A transposable element into the human long noncoding RNA CARMEN is a switch for cardiac precursor cell specification.

AIMS The major cardiac cell types composing the adult heart arise from common multipotent precursor cells. Cardiac lineage decisions are guided by extrinsic and cell-autonomous factors, including recently discovered long noncoding RNAs (lncRNAs). The human lncRNA CARMEN, which is known to dictate specification towards the cardiomyocyte (CM) and the smooth muscle cell (SMC) fates, generates a diversity of alternatively spliced isoforms. METHODS AND RESULTS The CARMEN locus can be manipulated to direct human primary cardiac precursor cells (CPCs) into specific cardiovascular fates. Investigating CARMEN isoform usage in differentiating CPCs represents therefore a unique opportunity to uncover isoform-specific function in lncRNAs. Here, we identify one CARMEN isoform, CARMEN-201, to be crucial for SMC commitment. CARMEN-201 activity is encoded within an alternatively-spliced exon containing a MIRc short interspersed nuclear element. This element binds the transcriptional repressor REST (RE1 Silencing Transcription Factor), targets it to cardiogenic loci, including ISL1, IRX1, IRX5, and SFRP1, and thereby blocks the CM gene program. In turn, genes regulating SMC differentiation are induced. CONCLUSIONS These data show how a critical physiological switch is wired by alternative splicing and functional transposable elements in a long noncoding RNA. They further demonstrated the crucial importance of the lncRNA isoform CARMEN-201 in SMC specification during heart development

Alternative splicing of a transposable element into the human long noncoding RNA CARMEN is a switch for cardiac precursor cell specification

International audienceIntroductionDevelopmental long noncoding RNAs (lncRNAs) emerge as key regulators of specification in the cardiovascular system. In this context, alternative transcription start sites and terminations as well as alternative splicing expand the diversity of lncRNAs.ObjectivesWe take advantage of primary cardiac precursor cells (CPCs) isolated from the fetal and adult human heart to investigate the relevance of lncRNA isoforms during cardiovascular lineage commitment.Materials and methodsThe locus encoding the enhancer-associated lncRNA CARMN plays crucial roles in dictating specification into the cardiomyocyte (CM) and the smooth muscle cell (SMC) lineage. We use therefore an integrated approach combining transcriptomic and proteomic analyses to dissect the molecular mechanisms mediated by specific CARMN isoforms.ResultsWe identify CARMN-201 as crucial for SMC specification. Mechanistically, CARMN-201 associates with the transcriptional repressor REST (RE1 Silencing Transcription Factor aka Neuron Restrictive Silencer Factor), targets the repressor to cardiogenic loci, namely IRX1, IRX5, SFRP1 and ISL1, and blocks the emergence of a CM gene program. Binding to REST depends on the inclusion of the second exon in the CARMN-201 transcript. This exon contains itself a short interspersed nuclear element of the MIR family of transposable element indispensable for the capacity of the isoform to determine the SMC fate. Furthermore, two other CARMN-201 protein partners have been identified, i.e. the RNA methyltransferase NOP2/Sun RNA Methyltransferase 6 (NSUN6), and the Replication Protein A1 (RPA1), a protein implicated in stabilization of single-stranded DNA, suggesting CARMN-201 represses loci via forming RNA:DNA triplex at target promoters. In contrast, in CPCs adopting a CM fate, CARMN-201 is not expressed, allowing cardiac factors to be expressed and enabling cardiogenic specification and differentiation.ConclusionThese data show how a critical physiological switch controlling cardiac precursor specification is wired by alternative splicing and a functional transposable element in a long noncoding RNA

A transposable element into the human long noncoding RNA CARMEN is a switch for cardiac precursor cell specification

International audienceAbstract Aims The major cardiac cell types composing the adult heart arise from common multipotent precursor cells. Cardiac lineage decisions are guided by extrinsic and cell-autonomous factors, including recently discovered long noncoding RNAs (lncRNAs). The human lncRNA CARMEN, which is known to dictate specification toward the cardiomyocyte (CM) and the smooth muscle cell (SMC) fates, generates a diversity of alternatively spliced isoforms. Methods and results The CARMEN locus can be manipulated to direct human primary cardiac precursor cells (CPCs) into specific cardiovascular fates. Investigating CARMEN isoform usage in differentiating CPCs represents therefore a unique opportunity to uncover isoform-specific functions in lncRNAs. Here, we identify one CARMEN isoform, CARMEN-201, to be crucial for SMC commitment. CARMEN-201 activity is encoded within an alternatively spliced exon containing a MIRc short interspersed nuclear element. This element binds the transcriptional repressor REST (RE1 Silencing Transcription Factor), targets it to cardiogenic loci, including ISL1, IRX1, IRX5, and SFRP1, and thereby blocks the CM gene program. In turn, genes regulating SMC differentiation are induced. Conclusions These data show how a critical physiological switch is wired by alternative splicing and functional transposable elements in a long noncoding RNA. They further demonstrated the crucial importance of the lncRNA isoform CARMEN-201 in SMC specification during heart development