Abstracts

Rationale: Several proteins involved in epigenetic regulation cause seizure-associated syndromic neurodevelopmental disorders when the human genes are mutated [e.g. MeCP2 in Rett syndrome (RTT)]. However, more general involvement of epigenetic mechanisms in these phenotypes is unclear. We investigated whether DNA methylation profiles of lymphoblastoid cell lines (LCLs) could be used to differentiate related neurodevelopmental disorders into etiological categories and identify specific genes that might be relevant to pathophysiology. Methods: DNA methylation profiling was performed on bisulfate converted DNA from LCLs in discovery (n=20) and replication cohorts (n=40) of females with Rett syndrome (RTT), autism (AUT), seizure disorder (SEZ), and controls (CTL) using Illumina HumanMethylation27 arrays. DNA methylation within the TAC1 promoter was confirmed using a Sequenom EpiTYPER assay, and gene expression by qRT-PCR was assessed in postmortem brain from RTT and AUT+SEZ females. Chromatin immunoprecipitation was performed in HEK cells using an anti-MeCP2 antibody, followed by qPCR at the TAC1 promoter. Results: In the discovery cohort, hierarchical clustering and principal component analysis using 40 CpG sites with significant differential methylation between etiological groups (P<0.01, FDR<0.10) distinguished RTT and AUT from CTL, but not from each other. These 40 CpG sites did not differentiate etiological groups in the replication cohort, but a pooled analysis identified four significant differentially methylated CpG sites (P<0.0005, FDR<0.005), including a CpG site in the TAC1 promoter. TAC1 hypermethylation in AUT and SEZ (P<0.005) was independently confirmed. In postmortem brain from RTT and AUT+SEZ females, TAC1 expression was significantly reduced in cingulate cortex (P=0.003), but not in striatum or temporal cortex (P>0.05). Chromatin immunoprecipitation in HEK cells confirmed MeCP2 binding at the TAC1 promoter, which was abolished following acute valproic acid treatment. Conclusions: These data newly identify TAC1 as a MeCP2 target gene and suggest underlying seizure disorder and/or antiepileptic treatment affects DNA methylation peripherally, which is associated with region-specific gene expression changes in the brain.