The lncRNA HOTAIR impacts on mesenchymal stem cellsviatriple helix formation

Abstract

There is a growing perception that long non-coding RNAs (lncRNAs) modulate cellular function. In this study, we analyzed the role of the lncRNA HOTAIR in mesenchymal stem cells (MSCs) with particular focus on senescence-associated changes in gene expression and DNA-methylation (DNAm). HOTAIR binding sites were enriched at genomic regions that become hypermethylated with increasing cell culture passage. Overexpression and knockdown of HOTAIR inhibited or stimulated adipogenic differentiation of MSCs, respectively. Modification of HOTAIR expression evoked only very moderate effects on gene expression, particularly of polycomb group target genes. Furthermore, overexpression and knockdown of HOTAIR resulted in DNAm changes at HOTAIR binding sites. Five potential triple helix forming domains were predicted within the HOTAIR sequence based on reverse Hoogsteen hydrogen bonds. Notably, the predicted triple helix target sites for these HOTAIR domains were also enriched in differentially expressed genes and close to DNAm changes upon modulation of HOTAIR. Electrophoretic mobility shift assays provided further evidence that HOTAIR domains form RNA–DNA–DNA triplexes with predicted target sites. Our results demonstrate that HOTAIR impacts on differentiation of MSCs and that it is associated with senescence-associated DNAm. Targeting of epigenetic modifiers to relevant loci in the genome may involve triple helix formation with HOTAIR.

Super-lncRNAs: identification of lncRNAs that target super-enhancers via RNA:DNA:DNA triplex formation

“…Separate studies on lncRNAs (such as Fendrr and Tug1) have indicated that they localize at various genomic regions by forming a RNA:DNA:DNA triplex. This conclusion was further validated by a matching result between genomewide triplexator's prediction of triplex helix DNA target sites by lncRNA HOTAIR and the peaks obtained using the CHIRP-seq experiment of HOTAIR (Kalwa et al 2016).…”

“…triplexator (Kalwa et al 2016). Our prediction of superlncRNAs binding sites is based on these preliminary studies that suggested that lncRNAs may bind to DNA via RNA: DNA:DNA triplex formation.…”

“…Our prediction of superlncRNAs binding sites is based on these preliminary studies that suggested that lncRNAs may bind to DNA via RNA: DNA:DNA triplex formation. To further cross-validate these claims, we first focused on studies where the interactions between lncRNA and DNA sites were experimentally validated (Hotair, Kalwa et al 2016;Tug1, Long et al 2016;Fendrr, Grote et al 2013;Meg3, Mondal et al 2015). We performed a prediction of the triplex-forming sites of the specific lncRNAs (Hotair, Fendrr, Meg3, and Tug1) involved in these previous studies following our strategy.…”

Long noncoding RNA HOTAIR is upregulated in an aggressive subgroup of gastrointestinal stromal tumors (GIST) and mediates the establishment of gene‐specific DNA methylation patterns

“…[10][11][12] A prominent example of an epigenetic-related long noncoding transcript is HOX transcript antisense RNA (HOTAIR) that exhibits an enormous regulatory potential in cells. 14 Nevertheless, although the role of HOTAIR in the establishment of histone methylation patterns has been well documented in numerous tumor entities, 11,13,[15][16][17][18][19] its possible involvement in the onset of aberrant DNA methylation is still poorly understood. It does so by serving as a molecular scaffold for Polycomb repressive complex 2 (PRC2) and the histone demethylase LSD1/CoR-EST/REST 13 and recruiting them to particular genomic loci likely via triple helix formation.…”

“…It does so by serving as a molecular scaffold for Polycomb repressive complex 2 (PRC2) and the histone demethylase LSD1/CoR-EST/REST 13 and recruiting them to particular genomic loci likely via triple helix formation. 14 Nevertheless, although the role of HOTAIR in the establishment of histone methylation patterns has been well documented in numerous tumor entities, 11,13,[15][16][17][18][19] its possible involvement in the onset of aberrant DNA methylation is still poorly understood. Only few experimental studies have reported on the principle possibility of HOTAIRmediated aberrations of DNA methylation for several candidate genes in pathology.…”