|dc.description.abstract||Amyotrophic lateral sclerosis (ALS) is a deadly neurodegenerative disease with no cure, characterised by the progressive death of spinal motor neurons and subsequent paralysis. A hexanucleotide repeat expansion (HRE) in an intronic region of C9ORF72 is the most common known cause of familial ALS in European populations. By an unconventional form of ATG-independent translation, HRE RNA gives rise to a series of dipeptide repeats (DPRs) out of which two, poly(PR) and poly(GR), are toxic. These arginine-rich peptides were shown to accumulate at nucleoli, repress ribosome biogenesis, and drive multiple disturbances in almost all aspects of RNA and nucleolar metabolism. However, the precise mechanism of their toxicity, and whether it could be chemically or genetically alleviated remained unknown.
During the first part of this thesis we focused on resolving the mechanism to explain the pleiotropic pathological effects driven by arginine-rich DPRs. Our work revealed that this is due to a generalised DPR coating of DNA and RNA, which leads to protein displacement from nucleic acids, and thus to widespread inhibition of all reactions using nucleic acid substrates. As such, we propose that cell death is the result of inhibition of multiple processes, the most prominent of which being ribosome biogenesis and translation, as well as the collective displacement of RNA-binding proteins (RBPs) and ribosomal proteins (r-proteins) from RNA.
Next, and to evaluate the effects of arginine-rich DPRs in vivo, we developed a novel mouse model that enables tissue-wide inducible expression of ninety-seven copies of PR. Systemic expression of the peptide led to a lethal premature ageing phenotype within 3 months. Molecular and cellular biology analyses revealed that this progeroid phenotype was associated to an accumulation of nucleolar stress and free r-proteins, suggesting that the pathology is reminiscent to a class of diseases known as ribosomopathies. Supporting this view, DPR toxicity in vitro is ameliorated by the mTOR inhibitor rapamycin, or by downregulation of MYC, both of which are known to reduce overall translation and to extend lifespan in mammals. Importantly, and similarly to what has been seen in ribosomopathies, rapamycin extends the survival of (PR)97 mice.
Altogether, our work has revealed the mechanism of toxicity driven by arginine-rich DPRs, provided a new mouse model which can be used as a preclinical tool for research in the field, revealed that the pathology driven by poly(PR) DPRs is reminiscent of ribosomopathies, and identified some initial therapies that can reduce the severity of these diseases||en_US