CAD increases the long noncoding RNA PUNISHER in small extracellular vesicles and regulates endothelial cell function via vesicular shuttling

• Main Authors: Mohammed Rabiul Hosen, Qian Li, Yangyang Liu, Andreas Zietzer, Katharina Maus, Philip Goody, Shizuka Uchida, Eicke Latz, Nikos Werner, Georg Nickenig, Felix Jansen
• Format: Article
• Language: English
• Published: Elsevier 2021-09-01
• Series: Molecular Therapy: Nucleic Acids
• Subjects: coronary artery disease; extracellular vesicles; long noncoding RNA; angiogenesis; cardiovascular disease
• Online Access: http://www.sciencedirect.com/science/article/pii/S2162253121001414

Long noncoding RNAs (lncRNAs) have emerged as biomarkers and regulators of cardiovascular disease. However, the expression pattern of circulating extracellular vesicle (EV)-incorporated lncRNAs in patients with coronary artery disease (CAD) is still poorly investigated. A human lncRNA array revealed that certain EV-lncRNAs are significantly dysregulated in CAD patients. Circulating small EVs (sEVs) from patients with (n = 30) or without (n = 30) CAD were used to quantify PUNISHER (also known as AGAP2-antisense RNA 1 [AS1]), GAS5, MALAT1, and H19 RNA levels. PUNISHER (p = 0.002) and GAS5 (p = 0.02) were significantly increased in patients with CAD, compared to non-CAD patients. Fluorescent labeling and quantitative real-time PCR of sEVs demonstrated that functional PUNISHER was transported into the recipient cells. Mechanistically, the RNA-binding protein, heterogeneous nuclear ribonucleoprotein K (hnRNPK), interacts with PUNISHER, regulating its loading into sEVs. Knockdown of PUNISHER abrogated the EV-mediated effects on endothelial cell (EC) migration, proliferation, tube formation, and sprouting. Angiogenesis-related gene profiling showed that the expression of vascular endothelial growth factor A (VEGFA) RNA was significantly increased in EV recipient cells. Protein stability and RNA immunoprecipitation indicated that the PUNISHER-hnRNPK axis regulates the stability and binding of VEGFA mRNA to hnRNPK. Loss of PUNISHER in EVs abolished the EV-mediated promotion of VEGFA gene and protein expression. Intercellular transfer of EV-incorporated PUNISHER promotes a pro-angiogenic phenotype via a VEGFA-dependent mechanism.