Methylation patterns of genes subject to and escaping X inactivation on active and inactive X chromosomes. (A) For genes subject to XCI, methylation differences between 46,XX and 45,X individuals occur specifically at gene promoters, with 46,XX individuals showing increased methylation compared to 45,X cases. As 83% of RefSeq transcripts listed on the X chromosome have a CGI within 1 kb of their TSS and CGIs are heavily methylated on the Xi versus the Xa, this enrichment of methylation at gene promoters on the Xi is a correlate of CGI methylation. (B) In contrast, for genes escaping XCI, there is a generalized increase in methylation in the bodies of genes in both 45,X and 46,XX individuals compared to genes subject to XCI, with the biggest difference between 46,XX and 45,X occurring at the promoter region. Additionally, a slight but consistent increase in methylation in the bodies of genes escaping XCI is evident in 46,XX compared to 45,X individuals. Two hundred sixteen RefSeq genes listed on chrX scored as subject to XCI (score = 0) and 52 RefSeq genes scored as escaping XCI (score = 8 or 9) by Carrel and Willard (2005) were analyzed. Where multiple splice forms were listed for a single gene, we chose the maximal boundary of all isoforms to define the transcribed region. Probes were then assigned into one of 40 bins depending on their relative position along the gene body, defined by the maximal transcription start and end coordinates. Similar data were observed when considering only those genes of intermediate length (10–100 kb) (Supplemental Figs. 11, 12).
