Fig. 2

Promoter methylation of CES1 in response to FA supplementation. A Schematic of CES1 gene in UCSC browser with genomic coordinates in hg19 genome release. EPIC array probes are shown at the top; maximum gain and loss for β is also shown (+ 0.02β = 2%, − 0.08β = 8% methylation). Corresponding introns (blue lines with arrows marking the direction of transcription) and exons (blue rectangles) of the gene are marked. The position of the CGI within the region is highlighted (green). Pyro assay product used for DNA methylation analysis in the CB samples and cell lines (black) and RT-qPCR product (black rectangles presenting exons and lines with arrows-spliced introns marking orientation of the PCR reaction) used for assessing gene expression in cell lines are shown in track at the bottom. B CES1 promoter loses methylation after FA supplementation. Mean methylation percentage within highlighted CG sites in FA-treated and placebo CB samples by EPIC array. C Confirmation of mean DNA methylation of CES1 promoter within region shown in A comparing EPIC array and pyrosequencing. Placebo, FA-mean average of the individual means in the placebo and FA-treated groups; change of methylation–methylation difference between the groups; P-probability by Student’s t test. D Methylation levels at individual CG sites covered by the pyroassay in WT (HCT116, blue) and knockout (DKO, orange) cells. Values are shown as mean +/− SD for each site: *p < 0.05; **p < 0.01; ***p < 0.001. E RT-qPCR showing upregulation of gene in HCT116/DKO cells using the primers indicated in A, values normalized to HPRT. F Methylation levels using pyroassay as in D but in SHSY5Y (WT, green) compared to aza-treated SH-SY5Y cells (Aza, blue). Values are shown as mean +/− SD for each site: *p < 0.05; **p < 0.01; ***p < 0.001. G Upregulation of gene expression by RT-qPCR in aza-treated SH-SY5Y cells. All values are shown as mean +/− SD for each site or cell line: *p < 0.05; **p < 0.01; ***p < 0.001