B10: Decoding the targets of CVD-relevant IncRNAs and genetic deciphering of their mechanisms of action in vivo

The X-linked Firre locus produces a conserved trans-acting lncRNA involved in multiple biological processes, and an antisense lncRNA Crossfirre, which we discovered as the only imprinted gene on the X chromosome. Moreover, the Firre locus forms a conserved long-range interaction with an uncharacterized lncRNA locus Dxz4. To investigate the function of Firre and its potential interaction partners in the heart, we generated mice lacking Crossfirre, Firre, and Dxz4 individually and in combination. By assessing the transcriptome of these mutants, we found that deletion of the Firre locus results in the upregulation of mitochondrial and ribosomal pathways in multiple organs, including the heart. To directly test the role of these lncRNA loci in cardiovascular disease (CVD), we subjected our triple knockout (ΔTKO) mutants to the established myocardial infarction (MI) model and observed significant improvement of left ventricular ejection fraction. Furthermore, we observed downregulation of Firre in cardiac fibroblasts and leukocytes post-MI, while Crossfirre and Dxz4 were unaffected. Building on these findings, we will perform a Firre loss- and gain-of-function or rescue experiment and perform an extensive transcriptomic and phenotypic analysis post-MI. To gain mechanistic insights into how Firre controls its targets in trans, we will evaluate triplex-forming potential nearby the identified disease-specific Firre targets. To identify additional CVD-relevant lncRNA candidates, we developed a strategy that utilizes allele-specific expression to link allele-specific lncRNAs to nearby allelic targets. We implemented this concept into a bioinformatic tool termed Allelome.LINK, analysed adult mouse hearts, and found that 128 cardiac lncRNAs can be linked to their neighbouring putative target genes. To identify lncRNAs relevant for cardiovascular malformations, we overlapped the identified lncRNA-to-target links with the CVD-GWAS catalogue, resulting in 43 disease-relevant candidates. We will extend this analysis in mouse disease models at cardiac cell-type resolution and by exploring a large cohort of human left ventricle data. Considering the lncRNA-to-target links can be successfully validated, this strategy will elucidate CVD-relevant lncRNAs and their mechanism of action, bringing us closer to understanding the impact of the non-coding genome on heart disease.