Friedreich ataxia (FRDA) is typically caused by homozygosity for an expanded GAA triplet-repeat in intron 1 of the FXN gene, which results in transcriptional deficiency via epigenetic silencing. Most patients are homozygous for alleles containing > 500 triplets, but a subset (~20%) have at least one expanded allele with < 500 triplets and a distinctly milder phenotype. The authors show that in FRDA DNA methylation spreads upstream from the expanded repeat, further than previously recognized, and establishes an FRDA-specific region of hypermethylation in intron 1 (~90% in FRDA versus < 10% in non-FRDA) as a novel epigenetic signature. Hypermethylation of this differentially methylated region (FRDA-DMR) was observed in a variety of patient-derived cells, it significantly correlated with FXN transcriptional deficiency and age of onset, and it reverted to the non-disease state in isogenically-corrected iPSC-neurons. Bisulfite deep sequencing of the FRDA-DMR in PBMCs from 73 FRDA patients revealed considerable intra-individual epiallelic variability, including fully methylated, partially methylated, and unmethylated epialleles. Whereas unmethylated epialleles were rare (median=0.33%) in typical patients homozygous for long GAA alleles with > 500 triplets, a significantly higher prevalence of unmethylated epialleles (median=9.8%) was observed in patients with at least one allele containing < 500 triplets, less severe FXN deficiency (>20%) and later onset (>15y). The higher prevalence in mild FRDA of somatic FXN epialleles devoid of DNA methylation is consistent with variegated epigenetic silencing mediated by expanded triplet-repeats. The proportion of unsilenced somatic FXN genes is an unrecognized phenotypic determinant in FRDA, and has implications for the deployment of effective therapies.

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