C9orf72 expansion disrupts ATM-mediated chromosomal break repair

Abstract

A hexanucleotide repeat expansion represents the most common genetic cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia, though the mechanisms by which the expansion cause neurodegeneration are poorly understood. We report elevated levels of DNA/RNA hybrids (R-loops) and double-strand breaks (DSBs) in rodent neurons, human cells, and in C9orf72-ALS patient spinal cord tissues. Accumulation of endogenous DNA damage is concomitant with defective ATM-mediated DNA repair signalling and accumulation of protein-linked DNA breaks. We further reveal that defective ATM-mediated DNA repair is a consequence of p62 accumulation, which impairs H2A ubiquitylation and perturbs ATM signalling. Adeno-associated virus-mediated expression of C9orf72-related RNA and dipeptide repeats in the murine central nervous system causes elevated DSBs, ATM defects, and triggers neurodegeneration. These findings identify R-Loops, DSBs, and defective ATM-mediated repair as pathological consequences of C9orf72 expansions, and suggest that C9orf72-linked neurodegeneration is driven, at least in part, by genomic instability.

C9orf72-FTD/ALS pathogenesis: evidence from human neuropathological studies

“…C9orf72 -ALS patient derived cell lines showed increased RNA G-quadruplex foci using an antibody against DNA/RNA G-quadruplex structures [20], however the existence of these expanded repeat-containing secondary structures in patient neuronal tissue remains unproven. A recent study showed increased R-loops (DNA-RNA hybrids) in spinal lower motor neurons from patients with C9orf72 -ALS compared to controls [157], but this study could not confirm that the R-loops contained elements of the repeat expansion. Higher order RNA structures could enhance sequestration of various RNA-binding proteins, leading to loss of function.…”

Reduced autophagy upon C9ORF72 loss synergizes with dipeptide repeat protein toxicity in G4C2 repeat expansion disorders

“…Second, sense G4C2 and antisense C4G2 expanded repeats are repeat‐associated non‐AUG (RAN) translated into dipeptide repeat (DPR)‐containing proteins, which form inclusions throughout the brain of patients with C9‐ALS/FTD (Ash et al , ; Gendron et al , ; Mori et al , ; Zu et al , ), as well as in mice expressing expanded G4C2 repeats (Chew et al , ; O'Rourke et al , ; Peters et al , ; Jiang et al , ; Liu et al , ). Overexpression of DPR proteins using artificial ATG codons for their translation initiation leads to neurodegeneration in cell and animal models, notably through alteration of mitochondria (Dafinca et al , ; Lopez‐Gonzalez et al , ; Choi et al , ), DNA repair (Walker et al , ; Lopez‐Gonzalez et al , ), nuclear and nucleolar organization (White et al , ; Zhang et al , ), and/or nucleocytoplasmic transport (Kwon et al , ; May et al , ; Mizielinska et al , ; Wen et al , ; Zhang et al , , ; Freibaum et al , ; Jovičić et al , ; Tao et al , ; Boeynaems et al , ; Khosravi et al , ). Third, expanded G4C2 repeats promote DNA epigenetic changes that lead to decreased expression of C9ORF72 mRNA and protein levels in C9‐ALS/FTD individuals (DeJesus‐Hernandez et al , ; Gijselinck et al , ; Almeida et al , ; Waite et al , ; van Blitterswijk et al , ; Xiao et al , ; Saberi et al , ; Frick et al , ; Viodé et al , ).…”

RNA Degradation in Neurodegenerative Disease

“…These structures may contribute to the pathology of expansion diseases in several ways: by blocking translation [238], disrupting chromatin remodeling [239], or promoting genomic instability at the repeat expansion site [235]. In support of the pathogenic effects of R-loops, mutations in the gene encoding senataxin (SETX), a helicase that helps resolve R-loops [240], cause juvenile ALS (ALS4), while SETX overexpression prevents neurodegeneration in ALS models [241]. …”