PROJECT SUMMARY Cardiovascular disease (CVD) remains the leading cause of death, and there is a critical need to determine underlying mechanisms and develop novel therapies for treatment. Small Cajal body-specific RNAs (scaRNAs) are evolutionarily conserved non-coding RNAs that guide biochemical modification on specific nucleotides such as pseudouridylation (Ψ) and 2′-O-ribose methylation (2′-OMe). These modifications are essential in the post- transcriptional changes and functions of spliceosomal small nuclear RNAs (snRNA). Recent studies highlight a crucial role of scaRNA in preserving spliceosome fidelity and cardiac development; however, the role of scaRNAs in acute cardiac injury and its repair has not been established. This proposal aims to determine scaRNA’s role in cardiovascular processes in a mouse myocardial infarction (MI) model that could develop new tools for treating MI. Our central hypothesis is that overexpression of scaRNA18 in the post-MI heart promotes positive cardiac remodeling by inducing therapeutic angiogenesis via 2′-OMe of U4 snRNA and regulating Wilms tumor1 (WT1) expression, thereby conferring cardioprotection and improving cardiac function. Our preliminary data demonstrates 1) decreased scaRNA18 expression in post-MI mouse hearts using unbiased small RNA sequencing, 2) scaRNA18 expression exclusively downregulated in isolated cardiac endothelial cells (EC) in post-MI mouse hearts, 3) scaRNA18 knockdown (KD) impairs tube formation and induces apoptosis in mouse cardiac ECs without induced stress, 4) scaRNA18 overexpression enhances EC angiogenesis and inhibits stress-induced EC apoptosis in vitro, 5) scaRNA18-overexpressing cardiac EC extracellular vesicles protect iPSC-cardiomyocyte cell death via angiogenic proteins, 6) 2’-O-methylation levels were reduced in MI hearts and in cardiac ECs from scaRNA18 KD mice, 7) mutant scaRNA18 lacking binding sites to guide 2′-OMe on U4 snRNA induces EC cell apoptosis without induced stress, 8) unbiased mass spectrometry results revealed that scaRNA18 reduces WT1 expression levels in scaRNA18 KD cardiac ECs, 9) WT-1 KD in the presence of scaRNA18 overexpression nullified scaRNA18 mediated EC survival effects, and 10) overexpression of scaRNA18 by VE-cadherin-driven AAV9 reduces infarct size after ex vivo cardiac ischemia/reperfusion injury and improves cardiac function post-MI mice (in vivo). The hypothesis that scaRNA18 is a critical component in cardiac function and repair will be tested in the following three specific aims: in Aim 1, we will define the role of scaRNA18 in cardiac EC function and cardiomyocyte survival; in Aim 2, we will determine the molecular mechanisms by which scaRNA18 regulates cardiac EC function and cardiomyocyte survival, and in Aim 3 we will use post-MI mice to demonstrate the critical role of scaRNA18 in myocardial injury repair. The proposed studies could establish scaRNAs as novel therapeutic targets for MI.