The Primary Transcript of Microrna-208b Modulates Gene Expression in Cardiac Hypertrophy by Chromatin Interaction: Core 3. Genetics, Genomics and Congenital CV Disorders, Session Title: Epigenetic and Noncoding RNA Regulation in the Cardiovascular System, Abstract 10907: November 26, 2013, Vol 128, Issue suppl_22

Objective: The Polycomb-group (PcG) protein, Ezh2 methylates histone protein and is required for epigenetic gene silencing in the adult heart, but the mechanism of interaction at specific genes has not been explored. This study investigates the role of Ezh2 and ncRNA-mediated (non-coding RNA) PcG mechanisms in ventricular remodeling of the pathological heart.

Methods: We explored the expression as well as the interaction of ncRNAs in a mouse model to attenuate pressure overload caused by transverse aortic constriction (TAC) using the prototypical histone deacetylase (HDAC) inhibitor, Trichostatin A (TSA). In addition, loss of function studies was used to validate our findings. We examined the interaction of Ezh2 and ncRNA bound at gene promoters using chromatin immunoprecipitation (ChIP) assays.

Results: ChIP experiments in TAC hearts show release of Ezh2 at promoters of hypertrophy marker genes, Anp and Bnp, whereas increased binding of Ezh2 at the bidirectional promoter (bdP) of myosin heavy chain (MHC) genes. Administration of TSA attenuated Ezh2 binding to bdP and restored antisense (AS) β-MHC and α-MHC gene expression. Long ncRNAs (lncRNAs) are specifically regulated and RNA-ChIP experiments in TAC hearts show increased chromatin binding for the primary microRNA-208b (Pri-miR-208b) transcript. RNAi-mediated loss-of-function screens for lncRNAs (AS β-MHC, Pri-miR-208b) and Ezh2 identified Pri-miR-208b as a functional lncRNA that interact with Ezh2 at gene promoters of TAC hearts. We additionally showed the AS β-MHC transcript is critical in regulating cardiac MHC shift to pathological stress. All experiments independently performed (n≥6).

Conclusion: These results describe a previously uncharacterized interaction of a primary microRNA targeting specific chromatin modifications and regulation of genes implicated in pathological remodeling of ventricular hypertrophy. We propose lncRNAs function as transcriptional regulators of the hypertrophied heart and a better understanding of the complexity of epigenetic changes regulating gene expression in cardiac hypertrophy.