Evf2 (Dlx6as) lncRNA regulates ultraconserved enhancer methylation and the differential transcriptional control of adjacent genes

Abstract

SUMMARYSeveral lines of evidence suggest that long non-coding RNA (lncRNA)-dependent mechanisms regulate transcription and CpG DNA methylation. Whereas CpG island methylation has been studied in detail, the significance of enhancer DNA methylation and its relationship with lncRNAs is relatively unexplored. Previous experiments proposed that the ultraconserved lncRNA Evf2 represses transcription through Dlx6 antisense (Dlx6as) transcription and methyl-CpG binding protein (MECP2) recruitment to the Dlx5/6 ultraconserved DNA regulatory enhancer (Dlx5/6ei) in embryonic day 13.5 medial ganglionic eminence (E13.5 MGE). Here, genetic epistasis experiments show that MECP2 transcriptional repression of Evf2 and Dlx5, but not Dlx6, occurs through antagonism of DLX1/2 in E13.5 MGE. Analysis of E13.5 MGE from mice lacking Evf2 and of partially rescued Evf2 transgenic mice shows that Evf2 prevents site-specific CpG DNA methylation of Dlx5/6ei in trans, without altering Dlx5/6 expression. Dlx1/2 loss increases CpG DNA methylation, whereas Mecp2 loss does not affect Dlx5/6ei methylation. Based on these studies, we propose a model in which Evf2 inhibits enhancer DNA methylation, effectively modulating competition between the DLX1/2 activator and MECP2 repressor. Evf2 antisense transcription and Evf2-dependent balanced recruitment of activator and repressor proteins enables differential transcriptional control of adjacent genes with shared DNA regulatory elements.

Long Noncoding RNAs in Atherosclerosis

“…Berghoff and his colleagues 27) found that lncRNA-Evf2 could repress Dlx5/6 gene expression by regulating Dlx5/6 enhancer site-specific methylation. Martens et al 28) found that lncRNA-SRG1 transcript across the SER3 promoter interferes with the binding of activators, thus regulating SER3 repression.…”

Evf2 lncRNA/BRG1/DLX1 interactions reveal RNA-dependent chromatin remodeling inhibition

“…Studies on Evf2, the first lncRNA member of larger classes of ucRNAs/t-UCRs, show that Evf2 (Dlx6os1, Dlx6as) is transcribed on the opposite strand, antisense to Dlx6, and therefore also belongs to the class of OS/NATs (Feng et al, 2006). Although antisense transcription predicted that Evf2 would have cis-regulatory effects, multiple experiments suggest that Evf2 exhibits both trans and cis effects (Feng et al, 2006;Bond et al, 2009;Berghoff et al, 2013). Chromatin immunoprecipitation (ChIP) experiments show that Evf2 increases the binding of transcriptional activators (DLX1/2; Zerucha et al, 2000) and the repressor methyl-CpG binding protein 2 (MECP2; Nan et al, 1997) to key Dlx5/6 enhancers (Zerucha et al, 2000) with a repressive outcome (Bond et al, 2009).…”

The functions of long noncoding RNAs in development and stem cells

“…Using this model, it was shown that transcription through the Evf2 locus controls the levels of Dlx6 in cis; after disengaging the polymerase, Evf2 then acts in trans to modulate methylation of the Dlx5/6 enhancer and transcription of Dlx5. Therefore, by regulating the cellular levels of the Dlx5 and Dlx6 TFs, Efv2 controls GABAergic interneuron activity (Berghoff et al, 2013;Bond et al, 2009). …”