RATIONALE: Epigenetic marks are crucial for organogenesis, but their role in 
heart development is poorly understood. Polycomb repressive complex 2 (PRC2) 
trimethylates histone H3 at lysine 27, which establishes H3K27me3 repressive 
epigenetic marks that promote tissue-specific differentiation by silencing 
ectopic gene programs.
OBJECTIVE: We studied the function of PRC2 in murine heart development using a 
tissue-restricted conditional inactivation strategy.
METHODS AND RESULTS: Inactivation of the PRC2 subunit Ezh2 by Nkx2-5(Cre) 
(Ezh2(NK)) caused lethal congenital heart malformations, namely, compact 
myocardial hypoplasia, hypertrabeculation, and ventricular septal defect. 
Candidate and genome-wide RNA expression profiling and chromatin 
immunoprecipitation analyses of Ezh2(NK) heart identified genes directly 
repressed by EZH2. Among these were the potent cell cycle inhibitors Ink4a/b 
(inhibitors of cyclin-dependent kinase 4 A and B), the upregulation of which was 
associated with decreased cardiomyocyte proliferation in Ezh2(NK). 
EZH2-repressed genes were enriched for transcriptional regulators of 
noncardiomyocyte expression programs such as Pax6, Isl1, and Six1. EZH2 was also 
required for proper spatiotemporal regulation of cardiac gene expression, 
because Hcn4, Mlc2a, and Bmp10 were inappropriately upregulated in ventricular 
RNA. PRC2 was also required later in heart development, as indicated by 
cardiomyocyte-restricted TNT-Cre inactivation of the PRC2 subunit Eed. However, 
Ezh2 inactivation by TNT-Cre did not cause an overt phenotype, likely because of 
functional redundancy with Ezh1. Thus, early Ezh2 inactivation by Nk2-5(Cre) 
caused later disruption of cardiomyocyte gene expression and heart development.
CONCLUSIONS: Our study reveals a previously undescribed role of EZH2 in 
regulating heart formation and shows that perturbation of the epigenetic 
landscape early in cardiogenesis has sustained disruptive effects at later 
developmental stages.