S-EPMC7184162 - Long non-coding RNA CDKN2B-AS1 contributes to atherosclerotic plaque formation by forming RNA-DNA triplex in the CDKN2B promoter. - OmicsDI

Unknown

Dataset Information

Long non-coding RNA CDKN2B-AS1 contributes to atherosclerotic plaque formation by forming RNA-DNA triplex in the CDKN2B promoter.

ABSTRACT:

Background

Atherosclerosis involves a slow process of plaque formation on the walls of arteries, and comprises a leading cause of cardiovascular disease. Long non-coding RNAs (lncRNAs) have been implicated in the pathogenesis of atherosclerosis. In this study, we aim to explore the possible involvement of lncRNA 'cyclin-dependent kinase inhibitor 2B antisense noncoding RNA' (CDKN2B-AS1) and CDKN2B in the progression of atherosclerosis.

Methods

Initially, we quantified the expression of CDKN2B-AS1 in atherosclerotic plaque tissues and, in THP-1 macrophage-derived, and human primary macrophage (HPM)-derived foam cells. Next, we established a mouse model of atherosclerosis using apolipoprotein E knockout (ApoE^-/-) mice, where lipid uptake, lipid accumulation, and macrophage reverse cholesterol transport (mRCT) were assessed, in order to explore the contributory role of CDKN2B-AS1 to the progression of atherosclerosis. RIP and ChIP assays were used to identify interactions between CDKN2B-AS1, CCCTC-binding factor (CTCF), enhancer of zeste homologue 2 (EZH2), and CDKN2B. Triplex formation was determined by RNA-DNA pull-down and capture assay as well as EMSA experiment.

Findings

CDKN2B-AS1 showed high expression levels in atherosclerosis, whereas CDKN2B showed low expression levels. CDKN2B-AS1 accelerated lipid uptake and intracellular lipid accumulation whilst attenuating mRCT in THP-1 macrophage-derived foam cells, HPM-derived foam cells, and in the mouse model. EZH2 and CTCF were found to bind to the CDKN2B promoter region. An RNA-DNA triplex formed by CDKN2B-AS1 and CDKN2B promoter was found to recruit EZH2 and CTCF in the CDKN2B promoter region and consequently inhibit CDKN2B transcription by accelerating histone methylation.

Interpretation

The results demonstrated that CDKN2B-AS1 promotes atherosclerotic plaque formation and inhibits mRCT in atherosclerosis by regulating CDKN2B promoter, and thereby could be a potential therapeutic target for atherosclerosis.

SUBMITTER: Ou M

PROVIDER: S-EPMC7184162 | biostudies-literature | 2020 May

REPOSITORIES: biostudies-literature

altmetric image

Publications

Long non-coding RNA CDKN2B-AS1 contributes to atherosclerotic plaque formation by forming RNA-DNA triplex in the CDKN2B promoter.

Ou Minghui M Li Xia X Zhao Shibo S Cui Shichao S Tu Jie J

EBioMedicine 20200424

<h4>Background</h4>Atherosclerosis involves a slow process of plaque formation on the walls of arteries, and comprises a leading cause of cardiovascular disease. Long non-coding RNAs (lncRNAs) have been implicated in the pathogenesis of atherosclerosis. In this study, we aim to explore the possible involvement of lncRNA 'cyclin-dependent kinase inhibitor 2B antisense noncoding RNA' (CDKN2B-AS1) and CDKN2B in the progression of atherosclerosis.<h4>Methods</h4>Initially, we quantified the expressi ...[more]

Similar Datasets

Project description:The long noncoding RNA CDKN2B-AS1 harbors a major coronary artery disease risk haplotype, which is also associated with progressive forms of the oral inflammatory disease periodontitis as well as myocardial infarction (MI). Despite extensive research, there is currently no broad consensus on the function of CDKN2B-AS1 that would explain a common molecular role of this lncRNA in these diseases. Our aim was to investigate the role of CDKN2B-AS1 in gingival cells to better understand the molecular mechanisms underlying the increased risk of progressive periodontitis. We downregulated CDKN2B-AS1 transcript levels in primary gingival fibroblasts with LNA GapmeRs. Following RNA-sequencing, we performed differential expression, gene set enrichment analyses and Western Blotting. Putative causal alleles were searched by analyzing associated DNA sequence variants for changes of predicted transcription factor binding sites. We functionally characterized putative functional alleles using luciferase-reporter and antibody electrophoretic mobility shift assays in gingival fibroblasts and HeLa cells. Of all gene sets analysed, collagen biosynthesis was most significantly upregulated (Padj=9.7 × 10- 5 (AUC > 0.65) with the CAD and MI risk gene COL4A1 showing strongest upregulation of the enriched gene sets (Fold change = 12.13, Padj = 4.9 × 10- 25). The inflammatory "TNFA signaling via NFKB" gene set was downregulated the most (Padj=1 × 10- 5 (AUC = 0.60). On the single gene level, CAPNS2, involved in extracellular matrix organization, was the top upregulated protein coding gene (Fold change = 48.5, P < 9 × 10- 24). The risk variant rs10757278 altered a binding site of the pathogen responsive transcription factor STAT1 (P = 5.8 × 10- 6). rs10757278-G allele reduced STAT1 binding 14.4% and rs10757278-A decreased luciferase activity in gingival fibroblasts 41.2% (P = 0.0056), corresponding with GTEx data. CDKN2B-AS1 represses collagen gene expression in gingival fibroblasts. Dysregulated collagen biosynthesis through allele-specific CDKN2B-AS1 expression in response to inflammatory factors may affect collagen synthesis, and in consequence tissue barrier and atherosclerotic plaque stability.