Abstract 14259: Novel Long Noncoding RNA, Ppp1r1b-LncRNA , Epigenetically Promotes Myogenic Differentiation Program in Cardiac and Skeletal Myocytes

Abstract

Introduction: Emerged as critical regulators of the transcriptome, the lncRNAs play important roles in cardiac development and might be targeted to treat human cardiomyocyte dysfunction in congenital heart disease. In our work, we identified a novel lncRNA, Ppp1r1b-lncRNA , as an essential, functionally conserved, positive regulator of myogenic differentiation. Hypothesis: Ppp1r1b-lncRNA promotes myogenic differentiation program of cardiac and skeletal myocytes via epigenic mechanisms. Methods: Antisense oligonucleotides (GapmeR) were used to suppress Ppp1r1b-lncRNA . Chromatin immunoprecipitation (CHIP) and chromatin isolation by RNA purification (CHIRP) were used to analyze gene specific histone modification level and lncRNA: chromatin interaction, respectively. RNA pull-down and RNA immunoprecipitation (RIP) were used to identify Ppp1r1b-lncRNA : protein interactome. Results: By silencing Ppp1r1b-lncRNA, skeletal myoblasts and human-induced pluripotent stem cell derived cardiomyocytes (hiPSCs-CMs) failed to develop fully differentiated myotubes. Analysis of GapmeR injected neonatal mouse heart further confirmed the role of Ppp1r1b-lncRNA in normal myogenesis. Members of muscle specific transcription factors, including MyoD1 and TBX5, were not induced during myogenic differentiation in the Ppp1r1b-lncRNA depleted cardiac and skeletal muscle precursor cells. Histone modification analysis revealed enrichment of H3K27 tri-methylation at MyoD1 and TBX5 promoters in GapmeR treated skeletal myoblasts. Furthermore, Ppp1r1b-lncRNA was found to bind to Ezh2, and ChIRP analysis revealed enriched interaction of Ppp1r1b-lncRNA with Myod1 and Tbx5 promoters, suggesting that Ppp1r1b-lncRNA induces transcription of myogenic regulatory factors by interacting with the polycomb repressive complex 2 (PRC2) at the chromatin interface. Correspondingly, the silencing of Ppp1r1b-lncRNA increased EZH2 binding at promoter regions of myogenic transcription factors. Conclusions: Ppp1r1b-lncRNA promotes myogenic differentiation by inhibiting histone methylation on the promotors of myogenic master regulators via competing for PRC2 binding with chromatin during heart and skeletal muscle development.