These results indicate that methylation attenuates NGFI-A induction of gene expression through the NR3C1 promoter. However, the decreased glucocorticoid receptor transcription observed in suicide victims with a history of childhood abuse was associated with differences in methylation levels occurring only at specific sites in the exon 1F NR3C1 promoter (Fig. 2b). An ANOVA examining the methylation of CpG dinucleotides across the exon 1F NR3C1 promoter revealed a significant effect of CpG site (F = 13.86, P < 0.0001), a significant effect of group (F = 17.12, P < 0.0001) and a significant interaction between CpG site and group (F = 13.44, P < 0.0001). In NGFI-A recognition elements, methylation was observed at CpG sites 12, 13, 30, 31 and 32 (Fig. 2b). We therefore examined whether such selective, site-specific differences in methylation could alter transcriptional activation through the NR3C1 promoter. Two deletion constructs of the NR3C1 promoter were generated in which selected CpG dinucleotides were patch-methylated (Fig. 3a). CpG sites 12 and 13 were methylated in the 255-bp construct, whereas the 125-bp promoter construct was methylated at CpG sites 30, 31 and 32. Thus, each deletion construct included at least one known or putative NGFI-A binding site28.
