The hexanucleotide GGGGCC repeat expansion in the intronic region of C9orf72 is the most common cause of Amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). The repeat expansion-generated toxic RNAs and dipeptide repeats (DPRs) including poly-GR, have been extensively studied in neurodegeneration. Moreover, haploinsufficiency has been implicated as a disease mechanism but how C9orf72 deficiency contributes to neurodegeneration remains unclear. Here, we show that C9orf72 deficiency exacerbates poly-GR-induced neurodegeneration by attenuating non-homologous end joining (NHEJ) repair. We demonstrate that C9orf72 localizes to the nucleus and is rapidly recruited to sites of DNA damage. C9orf72 deficiency resulted in impaired NHEJ repair through attenuated DNA-PK complex assembly and DNA damage response (DDR) signaling. In mouse models, we found that C9orf72 deficiency exacerbated poly-GR-induced neuronal loss, glial activation, and neuromuscular deficits. Furthermore, DNA damage accumulated in C9orf72-deficient neurons that expressed poly-GR, resulting in excessive activation of PARP-1. PARP-1 inhibition rescued neuronal death in cultured neurons treated with poly-GR peptides. Together, our results support a pathological mechanism where C9orf72 haploinsufficiency synergizes with poly-GR-induced DNA double-strand breaks to exacerbate the accumulation of DNA damage and PARP-1 overactivation in C9orf72 ALS/FTD patients.
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The data generated to support the finding of this study are available from the corresponding author upon reasonable request.
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We thank XL, CF, and ZY from the Molecular Imaging Core Facility (MICF); PH, and CZ from the Multi-Omics Core Facility (MOCF); and YX, XR, and JL from the Molecular and Cell Biology Core Facility (MCBCF) at the School of Life Science and Technology, ShanghaiTech University for providing technical support. We also thank W. Wang (Interdisciplinary Research Center on Biology and Chemistry, China) for providing C9ALS iPSCs, V. Gorbunova (University of Rochester, USA) for providing NHEJ reporter constructs, and D. Spector (Cold Spring Harbor Laboratory, USA) for providing U2OS-LacO cells. This study was supported by grants from the National Natural Science Foundation of China (91949117, 31871044 to LL).
The authors declare no competing interests.
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He, L., Liang, J., Chen, C. et al. C9orf72 functions in the nucleus to regulate DNA damage repair. Cell Death Differ 30, 716–730 (2023). https://doi.org/10.1038/s41418-022-01074-0