CpG site; (iv) methylated in both sites; or (v) mutated at the two CpGs with an adenosine replacing the cytosines. NGFIA formed a protein-DNA complex with the nonmethylated oligonucleotide, while the protein is unable to form a complex with either a fully methylated sequence or a sequence methylated at the 5' CpG site. NGFIA binding to its response element was only slightly reduced with the sequence methylated at the 3' CpG site. The results indicate that while methylation of the cytosine within the 5' CpG dinucleotide reduces NGFIA protein binding to the same extent as methylation in both CpG sites, methylation of the cytosine within the 3' CpG dinucleotide only partially reduces NGFIA protein binding. These data support the hypothesis that methylation of the cytosine within the 5' CpG dinucleotide in the NGFIA response element of the exon 17 GR promoter region in the offspring of low-LG mothers inhibits NGFIA protein binding.
