Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease resulting in loss of motor neurons in the spinal cord, motor cortex and brainstem. As a result of neuronal loss muscle atrophy and signs of brain nerve weakness occur. The exact cause of ALS is yet to be determined, but scientific evidence has shown that genetics and environment have a role in degeneration of motor neurons and development of the disease. On the other side of the neuropathological spectrum is the disease frontotemporal dementia (FTD), which affects the frontal and temporal part of the brain and shows changes in behaviour and personality. Just like with the ALS, the cause of FTD is still unknown. In patients with ALS as wel as FTD, TDP–43 pathology was observed, where the RNA binding protein TDP–43 aggregates in the cytoplasm. Other RNA binding proteins are included in the development of ALS. The primary genetic cause for the diseases is the mutation in the C9orf72 gene, where a hexanucleotide repeat expansion GGGGCC appears in the first intron. Healthy individuals have around 30 repeats whereas individuals affected with the disease have ranging from ten to several thousand repeats. There are three proposed mechanisms of how the disease occurs. The first one is loss–of–function of the protein C9orf72, second is RNA toxicity, where the sense and antisense transcripts of the gene form RNA foci and last is DPR toxicity. Sense and antisense RNA can be translated through the process of RAN translation.
In this thesis, research was focused on structural determination of longer sense and antisense RNA repeat expansion, as a result of the mutation in C9orf72 gene. The thesis shows detailed research of the (G4C2)n and (C4G2)n secondary structures of different repeat lengths, which would give a better understanding how RNA toxicity mechanism works. Using in vitro transcription, we prepared RNA constructs for examination. We used NMR spectroscopy to show that r(G4C2)8 and r(G4C2)48 form G–quadruplexes in the presence of K+ ions.
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