Abstract P3054: Further Delineation And Exploration Of The Cardiac MicroRNA-Target Interactome

Bailey Peck; Jared M McLendon; Ryan L Boudreau

doi:10.1161/res.133.suppl_1.P3054

Abstract only MicroRNAs (miRs) have emerged as key players in cardiac biology and disease. These non-coding RNAs are loaded into Argonaute (Ago) proteins to direct suppression of target mRNAs via base-pairing. Despite a mass of research on cardiac miRs in model systems and humans, the translational impacts have been limited since miR-mRNA target interactions in heart tissues have not been adequately defined at the “-omics” level. Researchers heavily rely on predicted miR targets and are often misled by false predictions, since imperfect base-pairing is sufficient for miR binding. Our goal is to comprehensively define miR targeting events and their biological relevance in the heart and to understand the clinical significance of genetic variants (e.g. SNPs) that modulate cardiac miR functions. We previously used high-throughput biochemical means (Ago2 CLIP-seq) to generate the first transcriptome-wide map of miR binding sites in non-failing human hearts, yielding ~4000 sites across >2000 genes. While exploring this dataset, we discovered translationally relevant miR-SNP interactions that associate with heart failure patient outcomes, most notably in SCN5A . We have since identified multiple common missense variants in TTN that disrupt miR binding. These associate with heart structure/function measures in the general population and could have significant implications for cardiomyopathy patients with TTN mutations. Beyond SNPs, we made the surprising discovery that TTN mRNA harbors >20 miR-133 binding sites and has the potential to bind up to 20% of the miR-133 pool in cardiomyocytes, perhaps serving as an endogenous miR “sponge”. We experimentally validated the potential for this using a TTN mini-gene harboring 18 of the most strongly bound miR-133 target sites; we found that this construct was capable of inhibiting miR-133 activity in cells. Finally, to further expand these studies, we optimized our protocol by adapting state-of-the-art CLIP methods, which recently yielded our 2 nd generation map of human cardiac miR binding sites (now in failing hearts), leading to the identification of >12,000 bindings sites across >6000 genes. We are now interrogating these data and processing additional samples to investigate how miR binding profiles rewire in failing hearts.

Abstract P3054: Further Delineation And Exploration Of The Cardiac MicroRNA-Target Interactome

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