The ALS/FTD-related C9orf72 hexanucleotide repeat expansion forms RNA condensates through multimolecular G-quadruplexes

• Published in: Nature communications Vol. 14; no. 1; p. 8272
• Main Authors: Raguseo, Federica, Wang, Yiran, Li, Jessica, Petrić Howe, Marija, Balendra, Rubika, Huyghebaert, Anouk, Vadukul, Devkee M, Tanase, Diana A, Maher, Thomas E, Malouf, Layla, Rubio-Sánchez, Roger, Aprile, Francesco A, Elani, Yuval, Patani, Rickie, Di Michele, Lorenzo, Di Antonio, Marco
• Format: Journal Article
• Language: English
• Published: England Nature Publishing Group 13.12.2023; Nature Publishing Group UK; Nature Portfolio
• Subjects: Amyotrophic lateral sclerosis; Amyotrophic Lateral Sclerosis - genetics; C9orf72 Protein - genetics; Condensates; Dementia; Dementia disorders; DNA Repeat Expansion - genetics; Fluorescent indicators; Frontotemporal dementia; Frontotemporal Dementia - genetics; G-Quadruplexes; Gene sequencing; Humans; Motor neurons; Neurodegenerative diseases; Neurons; Nucleic acids; Proteins; Ribonucleic acid; RNA; RNA - chemistry; RNA - genetics; Therapeutic targets
• ISSN: 2041-1723; 2041-1723
• DOI: 10.1038/s41467-023-43872-1

Amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) are neurodegenerative diseases that exist on a clinico-pathogenetic spectrum, designated ALS/FTD. The most common genetic cause of ALS/FTD is expansion of the intronic hexanucleotide repeat (GGGGCC) in C9orf72. Here, we investigate the formation of nucleic acid secondary structures in these expansion repeats, and their role in generating condensates characteristic of ALS/FTD. We observe significant aggregation of the hexanucleotide sequence (GGGGCC) , which we associate to the formation of multimolecular G-quadruplexes (mG4s) by using a range of biophysical techniques. Exposing the condensates to G4-unfolding conditions leads to prompt disassembly, highlighting the key role of mG4-formation in the condensation process. We further validate the biological relevance of our findings by detecting an increased prevalence of G4-structures in C9orf72 mutant human motor neurons when compared to healthy motor neurons by staining with a G4-selective fluorescent probe, revealing signal in putative condensates. Our findings strongly suggest that RNA G-rich repetitive sequences can form protein-free condensates sustained by multimolecular G-quadruplexes, highlighting their potential relevance as therapeutic targets for C9orf72 mutation-related ALS/FTD.