The C9orf72 repeat expansion itself is methylated in ALS and FTLD patients

• Published in: Acta neuropathologica 2015-05, Vol.129 (5), p.715-727
• Main Authors: Xi, Zhengrui, Zhang, Ming, Bruni, Amalia C., Maletta, Raffaele G., Colao, Rosanna, Fratta, Pietro, Polke, James M., Sweeney, Mary G., Mudanohwo, Ese, Nacmias, Benedetta, Sorbi, Sandro, Tartaglia, Maria Carmela, Rainero, Innocenzo, Rubino, Elisa, Pinessi, Lorenzo, Galimberti, Daniela, Surace, Ezequiel I., McGoldrick, Philip, McKeever, Paul, Moreno, Danielle, Sato, Christine, Liang, Yan, Keith, Julia, Zinman, Lorne, Robertson, Janice, Rogaeva, Ekaterina
• Format: Article
• Language: English
• Subjects: Aged; Alleles; Amyotrophic lateral sclerosis; Amyotrophic Lateral Sclerosis - genetics; C9orf72 Protein; CpG Islands; Disease; DNA Methylation; DNA Repeat Expansion; Epigenetics; Female; Frontotemporal Lobar Degeneration - genetics; Genetic testing; Heterozygote; Humans; Male; Medicine; Medicine & Public Health; Methylation; Middle Aged; Neurology; Neurosciences; Original Paper; Pathology; Proteins - genetics; Restriction Mapping - methods; Sequence Analysis, DNA
• ISSN: 0001-6322; 1432-0533
• DOI: 10.1007/s00401-015-1401-8

The most common cause of both amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD) is a G 4 C 2 -repeat expansion in C9orf72. However, the lower limit for pathological repeats has not been established and expansions with different sizes could have different pathological consequences. One of the implicated disease mechanisms is haploinsufficiency. Previously, we identified expansion-specific hypermethylation at the 5′ CpG-island near the G 4 C 2 -repeat, but only in a fraction of carriers (up to 36 %). Here, we tested the hypothesis that the G 4 C 2 -repeat itself could be the main site of methylation. To evaluate (G 4 C 2 ) n -methylation, we developed a novel assay, which was validated by an independent methylation-sensitive restriction enzyme assay. Notably, both assays are qualitative but not quantitative. Blood DNA was available for 270 unrelated individuals, including 71 expansion carriers. In addition, we investigated blood DNA from family members of 16 probands, and 38 DNA samples from multiple tissues of 10 expansion carriers. Finally, we tested DNA from different tissues of an ALS patient carrying a somatically unstable 90-repeat. We demonstrated that the G 4 C 2 -expansion is generally methylated in unrelated carriers of alleles >50 repeats (97 %), while small (