BACKGROUND

Vascular homeostasis is maintained by the differentiated phenotype of vascular smooth muscle cells (VSMCs). The landscape of protein coding genes comprising the transcriptome of differentiated VSMCs has been intensively investigated but many gaps remain including the emerging roles of noncoding genes.

METHODS

We reanalyzed large-scale, publicly available bulk and single-cell RNA sequencing datasets from multiple tissues and cell types to identify VSMC-enriched long noncoding RNAs. The in vivo expression pattern of a novel smooth muscle cell (SMC)-expressed long noncoding RNA, (cardiac mesoderm enhancer-associated noncoding RNA), was investigated using a novel green fluorescent protein knock-in reporter mouse model. Bioinformatics and quantitative real-time polymerase chain reaction analysis were used to assess expression changes during VSMC phenotypic modulation in human and murine vascular disease models. In vitro, functional assays were performed by knocking down with antisense oligonucleotides and overexpressing by adenovirus in human coronary artery SMCs. Carotid artery injury was performed in SMC-specific knockout mice to assess neointima formation and the therapeutic potential of reversing loss was tested in a rat carotid artery balloon injury model. The molecular mechanisms underlying function were investigated using RNA pull-down, RNA immunoprecipitation, and luciferase reporter assays.

RESULTS

We identified , which was initially annotated as the host gene of the cluster and recently reported to play a role in cardiac differentiation, as a highly abundant and conserved, SMC-specific long noncoding RNA. Analysis of the GFP knock-in mouse model confirmed that is transiently expressed in embryonic cardiomyocytes and thereafter becomes restricted to SMCs. We also found that is transcribed independently of . expression is dramatically decreased by vascular disease in humans and murine models and regulates the contractile phenotype of VSMCs in vitro. In vivo, SMC-specific deletion of significantly exacerbated, whereas overexpression of markedly attenuated, injury-induced neointima formation in mouse and rat, respectively. Mechanistically, we found that physically binds to the key transcriptional cofactor myocardin, facilitating its activity and thereby maintaining the contractile phenotype of VSMCs.

CONCLUSIONS

is an evolutionarily conserved SMC-specific long noncoding RNA with a previously unappreciated role in maintaining the contractile phenotype of VSMCs and is the first noncoding RNA discovered to interact with myocardin.