Figure 4

Regional DNA methylation in FBXL5 promoter. A) Screenshot from the UCSC genome browser showing location of FBXL5 promoter exon1, intron1, CpG island, Illumina microarray probes and pyrosequencing assays and tracks for H3K4me1 and H3K4me3 in lymphoblastoid cell line (GM12878) and ES cell line (H1 h-ES) (Broad Institute Histone data). B&C) Boxplots of Illumina DNA methylation data for two microrray probes within FBXL5 gene. The Y axis shows DNA methylation levels presented as C/C + T and ranging from 0 to 1. The bottom and the top of the box are 25th and 75th percentiles respectively; the whiskers are within the 1.5 interquartile range (IQR) of the data, and the circles, are outlier data points above or below 1.5 IQR. C are controls (N = 19), K are cases with KDM5C mutations (N = 10), V are cases with p.R1546Q sequence variant. D) DNA methylation upstream of FBXL5 transcription start site as determined by pyrosequencing assays 1&2 (pyro1&2). The order of CpG sites are shown from the furthest upstream to the transcription start site. Each column is an average for each of three groups of 1)10 cases with KDM5C mutations (mutation), 2)19 controls (control), and 3) two individuals with the p.R1546Q variant of unknown significance (VUS). The arrow shows the CpG sites overlapping the microarray probe. P-values were determined by Kruskal-Wallis test between mutation cases and controls. **** is p <0.0001. E) DNA methylation upstream of FBXL5 transcription start site and downstream of pyrosequencing assays 1&2 as determined by pyrosequencing assay 3 (pyro 3). The order of CpG sites are shown from the furthest upstream to the transcription start site. Each column is an average for each of two groups of 1) 5 cases with KDM5C mutations (mutation), 2) 5 controls (control). Error bars are standard deviations.