To profile EtOH-induced DNA methylation change on individual chromosomes, we calculated the average level of DNA methylation as shown in Figure 3. We also compared EtOH effects on DNA methylation in undifferentiated hESCs versus EBs. EtOH treatment of undifferentiated hESCs induced increases in the average levels of DNA methylation in the promoter regions and CpG islands of each chromosome (Fig. 3A and 3B, top row). The level of DNA methylation changes specific to EtOH treatment in differentiated hESC EBs was also examined (Fig. 3A and 3B, middle row). Interestingly, EtOH had a much smaller effect on DNA methylation in differentiated EBs. The extent of EtOH's effect on DNA methylation on each chromosome was evaluated by calculating a fold-change in DNA methylation level after EtOH treatment against untreated controls (Fig. 3, bottom row). Undifferentiated hESCs showed higher fold changes in DNA methylation upon EtOH treatment than differentiated hESC EBs. We also found higher fold changes in EtOH-induced DNA methylation in the promoter regions compared to CGIs. Chromosomes 2, 16 and 18 in undifferentiated hESCs showed the highest increases in DNA methylation in the promoter regions compared to other chromosomes.
