Site-selective differences in methylation of CpG regions in promoters can alter gene transcription by affecting transcription factor binding24. We carried out chromatin immunoprecipitation assays on samples that were obtained using the transfection model described above to examine the association of NGFI-A with the methylated and the nonmethylated NR3C1 promoter constructs. The results revealed a significant effect of methylation status on NGFI-A association with the exon 1F NR3C1 promoter constructs transfected with EGR1 (F = 242.92, P < 0.0001; Fig. 4a). Post hoc tests showed that, in comparison with the nonmethylated construct, patch-methylation inhibited NGFI-A binding to the exon 1F NR3C1 promoter (P < 0.005; Fig. 4a). For the 125-bp promoter construct, there was a significant effect of methylation status (F = 102.28, P < 0.001) and NGFI-A treatment (F = 209.99, P < 0.0005) and a significant interaction between methylation status and the presence of NGFI-A (F = 72.71, P < 0.005). Similarly, for the 255-bp promoter construct, there was a significant effect of methylation status (F = 95.18, P < 0.001) and NGFI-A treatment (F = 152.13, P <
