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ABSTRACTUsing Illumina 450K arrays, 1.85% of all analyzed CpG sites were significantly hypermethylated and 0.31% hypomethylated in fetal Down syndrome (DS) cortex throughout the genome. The methylation changes on chromosome 21 appeared to be balanced between hypo- and hyper-methylation, whereas, consistent with prior reports, all other chromosomes showed 3–11 times more hyper- than hypo-methylated sites. Reduced NRSF/RESTexpression due to upregulation of DYRK1A(on chromosome 21q22.13) and methylation of REST binding sites during early developmental stages may contribute to this genome-wide excess of hypermethylated sites. Upregulation of DNMT3L(on chromosome 21q22.4) could lead to de novomethylation in neuroprogenitors, which then persists in the fetal DS brain where DNMT3Aand DNMT3Bbecome downregulated. The vast majority of differentially methylated promoters and genes was hypermethylated in DS and located outside chromosome 21, including the protocadherin gamma (PCDHG) cluster on chromosome 5q31, which is crucial for neural circuit formation in the developing brain. Bisulfite pyrosequencing and targeted RNA sequencing showed that several genes of PCDHGsubfamilies A and B are hypermethylated and transcriptionally downregulated in fetal DS cortex. Decreased PCDHGexpression is expected to reduce dendrite arborization and growth in cortical neurons. Since constitutive hypermethylation of PCDHGand other genes affects multiple tissues, including blood, it may provide useful biomarkers for DS brain development and pharmacologic targets for therapeutic interventions.

Large genomic reference data sets reveal a spectrum of pathogenicity in the prion protein gene and provide genetic validation for a therapeutic strategy in prion disease.