As mentioned earlier, DNA methylation changes at the overall MECP2 promoter and individual CpG sites within the MECP2/Mecp2 promoter are associated with altered MECP2/Mecp2 expression [2,19,20]. Therefore, we investigated whether altered expression of Mecp2 isoforms in our NSC system are associated with change in DNA methylation at the Mecp2 REs found within the Mecp2 promoter and intron 1. Bisulfite pyrosequencing analysis showed that decitabine treatment (D2) caused no significant change in the percentage DNA methylation at the Mecp2 promoter R1 and R3 (Figure 6A, a, c). However, decitabine caused demethylation of all individual CpG dinucleotides at the R2 (CpG1, 3.5%; CpG2, 4.4%; CpG3, 3.1%; CpG4, 4.28%) (Figure 6A, b), as well as the average R2 percentage DNA methylation (3.83%, P <0.05) (Figure 6B, a). Similarly, decitabine caused demethylation of individual CpG sites at the intron 1 R4 (15.8%, P <0.05), R5 (13.08%, P <0.05), and R6 (CpG1, 8.01%, P <0.01; CpG2, 3.8%, P = 0.4) (Figure 6A, d-f), with significant demethylation at the entire intron 1 (10.37%, P <0.05) (Figure 6B, b). These results indicated that decitabine induced significant DNA
