Figure 4

Methylation statuses of MOR gene proximal promoter (PP) in DRG. (A) The PP region of the MOR gene contains 20 putative methyl CpG sites from -569 to +33 (with the ATG start codon designated +1). The numbers at the top of the figure (No. CpG) are arbitrary designations to indicate each methyl CpG site. ppTIS indicates the TISs of the major MOR PP containing four sites. Each row of circles represents a single cloned allele, and each circle indicates a single CpG site at a specific location. The methylation statuses of 20 individual clones were analyzed for each mouse. The filled and open circles represent the methylated and unmethylated CpG sites, respectively. (B) Methylation changes within the PP region of MOR gene. The percentages of methylation at -388, -344 and -255 CpG sites significantly increased in DRG after nerve injury and reduced after 5-aza-dC treatment (5 μg, daily for 3 consecutive days, starting 30 minutes before surgery). **P<0.01compared with Sham-Vehicle mice; ^## p <0.01 compared with CCI-Vehicle mice, n=4 in each group. (C) Samples of the sequence fluorograms obtained using bisulfite genomic sequencing of DNA isolated from the DRG of Sham-Vehicle or CCI-5-aza-dC and CCI-Vehicle mice are shown. Arrows indicate methylated and nonmethylated sequences of CpG site -388, -344, and -255.