A high proliferation rate has been observed to facilitate somatic cell reprogramming, but the pathways that connect proliferation and reprogramming have not been reported. DNA methyltransferase 1 (DNMT1) methylates hemimethylated CpG sites produced during S phase and maintains stable inheritance of DNA methylation. Impairing this process results in passive DNA demethylation. In this study, we show that the cell proliferation rate positively correlated with the expression of Dnmt1 in G1 phase. In addition, as determined by whole-genome bisulfate sequencing and high-performance liquid chromatography, global DNA methylation of mouse embryonic fibroblasts was significantly higher in G1 phase than in G2/M phase. Thus, we suspected that high cellular proliferation requires more Dnmt1 expression in G1 phase to prevent passive DNA demethylation. The methylation differences of individual CpG sites between G1 and G2/M phase were related to the methylation status and the positions of their surrounding CpG sites. In addition, larger methylation differences were observed on the promoters of pluripotency-related genes; for example, Oct4, Nanog, Sox2, Esrrb, Cdh1, and Epcam. When such methylation differences or passive DNA demethylation accumulated with Dnmt1 suppression and proliferation acceleration, DNA methylation on pluripotency-related genes was decreased, and their expression was up-regulated, which subsequently promoted pluripotency and mesenchymal–epithelial transition, a necessary step for reprogramming. We infer that high cellular proliferation rates promote generation of induced pluripotent stem cells at least partially by inducing passive DNA demethylation and up-regulating pluripotency-related genes. Therefore, these results uncover a connection between cell reprogramming and DNA methylation.
cell proliferation
DNA methylation
DNA methyltransferase
p53
reprogramming
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This work was supported by National Natural Science Foundation of China Grants 91519305, 31422032, 31671475, U1601228, and 31421004; Special Research Project for Frontier Science of the Chinese Academy of Sciences Grant QYZDB-SSW-SMC031; Guangdong Natural Science Foundation Grant 2014A030308002; Guangzhou Science and Technology Program Grant 201607010239; Guangzhou Health Care Collaborative Innovation Program Grant 201508020250; and the Guangdong Special Support Program Grant 2014TQ01R157. The authors declare that they have no conflicts of interest with the contents of this article.
The WGBS, RRBS, and RNA-seq data were deposited in the Gene Expression Omnibus under accession numbersGSE92903, GSE93058, andGSE93416, respectively. These high-throughput sequencing results are available under SuperSeries accession numberGSE93417.
