In-vivo Function and Mechanism of the Long non-coding RNA Locus Handsdown

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Heart malfunction is one of the major causes of morbidity and lethality in modern society and nearly 1 in 100 newborns exhibit developmental malformations of the heart. In addition, heart injuries such as myocardial infarction impose an enormous societal and economic burden as they often result in maladaptive heart remodeling leading to congestive heart failure. While the heart has very limited regenerative potential, numerous efforts are currently undertaken to enhance regeneration or transplant cells with regenerative potential to patients. However, an in-depth knowledge of how the heart develops under normal circumstances is crucial for the development of effective regenerative therapies for heart disease.The heart is the first organ to develop during embryogenesis, containing functional cells in the form of contractile cardiomyocytes from very early on. Shortly after birth, the heart of both mice and humans has the temporally-restricted potential to regenerate injured heart muscle tissue into fully functional contractile tissue. While the role of many protein-coding RNAs in heart development has been well-studied, the class of long non-coding RNAs (lncRNAs) and their role in heart development and disease has only recently shifted into focus. Actually, the most abundant RNA transcripts in the vertebrate transcriptome are long noncoding RNAs (lncRNAs), transcripts that are longer than 200 nucleotides and do not encode proteins. LncRNAs have been associated with epigenetic changes, developmental defects and cancer, and knockouts are sometimes lethal. We showed previously that the lncRNA Fendrr interacts with the Polycomb repressive complex 2 (PRC2) and thereby regulates essential genes in the early embryonic cardiac cell lineage. We identified several novel lncRNAs with specific expression in the early embryonic heart and show that one particular locus is essential for embryonic development. To that end, we deleted the lncRNA locus in mouse embryonic stem cells (ESCs) using CRISPR/Cas9 genome editing and generated mouse embryos from these ESCs. The resulting embryos are embryonic lethal with, amongst other defects, mis-development of the heart. We propose here to analyze this novel heart specific lncRNA locus in vivo using state-of the art genomic tools and the generation of embryos and mice directly from genetically engineered stem cells. We will furthermore define the molecular mechanism of this lncRNA locus with respect to the RNA transcripts generated from it or the underlying genomic context.By understanding how lncRNAs function during normal heart development, we are likely to define new lncRNAs that may be valid targets to reactivate regenerative potential in adult hearts or enhance the regenerative potential of transplanted cells. Furthermore, successful downregulation of the lncRNA transcript by antisense RNA technology (GapmerR) might allow for the upregulation of heart specific genes, beneficial for cardiac repair after injury.