Figure 3.

DR4 promoter methylation in SH-SY5Y cell lines cultured in different mediums. A. DNA methylation assay results of DR4 promoter. The sequences including the framed CpG sites were shown in Figure 2B. Solid circles indicate methylated CpG sites, and hollow circle nonmethylated. “Folate-” labels indicate cells cultured in medium supplemented with folate concentration 0 μg/ml, and “normal” labels folate concentration 4 μg/ml; B. Sequences of the 12^th and 13^th CpG sites of CpG island 2. They are both unmethylated in folate deficient cells and methylated in normal cells; C. Overall methylation % of CpG sites in CpG island 1, CpG island 2, and both islands in the promoter of DR4. *P < 0.05; **P < 0.01; N=3; D. CpG methylation pattern of each CpG site of DR4 promoter in CpG island 1. The methylation rate of the 6^th CpG site is lower in folate deficient cells than that in normal cells (P < 0.05); E. CpG methylation pattern of each CpG site of DR4 promoter in CpG island 2. The methylation rates of the 5^th, 8^th, 9^th,10^th, and 13^th CpG site are lower in folate deficient cells than those in normal cells (P < 0.05); F. Results of search transcription factor binding sites using TFSEARCH software in fragments including target CpG sites.