Relatively little is known about the epigenetic control mechanisms that guide postnatal organ maturation. The goal of this study was to determine whether DNA methylation plays an important role in guiding transcriptional changes during the first 2 wk of mouse heart development, which is an important period for cardiomyocyte maturation, loss of proliferative capacity and loss of regenerative potential. Gene expression profiling (RNA‐seq) and genome‐wide sequencing of methylated DNA (MBD‐seq) identified dynamic changes in the cardiac methylome during postnatal development [2545 differentially methylated regions (DMRs) from P1 to P14 in the mouse]. The vast majority (~80%) of DMRs were hypermethylated between P1 and P14, and these hyper‐methylated regions were associated with transcriptional shut down of important developmental signaling pathways, including Hedgehog, bone morphogenetic protein, TGF‐β, fibroblast growth factor, and Wnt/β‐catenin signaling. Postnatal inhibition of DNA methylation with 5‐aza‐2'‐deoxycytidine induced a marked increase (~3‐fold) in cardiomyocyte proliferation and ~50% reduction in the percentage of binucleated cardiomyocytes compared with saline‐treated controls. This study provides novel evidence for widespread alterations in DNA methylation during postnatal heart maturation and suggests that cardiomyocyte cell cycle arrest during the neonatal period is subject to regulation by DNA methylation.—Sim, C. B., Ziemann, M., Kaspi, A., Harikrishnan, K. N., Ooi, J., Khurana, I., Chang, L., Hudson, J. E., El‐Osta, A., Porrello, E. R. Dynamic changes in the cardiac methylome during postnatal development. FASEB J. 29, 1329‐1343 (2015). www.fasebj.org