Objective: Long-noncoding RNAs (lncRNAs) are potent regulators of the vascular system. The lncRNA HIF1a-AS1 is located on the antisense strand of the important Hypoxia-inducible factor 1a gene and is induced in aortic aneurysms. The function and the mode of action of the lncRNA are, however, unknown and were identified here.

Results: HIF1a-AS1 was reduced in endothelial cells isolated from glioblastoma and from lungs of patients with pulmonary arterial hypertension. In contrast, reoxygenation after hypoxia induced HIF1a-AS1. Functionally, knockdown of HIF1a-AS1 in endothelial cells increased their angiogenic capacity. HIF1a-AS1 localized to condensed nuclear regions and associated with double stranded DNA. Triplex-Sequencing, triplex domain finding and RNA immunoprecipitation indicated that HIF1a-AS1 forms DNA:DNA:RNA triplexes on specific repressive sites of the genome, among them the HIF1a and EPH receptor A2 (EPHA2) genes. Exchange of the triplex forming region of HIF1a-AS1 with other known triplex forming regions by CRISPR Arcitect abolished its effects on gene expression. Protein interaction studies revealed that HIF1a-AS1 interacts with the human silencing hub (HUSH) complex, which contains the epigenetic repressor MPP8. HIF1a-AS1 recruits the HUSH complex to sites of triplex formation to pro-angiogenic genes like EPHA2.

Conclusions: These results suggest that HIF1a-AS1 forms DNA:DNA:RNA triple helices in endothelial cells to recruit epigenetic silencer complexes. This mechanism is operative in vascular tissue and controls the expression of the highly important genes HIF1a and ephrin receptor A2. Through this mechanism, HIF1a-AS1 limits the aniogenic response.