is a frequent cause of defective cardiogenesis, whereas the reduced regenerative potential of the adult heart after injury can be explained by the fact that adult CMs lack proliferative potential. CMs switch from a proliferative state in embryos, to a non-replicative state shortly after birth. Understanding molecular and cellular mechanisms underlying this transition is critical for deciphering molecular origins of cardiac malformations and develop new therapies for cardiac regeneration in adult. Stage and cell-type-specific gene expression programs are established by the coordinated action of transcription factors and epigenetic enzymes. Epigenetic modifications are crucial for stabilizing transcriptional programs, but little is known about the “epigenetic code” of histone methylation during cardiogenesis. Previous studies in mouse embryonic stem cell-derived CMs suggested that the histone methyl transferase DOT1L is required for proper CM differentiation (Cattaneo et al. Cell Death Diff 2016). DOT1L is the sole enzyme that catalyzes methylation (mono, di or tri) at H3K79. Different degrees of H3K79me lead to distinct transcriptional readouts (gene activation, repression, or no changes of transcription) in a context dependent manner.
Following up on observations from these in vitro studies, the hypothesis to test with the “Cardiac Code” project was that DOT1L plays a key role in embryonic heart development, defining a H3K79 dependent epigenetic landscape essential for cardiomyocyte differentiation and cardiomyocyte cell cycle regulation.
To test this hypothesis, the following Specific Aims have been proposed: 1) To investigate effects of early cardiomyocyte-specific deletion of Dot1L on cardiac morphogenesis and cardiomyocyte cell biology in vivo.
2) To identify genes directly regulated, positively or negatively, by DOT1L mediated-H3K79 methylation in vivo.
Dr. Cattaneo used the xMLC2cre line to selectively ablate DOT1L in CMs from embryonic day E7.5 in mice and performed a detailed characterization of the cardiac phonotype. She observed that mutant pups die between postnatal day 2 and 10. Interestingly, this window of lethality corresponds to the period in which CMs withdraw from cell cycle. In the absence of DOT1L, mutants display enlarged hearts, increased CM proliferation and reduced binuclation, demonstrating an essential role for DOT1L in cardiogenesis and CM cell cycle regulation in vivo.
To determine the mechanistic basis of these cardiac phenotypes, Dr. Cattaneo collected purified CMs derived from xMLC2cre;Dot1L f/f mutant and control mice to determine, by RNA-seq, genes differentially expressed in the absence of DOT1L. In addition, she also collected samples to perform ChIP-seq studies to determine genomic loci containing peaks of H3K79me. During the return phase, datasets produced by such experiments will be analyzed and cross-compared to identify genes directly regulated by DOT1L in CMs and identify specific key target genes that are directly affected by the ablation of Dot1L. Such analyses will shed light into the mechanisms by which DOT1L regulates CM differentiation and proliferation and might help designing new diagnostic and/or therapeutic approaches for congenital and adult heart disease. The conclusion of the “Cardiac Code” project will significantly increase our knowledge of mechanisms by which DOT1L- mediated methylation is critical for heart development, and mechanisms underlying CM cell cycle regulation. Proposed studies will result in a coherent picture of the potential role of DOT1L and H3K79 methylation in CM cell cycle regulation and differentiation. This knowledge will help in designing regenerative therapies for the heart, and will also yield fundamental insights into how epigenetic mechanisms regulate gene expression in cardiomyocytes.

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Life Sciences
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