Congenital heart defects (CHDs) are the most common congenital malformations. 5% of CHDs comprise atrioventricular septal defects (AVSDs). However, the molecular etiology underlying most AVSDs are not understood. Furthermore, CHDs can cause cardiovascular diseases later in life, resulting in arrhythmias, stroke, and premature death. In order to develop novel therapies able to prevent CHDs and heal specific cardiovascular tissues, it is critical to garner understanding of fundamental mechanisms directing normal cardiac chamber development and regeneration. Therefore, long-term goals of our lab are to understand conserved mechanisms that direct the development of the individual cardiac chambers and chamber-specific mechanisms of regeneration in vertebrates. Few signals are known to be required that specifically direct atrial development, with specific regulators of atrial regeneration not being understood.
The specific aims of this proposal are to elucidate the mechanisms by which a syntenic long non-coding RNA (lncRNA) as-oca restricts atrial development and regeneration through inhibition of Nr2f1a translation in zebrafish. The studies in this proposal are relevant to human health as recent genomic analysis indicates that mutations in the orphan nuclear receptor Nr2f2 are associated with AVSDs in humans. While Nr2f2 knockout mice and in vitro studies with human stem cells have revealed requirements for both Nr2f1 and Nr2f2 in atrial development, the mechanisms by which Nr2f proteins direct proper atrial development are not completely understood. Importantly, there is currently no understanding of epigenetic lncRNA-dependent mechanisms regulating Nr2f proteins. Our preliminary analysis of the novel zebrafish mutant acorn worm (aco) indicate that excess expression of as-oca specifically restricts the addition of later differentiating second heart field (SHF)-derived atrial cells.
In Aim 1, we will use blastula cell transplantation, in vivo cardiomyocyte differentiation assays, and genome editing to determine the cellular requirements underlying the atrial defects and cause of increased as- oca expression in aco mutants. We do not understand how as-oca inhibits Nr2f1a translation in aco mutants.
In Aim 2, we will use RNA and ribosomal association techniques and loss of function methods to determine if as- oca inhibits Nr2f1a translation through interactions with the nr2f1a 3' untranslated region. In addition to their requirements during development, we find that as-oca and nr2f1a are specifically expressed in the atria of adult zebrafish.
In Aim 3, we will test the effects of as-oca on embryonic and adult models of cardiac regeneration. Because Nr2f transcription factors play conserved roles in atrial development of all vertebrates, these studies will dramatically improve our understanding of posttranscriptional mechanisms regulating normal vertebrate atrial development and the molecular etiology of AVSDs and ASDs in humans. Ultimately, these studies will generate a foundation for improved therapies capable of preventing CHDs in children and efficiently repairing injured hearts in adults.
We currently do not understand the mechanisms that underlie the majority of congenital heart defects, which are the most common birth defects. This project will provide a greater understanding of the causes of congenital heart defects and normal heart development. Ultimately, the information gleaned will allow the generation of novel therapies able to prevent congenital heart defects and regenerate injured hearts in children and adults.
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|Duong, Tiffany B; Ravisankar, Padmapriyadarshini; Song, Yuntao Charlie et al. (2018) Nr2f1a balances atrial chamber and atrioventricular canal size via BMP signaling-independent and -dependent mechanisms. Dev Biol 434:7-14|