Genome-wide DNA methylation analysis reveals dynamic changes in the cardiac methylome during post-natal heart development (RNA-Seq)

Epigenetic modifications have emerged as central players in the coordination of gene expression networks during cardiac development. While several studies have investigated the role of histone modifications during heart development, relatively little is known about the role of DNA methylation. The purpose of the current study was to determine whether DNA methylation plays an important role in guiding transcriptional changes during the neonatal period, which is an important developmental window for cardiac maturation and cardiomyocyte cell cycle arrest. We used methyl binding domain protein sequencing (MBD-seq) and mRNA-seq to profile DNA methyation and gene expression respectively in neonatal hearts at P1 and P14 stages. Thousands of differentially methylated regions (DMRs) were identified between P1 and P14, the vast majority of which were hypermethylated. Gene ontology analysis revealed that these hypermethylated genes were associated with transcriptional regulation of important developmental signaling pathways, including Hedgehog, BMP, TGF beta, FGF and Wnt/b-catenin signaling. A significant enrichment for myogenic transcription factors and Smad2/3/4 binding sites was also noted among differentially methylated peaks at P14. This study provides novel evidence for widespread alterations in DNA methylation during post-natal heart maturation and suggests that DNA methylation plays an important role in cardiomyocyte cell cycle arrest during the neonatal period. mRNA-seq to profile gene expression in neonatal hearts at P1 and P14 stages (post-natal day 1 and 14 respectively) in three biological replicates.表观遗传修饰（epigenetic modifications）已成为心脏发育过程中基因表达网络调控的核心参与者。尽管已有多项研究探讨了心脏发育过程中组蛋白修饰的作用，但目前对DNA甲基化的功能仍知之甚少。本研究旨在探究DNA甲基化是否在新生期这一心脏成熟与心肌细胞周期阻滞的关键发育窗口中，对转录组变化起到重要调控作用。我们分别采用甲基结合域蛋白测序（methyl binding domain protein sequencing, MBD-seq）与mRNA测序（mRNA-seq），对出生后第1天（P1）和第14天（P14）的新生小鼠心脏中的DNA甲基化水平与基因表达谱进行检测。研究在P1与P14心脏间鉴定出数千个差异甲基化区域（differentially methylated regions, DMRs），其中绝大多数呈现高甲基化状态。基因本体（gene ontology, GO）富集分析显示，这些高甲基化基因与多个重要发育信号通路的转录调控密切相关，包括刺猬信号通路（Hedgehog）、骨形态发生蛋白信号通路（BMP）、转化生长因子β信号通路（TGF beta）、成纤维细胞生长因子信号通路（FGF）以及Wnt/β-连环蛋白信号通路（Wnt/β-catenin）。此外，在P14阶段的差异甲基化峰中，还显著富集了肌源性转录因子与Smad2/3/4结合位点。本研究为出生后心脏成熟过程中DNA甲基化的广泛改变提供了全新证据，并表明DNA甲基化在新生期心肌细胞周期阻滞过程中发挥着关键调控作用。本数据集包含针对出生后第1天和第14天新生小鼠心脏的基因表达谱解析的mRNA测序数据，共设置3次生物学重复。