DNA methylation is developmentally regulated for genes essential for cardiogenesis.
Bottom Line: Quantitative real-time PCR analysis of 350 genes with differential DNA methylation showed that the expression of 181 genes is developmentally regulated, and 79 genes have correlative changes between methylation and expression, including hyaluronan synthase 2 (Has2).Required for heart valve formation, Has2 expression in the developing heart valves is downregulated at E14.5, accompanied with increased DNA methylation in its enhancer.Genetic knockout further showed that the downregulation of Has2 expression is dependent on DNA methyltransferase 3b, which is co-expressed with Has2 in the forming heart valve region, indicating that the DNA methylation change may contribute to the Has2 enhancer's regulating function.
Affiliation: Division of Hematology, Department of Genetics, Albert Einstein College of Medicine of Yeshiva University, Bronx, NY (A.A.C., M.L., A.A.M., Y.W., M.S., B.W., J.M.G., D.Z.).Show MeSH
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Mentions: Pearson correlation was used to evaluate the overall similarity of MSFE/MPS tag counts between E11.5 and E14.5 samples. A linear regression was used to fit the relationship between tag counts and DNA methylation levels at 14 selected ACGT sites. Differentially methylated (DM) sites were identified by the program EdgeR,38 with <5% FDR. A two‐sided t test was used to evaluate the difference of tag counts at ACGT sites located to different genomic contents, while hypergeometric test was used to evaluate the enrichment of DM sites in promoters and gene bodies. The statistical analysis of differential gene expression was performed using Microsoft Excel and the data were presented as mean±standard error (SE). Student t test was used for comparison between groups and P values <0.05 were considered as significant, Bonferonni's correction was applied to account for multiple testing in gene expression analysis. For the expression analysis of 350 genes, Mann–Whitney test was also performed (data not shown). While the significance observed for the top 15 up‐ and down‐regulated genes in Figure 5B remained, a few other differentially expressed genes included in Figure 5A would lose statistical support.
Affiliation: Division of Hematology, Department of Genetics, Albert Einstein College of Medicine of Yeshiva University, Bronx, NY (A.A.C., M.L., A.A.M., Y.W., M.S., B.W., J.M.G., D.Z.).